CA 02421319 2003-03-25
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CA 02421319 2003-03-25
WO 99146281 PCT/US99/OSOZ8
NOVEL POLYPEPTIDES AND NUCLEIC ACIDS ENCODING THE SAME
F~E~.D OF THE INVENTION
_, The present invention relates generally to the identification and isolation
of novel DNA and to the
recombinant production of novel polypeptides encoded by that DNA.
BACKGROUND OF THF~,NV~jVTION
Extracellular proteins play an important role in the formation,
differentiation and maintenance of
multicelluiar organisms. The fate of many individual cells, e.g.,
proliferation, migration, differentiation, or
interaction with other cells, is typically governed by information received
from other cells andlor the immediate
environment. This information is often transmitted by secreted polypeptides
(for instance, mitogenic factors, survival
factors, cytotoxic factors, differentiation factors, neuropeptides, and
hormones) which are, in turn, received and
interpreted by diverse cell receptors or membrane-bound proteins. These;
secreted polypeptides or signaling
molecules normally pass through the cellular secretory pathway to reach their
site of action in the exuacellular
envirottment.
Secreted proteins have various industrial applications, including
phatxttaceuticals, diagnostics, biosensors
and bioreactors. Most protein drugs available at present, such as thromboiytic
agents, interferons, interletikins,
erythropoietins, colony stimulating factors, and various other cytokines; are
secretory proteins. Their receptors,
which are membrane proteins, also have potential as therapeutic or diagnostic
agents. Efforts are being undertaken
by both ittdvstry and academia to identify new, native secreted proteins. Many
efforts are focused on the screening
of mammalian recombinant DNA libraries to identify the coding sequences for
novel secreted proteins. Examples
of screening methods and techniques are described in the literature jsee, for
example, Klein et al., Proc. Natl. Acacj,,
~1., g~:?i08-7113 (1996); U.S. Patent No. 5,536,637)].
Membrane-bound proteins and receptors can play an important role in the
formation, differentiation and
maintenance of multicellttlar organisms. The fate of many individual cells,
e.g., proliferation, migration,
di0'ertumation, or interaction with other cells, is typically governed by
information received from other cells andlor
the immediate environment. This information is often transmitted by secreted
polypeptides (for instance, mitogenic
factors, survival factors, cytotoxic factors, differentiation factors,
neuropeptides, and hormones) which are, in turn,
received and imerpreted by diverse cell receptors or membrane-bound proteins.
Such membrane-bound proteins and
~ll receptors include, but are not limited to, cytokine receptors, receptor
kinases, receptor phosphatases, receptors
involved in cell-cell interactions, and cellular adhesin molecules like
selectins and integrins. For instance,
ttansduction of signals that regulate cell growth and differentiation is
regulated in part by phosphorylation of various
_ cellular proteins. Protein tyrosine kinases, enzymes that catalyze that
process, can also act as growth factor
receptors. Examples include fibroblast growth factor receptor and nerve growth
factor receptor.
Membrane-bound proteins and receptor molecules have various industrial
applications, including as
pharmaceutical atxi diagnostic agents. Receptor immuttoadhesins, for instance,
can be employed as therapeutic agents
1
CA 02421319 2003-03-25
WO 99/x6281 PCT/US99105028
to block receptor-ligand interaction. The membrane-bound proteins can also be
employed for screening of potential
peptide or small molecule intu'bitors of the relevant receptorlligand
interaction. Efforts are being undertaken by both
industry and academia to identify new, native receptor proteins. Many efforts
are focused on the screening of
mammalian recombinant DNA libraries to identify the coding sequences for novel
receptor proteins.
We herein describe the identification and characterization of novel secreted
and transmembrane polypeptides
and novel nucleic acids encoding those polypeptides.
1. R 2
Human growth arrest-specific gene 6 (gash) encodes a protein that is expressed
in a variety of different
tissues and which has been reported to be highly expressed during periods of
serum starvation and negatively
regulated during growth induction. See Manfioletti et al., Mot. Celt. Biol.
13(8):4976-4985 (1993) and Stitt et al.,
Cell 80:661-670 (1995). ManfioIetti et al. (1993), supra, have suggested that
the gash protein is member of the
vitamin K-dependent family of proteins, wherein the members of the latter
family of proteins (which include, for
example, Protein S, Protein C and Factor X) all play regulatory roles in the
blood coagulation pathway, Thus, it has
been suggested that gash may play a role in the regulation of a protease
cascade relevant in growth regulation or in
the blood coagulation cascade.
Given the physiological importance of the gash protein, efforts are currently
being undertaken by both
industry and academia to identify new, native proteins which are homologous to
gash. Many of these efforts are
focused on the screening of mammalian recombinant DNA libraries to identify
the coding sequences for novel
secreted and membrane-bound receptor proteins, specifncally those having
homology to gash. Examples of such
screening methods and techniques are described in the literature [see, for
example, Klein et al., roc. Nail. Acad.
~,,, Q3:7108 7113 (1996); U.S. Patern No. 5,536,637)]. We herein describe the
identification of a novel polypeptide
which has homology to the gash polypeptide.
2. PR0274
The 7-transmembrane ("7TM") proteins or receptors, also referred to in the
literature as G-protein coupled
receptors, are specialized proteins designed for recognition of ligands and
the subsequent signal transduction of
information contained within those ligands to the machinery of the cell. The
primary purpose of cell surface receptors
is to discriminate appropniate.ligands from the various extracellular stimuli
which each cell encounters, then to
activate an effector system that produces an intracellular signal, thereby
controlling cellular processes. (Dohhnan,
H.,Arnn. ev. Biochem., øt'1:653 (1991)]. The ability of 7TM receptors to bind
ligand to a recognition domain and
allostetically transmit the information to an intracellular domain is a
specialized feature of 7TM proteins [Kenakin,
T., Pharmacol. Rev., 4$:413 (1996)]. The gene family which encodes the 7TM
receptors or G-protein linked
n~eceptors encode receptors which recognize a large number of ligands,
including but rat limited to, CSa, interleukin
8 and related chemokines. Research in this area suggests that distinct signals
at the cell surface feed into common
pathways of cell activation. [Genard, C. and Gerard, N..CUTr. Op. Immunol.,
6_:140 (1994), Gerard, C. and Gerard,
N.,Atm. Rev. hnttrunol., x:775 (1994)]. The superfamily of 7TM or G-protein
coupled receptors contains several
hundred members able to recognize various messages such as photons, ions and
amino acids among others [Schwartz,
T.W., et al., H.,TrenrtC In Phar~col. Sci., 7~]:213 (1996)].
(DohhTtan; H."inn. Rev. B'E~qchem., x:653 (1991)]. (Schwartz. T.W., ei al"
H.,~ur J Pharm Sci , 2:85 (1994)].
2
CA 02421319 2003-03-25
WO 99146281 PCTlUS99105028
We describe herein the identification of a novel polypeptide (designated
herein as PR0274) which has homology to
the 7 uansmembrane segment receptor proteins and the Fn54 protein.
3. PR030_0
The Dill' 33 protein is over-expressed in mouse testicular tumors. At present
its role is unclear, however;
it may play a role in cancer. Given the medical importance of understanding
the physiology of cancer, efforts are
currently being under fatten to identify new, native proteins which are
involved in cancer. We describe herein the
identification of a novel polypeptide which has homology to Diff 33,
designated herein as PR0300.
4. PR0284
Efforts acre currently being undertaken to identify and characterize novel
uartsmembrane proteins. We
herein describe the identification and characterization of a novel
transmembrane polypeptide, designated herein as
PR0284.
5. PR0296
Cancerous cells often express numerous proteins that are not expressed in the
corresponding normal cell
type or are expressed at different levels than in the corresponding normal
cell type. Many of these proteins are
involved in inducing the transformation from a nam~al cell to a cancerous cell
or in maintaining the cancer phenotype.
As such, there is significant interest in identifying and characterizing
proteins that are expressed in cancerous cells.
We herein describe the identification and characterization of a novel
polypeptide having homology to the sarcoma-
amplified protein SAS, designated herein as PR0296.
6. PR0329
Irnmunoglobulin molecules play roles in many important matntnalian
physiological processes. The structure
of immunoglobulin molecules has been extensively studied and it has been well
documented that intact
immunoglobulins possess distinct domains, one of which is the constant domain
or F~ region of the immunoglobulin
molecule. The F~ domain of an immunoglobulin, while not being directly
involved in antigen recognition and binding,
does mediate the ability of the immunoglobulin molecule, either uncomplexed or
complexed with its respective
antigen, to bind to F~ receptors either circulating in the serum or on the
surface of cells. The ability of an F~ domain
of an immunoglobulin to bind to an F~ receptor molecule results in a variety
of important activities, including for
example, in maiu>ting an immune response against unwanted foreign particles.
As such, there is substantial interest
in identifying novel F~ receptor proteins and subunits thereof. We herein
describe the identification and
characterization of a novel polypeptide having homology to a high affinity
immunoglobulin F~ receptor protein,
designated herein as PR0329.
7. PR036~
._ Colorectal carcinoma is a malignant neoplastic disease which has a high
incidence in the Western world,
particularly in the United States. Tumors of this type often metastasize
through lymphatic and vascular channels and
result in the death of some 62,000 persons in the Un9ted States annually.
3
CA 02421319 2003-03-25
wr
WO 99/4628I PCTIUS99I05028
Monoclonal antibody A33 (mAbA33) is a murine immunoglobulin that has undergone
extensive preclinical
analysis and localization studies in patients inflicted with colorectal
carcinoma (Welt et al., J. Clin. Oncol. 8:1894-
1906 (1990} and Welt et al., J. Clin. Oncol. 12:1561-1571 (1994)). mAbA33 has
been shown to bind to an antigen
found in and on the surface of normal colon cells and colon cancer cells. In
carcinomas originating from the colonic
tnucosa, the A33 antigen is expressed homogeneously in more than 95 % of the
cases. The A33 antigen, however,
S has not been detecting in a wide range of other nomtaI tissues, i.e., its
expression appears to be rather organ specific
Therefore, the A33 antigen appears to play an important role in the induction
of colorectal cancer.
Given the obvious importance of the A33 antigen in tumor cell formation andlor
proliferation, there is
Substantial interest in identifying homologs of the A33 antigen, In this
regard, we herein describe the identification
and characterization of a novel polypeptide having homology to the A33 antigen
protein, designated herein as
PR0362.
8. 03
The cell surface protein HCAR is a membrane-bound protein that acts as a
receptor for subgroup C of the
adenoviruses and subgroup B of the coxsackieviruses. Thus, HCAR may provide a
means for mediating viral
infection of cells in that the presence of the HCAR receptor on the cellular
surface provides a binding site for viral
particles, thereby facilitating viral infection.
In light of the physiological importance of membrane-bound proteins and
spcfcially those which serve a cell
surface receptor for viruses, efforts are currently being undertaken by both
industry and academia to identify new,
native membrane-bound reeptor proteins. Many of these efforts are focused on
the screening of mammalian
recombinant DNA libraries to identify the coding sequences for novel receptor
proteins. We herein describe a novel
membrane-bound polypeptide having homology to the cell surface protein HCAR
and to various tumor antigens
inchiding A33 and carcinoembryonic antigen, designated herein as PR0363,
wherein this polypeptide may be a novel
cell surface virus receptor or tumor antigen.
9. Rg 0868
Control of cell numbers in ma>rmoals is believed to be determined, in part, by
a balance between cell
proliferation and cell death. One form of cell death, sometimes referred to as
necrotic cell death, is typically
charaaerized as a pathologic form of cell death resulting from some uauma or
cellular injury. 1n coturast, there is
another, "physiologic" form of cell death which usually proceeds in an orderly
or controlled manner. This orderly
~ comrolled form of cell death is often referred to as "apoptosis" [see, e.g.,
liarr et al., ~olTec~ology, 12:487-493
(1994); Steller et al., S l ce, x:1445-1449 (1995)]. Apoptotic cell death
naturally occurs in many physiological
~, including embryonic development and clonal selection in the immune system
[Itch et al., Cell, x:233 243
(1991)]. Decreased levels of apoptotic cell death have been associated with a
variety of pathological conditions,
including cancer, lupus, and herpes virus infection ['fhompson, ci e, x:1456-
1462 (1995)]. Increased levels
of apoptotic cell death may be associated with a variety of other pathological
conditions, including AIDS, Alzheimer's
disease, Parkinson's disease, amyotrophic lateral sclerosis, multiple
sclerosis, retinitis pigmentosa, cerebellar
degeneration, aplastic anemia, myocardial infarction, stroke, reperfusion
injury, and toxin-induced liver disease [see,
Thompson, suvral.
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CA 02421319 2003-03-25
WO 99/46281 PCT/US99/05028
Apoptotic cell death is typically accompanied by one or more characteristic
morphological and biochemical
changes in cells; such as condensation of cytoplasm, loss of plasma membrane
microvilli, segmentation of the
nucleus, degradation of chromosomal DNA or loss of mitochondria) function. A
varien~ of extrinsic and intrinsic
signals are believed to trigger or induce such morphological and biochemical
cellular changes [Raft, ature, 3,y56:397-
400 (1992); Steller, ~unra; Sachs et al., B ood, 82:15 (1993}]. For instance,
they can be triggered by hormonal
stimuli, such as giucocorticoid hormones for immature thymocyCes, as well as
withdrawal of certain growth factors
(Watanabe-Fukunaga et al., Na r , ~5 :314-317 (1992)]. Also, some identified
oncogenes such as myc, re), and
EIA, and tumor suppressors, like p53, have been reported to have a role in
inducing apoptosis. Certain
chemotherapy. drugs and some forms of radiation have likewise been observed to
have apoptosis-inducing activity
[Thompson, su ra .
Various molecules, such as tumor necrosis factor-a ("TNF-a"), tumor necrosis
factor-~i ("TNF-Vii" or
"lymphotoxin-a"), lymphotoxin-p ("LT (3"), CD30 ligand, CD27 ligand, CD40
ligand, OX-40 ligand, 4-IBB ligand,
Apo-l ligand (also referred to as Fas Iigand or CD95 ligand), and Apo-2 ligand
(also referred to as TRAIL) have been
identified as members of the tumor necrosis factor ("TNF") family of cytokines
[See, e.g., Gruss and Dower, ood,
$5:3378-3404 (199; Pith et al., J. Biol. Chem., 271:12687-12690 (1996); Wiley
et al., Immunity, x:673-682 (1995);
Browning et al., Cue], x:847-856 (1993); Armitage et al. Nature, 3~7:80-82
(1992), WO 97101633 published January
16, 1997; WO 97)25428 published July 17, 1997]. Among these molecules, TNF-a,
TNF-(3, CD30 ligand, 4-1BB
ligand, Apo-1 ligand, and Apo-2 ligand (TRAIL) have been reported to be
involved in apoptotic cell death. Both
TNF-a and TNF-(3 have been reported to induce apoptotic death in susceptible
tumor cells [Schmid et al., Proc. Nat).
cad, c' , 83:1881 (1986); Dealtry et al., Eur. J. Immunol., X7:689 (1987)].
Zheng et al. have reported that TNF-a
is involved in post stimulation apoptosis of CD8-positive T cells [Zheng et
al., Na e, 377:348-351 (1995)]. Other
investigators have reported that CD30 ligand may be involved in deletion of
self reactive T cells in the thymus
(Amahawa et al., Cold Spring Harbor Laboratory Symposium on Programmed Cell
Death, Abstr. No. 10, (1995)].
Mutations in the mouse Fas/Apo-1 receptor or ligand genes (called ,)pr and
gld, respectively) have been
associated with some autoimmune disorders, indicating that Apo-1 ligand may
play a role in regulating the clonal
deletion of self reactive lymphocytes in the periphery jKratnmer et al., ~.
Op. hntr~unol., ø:279-289 (1994);
Nagata et al., Science, 07:1449-1456 (1995)). Apo-1 ligand is also reported to
induce post-stimulation apoptosis
in CD4-positive T lymphocytes and in B lymphocytes, and may be involved in the
elimination of activated
lymphocytes when their function is no longer needed [Krammer et al., supra;
Nagata et al., su . Agonist mouse
monoclonal antibodies specifically binding to the Apo-1 receptor have been
reported to exhibit cell killing activity
that is comparable to or similar to that ofTNF-a [Yonehara et a).,1 Exp. Med.,
169:1747-1756 (1989)]:
Induction of various cellular responses mediated by such TNF family cytokines
is believed to be initiated
by their binding to specific cell receptors. Two distinct TNF receptors of
approximately 55-kDa (TNFRl) and 75-
kDa (TNFR2) have been identified [Hohman et al:, j. Biol: Chem., x:14927-14934
(1989); Brockhaus et al., Proc.
Nat). Acad. Sci., $7:3127-3131 (/990); EP 417,563, published March 20, 1991]
and human and mouse cDNAs
corresponding to both receptor types have been isolated and characterized
[Loetscher et al., ~, ,øx:351 (1990);
Schall et al., ~I , ø1_:361 (1990); Smith et al., Science, ~$:1019-1023
(1990); Lewis et al., Proc. Natl. Acad. Sci.,
X8:2830-2834 (1991); Goodwin et al., Mol. Ce,[l. Biol., j;~:3020-3026 (1991)].
Extensive polymorphisms have been
associated with both TNF receptor genes (see, e.g., Takao et at.,
ImmunoQenetics, X7:199-203 (1993)]. Both TNFRs
share the typical strucwre of cell surface receptors including extracellular;
transmembrane and intracellular regions.
S
CA 02421319 2003-03-25
WO 99/46281 PCT/US99105028 .
The extracellular portions of both receptors are found naturally also as
soluble TNF-binding proteins [Nophar, Y.
et ai., EM~ 80 J., _9:3269 (1990): and Kohno, T. et ai., Proc. Natl. Aca~l.
Sci. U.S.A., x'1:8331 (1990)]. More
recently, the cloning of recombinant soluble TNF receptors was reported by
Hale et al. LJ. Cell Biochem.
Suppjement 15F, 1991, p. 113 (P424)].
The extracellular portion of type 1 and type 2 TNFRs (TNFRI and TNFR2)
contains a repetitive amino acid
sequence pattern of four cysteine-rich domaizLS (CRDs) designated 1 through 4,
starting from the NH,-terminus. Each
CRD is about 40 amino acids Iong and contains 4 to 6 cysteine residues at
positions which are well conserved [Schall
et al., supra; L~oetscher et al., ~nra; Smith et al., suvra; Nophar et al.,
supra; Kohno et al., ~_upral. In TNFRl, the
approximate boundaries of the four CRDs are as follows: CRD1- amino acids 14
to about 53; CRD2- amino acids
from about 54 to about 97; CRD3- amino acids from about 98 to about 138; CRD4-
amino acids from about 139 to
about 167. In TNFR2, CRD1 includes amino acids 17 to about 54; CRD2- amino
acids from about 55 to about 97;
CRD3- amino acids from about 98 to about 140; and CRD4- amino acids from about
141 to about 179 [Banner et al.,
.dell, x_3:431-435 (1993)]. The potential rose of the CRDs in ligand binding
is also described by Banner et al., suvra.
A similar repetitive pattern of CRDs exists in several other cell-surface
proteins, including the p75 nerve
growth factor receptor (NGFR) (Johnson et al., Celj, 47:545 (1986); Radeke et
al., N re, 325:593 (1987)], the B
cell antigen CD40 [Stamenkovic et al., ME BO J., _8:1403 (1989)), the T cell
antigen OX40 [Mallet et al., EMBO J.,
Q:1063 (1990)] and the Fas antigen [Yonehara et ai., supra and Itoh et al.,
ell, 66:233-243 (1991)]. CRDs are also
found in the soluble TNFR (sTNFR)-like T2 proteins of the Shope and myxoma
poxviruses [LJpton et al., Virolossv,
L~0-29 (1987); Smith et al., Biochem. Biophxs. Res. Commun., 176:335 (1991);
Upton et al., ViroloQV, 184:370
(1991)]. Optimal alignment of these sequences indicates that the positions of
the cysteine residues are well conserved.
These receptors are sometirues collectively referred Lo as members of the
TNFINGF receptor superfamily. Recent
studies on p75NGFR showed that the deletion of CRDI (lVeicher, A.A. et aL,
Proc. Natl. Acad. Sci. USA, $$:159-
163 (1991)] pr a 5-amino acid insertion in this domain [Yap, H. and Chao,
M.V., J. Biol. Chem., X66:12099-12104
(1991)] had little or no effect on NGF binding [Yap, H. and Chao, M.V., a ra ,
p75 NGFR contains a proiine-rich
stretch of about 60 amino acids, between its CRD4 and transmembrane region,
which is not involved in NGF binding
[Peetre, C. et al., Eur. J. Het~a~ol., 4_x,:414-419 (1988); Seckinger, P. et
al., J. Biol. Chem., x:11966-11973
(1989); Yan, H. and Chao, M.V., su . A similar proline-rich region is found in
TNFR2 but not in TNFRl.
Tl~ TNF family ligands identified to date, with the exception of lymphotoxin-
a, are type D transmembrane
proteins, whose C-terminus is extracellular. In contrast, most receptors in
the TNF receptor (TNFR) family identified
to date are type I transmembrane proteins. In both the TNF ligand and receptor
families, however, homology
identified between family members has been found mainly in the extracellular
domain ("F:CD"). Several of the TNF
family cytokirtes, including TNF-a, Apo-1 ligand and CD40 Iigand, are cieaved
proteolytically at the cell surface;
the resorting protein in each case typically forms a homotrimeric molecule
that functions as a soluble cytokine. TNF
receptor family proteins are also usually cleaved pmteolytically to reiease
soiubie receptor ECDs that can function
as inhibitors of the cognate cytokines.
Reoezuiy, other members of the TNFR family have been identified. Such newly
identified members of the
TNFR family include CARL, HVEM and osteoprotegerin (OPG) [Brojatsch et al.,
~1, 82:845-855 (1996);
Montgomery et al., ~, x'1:427-436 (1996); Marsters et at., J. Biol. Chem.,
x_2:14029-14032 (1997); Simonet et
al., ~1 , $Q:309-319 (1997)1. Unlike other kt~wn TNFR-like molecules. Simonet
et al., , report that OPG
contains no hydrophobic transmembrane-spanning seduence.
6
CA 02421319 2003-03-25
WO 99!46281 PCT/US99/05028
Moreover, a new member of the TNFINGF receptor family has been identified in
mouse, a receptor referred
to as "GTI'R" for "glucocorticoid-induced tumor necrosis factor receptor
family-related gene" [Nocentini et al., ]?roc.
,~latl. Acad. Sci. USA 94:6216-6221 (1997)]. The mouse GITR receptor is a 228
amino acid type I transmembrane
protein that is expressed in normal mouse T lymphocytes from thymus, spleen
and lymph nodes. Expression of the
mouse GITR receptor was induced in T lymphocytes upon activation with anti-CD3
antibodies, Con A or phorbol
12-myristate 13-acetate. It was speculated by the authors that the mouse GITR
receptor was involved in the
regulation of T cell receptor-mediated cell death.
In Marsters et al., Curr. Biol.; 6_:750 (1996), investigators describe a full
length native sequence human
polypeptide, called Apo-3, which exhibits similarity to the TNFR family in its
extracellular cysteine-rich repeats and
resembles TNFR1 and CD95 in that it contains a cytoplasmic death domain
sequence [see also Marsters et al., rr.
lo!., ø:1669 (1996)]. Apo-3 has also been referred to by other investigators
as DR3, wsl-1 and TRAMP
[Chinnaiyan et al., Sc' ce, 274:990 {1996); Kitson ec al., Nature, X4:372
(I996); Bodmer et al., Immunity, ø:79
(1997)] . '
Pan et al. have disclosed another TNF receptor family member referred to as
"DR4" [Pan et al., Sci ,
x,7ø:111-1 I3 (1997)J. The DR4 was reported to contain a cytoplasmic death
damain capable of engaging the cell
suicide apparatus. Pan et al. disclose that DR4 is believed to ix a r~eptor
for the ligand known as Apo-2 ligand or
TRAIL.
In Sheridan et al., c'ence, 277:818-821 (1997) and Pan et al., S ' ce, ?7:815-
818 (1997), another
molecule believed to be a receptor for the Apo-2 ligand (TRAIL) is described.
That molecule is referred to as DR5
(it has also been alternatively referred to as Apo-2). Like DR4, DRS is
reported to contain a cytoplasmic death
domain and be capable of signaling apoptosis.
In Sheridan et al., ,s~Dra, a receptor called DcRI (or alternatively, Apo-
2DcR) is disclosed as being a
potential decoy receptor for Apo-2 Iigand (TRAIL). Sheridan et al, report that
DcRl can inhibit Apo-2 ligand
function in vitro. See also, Pan et al., ~,upra, for disclosure on the decoy
receptor referred to as TRID.
For a review of the TNF family of cytokines and their receptors; see Gruss and
Dower. SUDta.
As presently understood, the cell death program contains at least three
important elements - activators,
inhibitors, and effectors; in C, elegaru, these elements are encoded
respectively by three genes, Ced-4, Ced 9 and
Ced 3 [Steller, ie , x'7:1445 (1995); Chinnaiyan et al., ci , X5:1122-1126
(1997); Wang et a1, t~, QQ:I-
20 (199'1)]. Two of the TNFR family members, TNFRI and FasIApol (CD95), can
activate apoptotic cell death
[Chinnaiyan and Dixit, Current Biology, ø:555-562 (1996); Fraser and Evan,
Wit; X5;781-784 (1996)]. TNFRl is
also known to mediate activation of the transcription factor, NF-xB (Tartaglia
et al" ~, 74:845-853 (I993); Hsu
et al., ~gj], $4:299-308 (1996)]. In addition to some ECD homology, these two
receptors share homology in their
intracellular domain øCD) in an oiigometization interface known as the death
domain ['fartagIia et al., supra; Nagata,
~, $x:355 (I997)]. Death domains are also found in several metazoan proteins
that regulate apoptosis, nar~ly,
the Drosophila protein, Reaper, and the mammalian proteins referred to as
FADDIMORTl , TRADD, and RIP
[Cleaveland and lttle, ~g[t, ,$],:479-482 (1995)].
Upon ligand binding and receptor clustering, TNFR1 and CD95 are believed to
recruit FADD into a death-
inducing signalling complex. CD95 purportedly binds FADD directly, while TNFRl
binds FADD indirectly via
TRADD (Chinnaiyan et al., t~, $[,:505-512 (1995); Boldin et al:, j~ol. Chem.,
7~t :387-391 (1995); Hsu et al.,
supra; Chinnaiyan et al., J. Bip~. Chem., x:4961-4965 (1996)]. It has been
reported that FADD serves as an
7
CA 02421319 2003-03-25
WO 99/46281 PCT/US99I05028
adaptor protein which recruits the Ced-3-related protease, MACI~iaIFLICE
(caspase 8), into the death signalling
complex [Boldin et al., Celi, 85:803-815 (1996); Muzio et al., Ce 1, 85:817-
827 (1996)]. MACHa/FLICE appears
to be the trigger that sets off a cascade of apoptotic proteases, including
the interleukin-1 p converting enzyme (ICE)
and CPP32IYatna, which may execute some critical aspects of the cell death
programme [Eraser and Evan, a ra .
It was recently disclosed that programmed cel! death involves the activity of
members of a family of cysteine
pmteases related to the C. etegans cell death gene, ced-3, and to the
mammalian IL-1-converting enzyme, ICE. The
activity of the ICE and CPP32IYama proteases can be inhibited by the product
of the cowpox virus gene, crmA [Ray
et aL, C~,11, 69:597-604 (i992); Tewari et al., Cell, 8:801-809 (1995)].
Recent studies show that CrmA can inhibit
TNFRl- and CD95-induced cell death [Enari et al., Na e, 37~. :?8-81 (1995);
Tewari et al., J. Biol. Chem.,
x:3255-3260 (1995)].
As reviewed recently by Tewari et al., TNFRl, TNFR2 and CD40 modulate the
expression of
proinflatnntatory and costimulatory cytoldnes, cytolcine receptors, and cell
adhesion molecules through activation of
the transcription factor, NF-xB (Tewari et al., burr. 0g. Genet. Develop.,
6_:39-44 (1996)]. NF-xB is the prototype
of a family of dimeric uanscription factors whose subunits contain conserved
Rel regions [Verma et al., Genes
Develop., x:2723-2735 (1996); Baldwin, Ann. Rev. Immunol., x_4:649-681
(1996)]. In its latent form, NF-tcB is
complexed with members of the IxB inhibitor family; upon inactivation of the
IxB in response to certain stimuli,
released NF-xB translocates to the nucleus where it binds to specific DNA
sequences and activates gene transcription.
10. PR0382
Proteases are enzymatic proteins which are involved in a large number of very
important biological
processes in mammalian and non-mammalian organisms. Numerous different
protease ertzyrnes from a variety of
different mammalian and non-matrunalian organisms have been both identified
and characterized, including the serine
proteases which exhibit specific activity toward various serine-containing
proteins. The mammalian protease enzymes
play important roles in biological processes such as, for example, protein
digestion, activation, inactivation, or
modulation of peptide hormone activity, and alteration of the physical
properties of proteins and enzymes.
In light of the important physiological roles played by protease enzymes,
efforts are currently being
undertaken by both industry and academia to identify new, native protease
homologs. Many of these efforts are
focused on the screening of mammalian recombinant DNA libraries to identify
the coding sequences for novel
membrane-bound receptor proteins. Examples of screertirtg methods and
techniques are described in the literature
[see, for example, Klein et al., Proc. Natl. Acad. Sci., 23:7108-7113 (1996);
U.S. Patent No. 5,536,637)]. We
herein describe the identification of novel polypeptides having homology to
serine protease enzymes, designated
herein as PR0382 polypeptides.
11. pR0545
The ADAM (A Disintegrin And Metalloprotease) family of proteins of which
meltrin is a member may have
an important role in cell interactions and in modulating cellular responses.
[see, for example, Gilpin et al., . Biol
Chem., 27311):157-166 {1998)]. The ADAM proteins have been implicated in
carcinogenesis. Meltrin-a (ADAM 12)
is a myoblast gene product reported to be required for cell fusion. [Harris et
al., J. Cell. Biochem., ~70:136-142
(1997), Yagami-Hiromasa et al., Nature, .x:652-656 (1995)]. The meltrins
contain disintegrin and metalloprotease
donrairts and are implicated in cell adhesive events involved in development,
through the integrin binding disintegrin
8
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domain, but also have an anti-adhesive function through a zinc-dependent
metalloprotease domain. [Alfandari et al.,
Devel. Biol., l 2 2 :314-330 (1997)]. Given the medical importance of cell
fusion and modulation of cellular
responses in carcinogenesis and other disease mechanisms, efforts are
currently being under taken to identify new,
native proteins which are involved in cell fusion and modulation of cellular
responses. We describe herein the
identification of a novel polypeptide which has homology to meltrin,
designated herein as PR0545.
I2. PR 7
CD24 is a protein that is associated with the cell surface of a variety of
different cells of the mammalian
immune system, including for example, neutrophils, monocytes and some
lymphocytes, for example, B lymphocytes.
CD24 has been shown to be a ligand for the platelet-associated surface
glycoprotein P-selectin (also known as granule
membrane protein-140 or GMP-140), a glycoprotein that is constitutively
synthesized in both platelets and endothelial
cells and becomes exposed on the surface of platelets when those cells become
activated. In this way, P-selectin
mediates the calcium-dependent adhesion of activated platelets and endothelial
cells to the various cells of the immune
system that express one or more ligands for the P-selectin molecule,
particularly CD24. This mechanism allows for
recruitment of immune system cells to locations where they are most needed,
for example, sites of injury. Thus,
there is substantial interest in identifying novel polypeptides that exhibit
homololry to the cell surface antigens of the
immune system cells. We herein describe the identification and
characterization of a novel polypeptide having
homology to the CD24 protein, wherein that novel polypeptide is herein
designated PR0617.
13. PR070a
Protein-disulfide isomerase (PDn is a catalyst of disulfide formation and
isomerization during protein
folding. It has two catalytic sites housed in two domains homologous to
thioredoxin, one near the N terminus and
the other near the C terminus. [See for example, Gilbert HF, J.Biol.Chem.,
47:29399-29402 (1997}, Mayfield KJ,
Science, 278:1954-1957 (1997) and Puig et al., ~Biol.Chem., 52:32988-32994
(1997)]. PDl is useful for formation
of natural type disulfide bonds in a protein which is produced in aprokaryotic
cell. (See also, U.S. Patent Nos.
5,700,659 and 5,700,678).
Thus, PDI and molecules related thereto are of interest, particularly for
ability to catalyze the fornnation of
disulfide bonds. Moreover, these molecules are generally of interest in the
study of redox reactions and related
processes. PDI and related molecules are further described in Darby, et al.,
Biochemistry 34; 11725-11735 {1995).
We herein describe the identification and characterization of novel
polypeptides having homology to protein disulfide
isomerase, designated herein as PR0700 polypeptides.
14. PR0702
Conglutmin is a bovine serum protein that was originally described as a
vertebrate lectin protein and which
belongs to the family of C-type lectins that have four characteristic domains,
(I) an N-terminal cysteine-rich domain,
(2) a collagen-Like domain, (3) a neck domain and (4} a carbohydrate
recognition domain (CRD). Recent reports have
demonstrated that bovine conglutinin can inhibit hemagglutination by influenza
A viruses as a result of their lectin
properties (Eda et al., Biochem. J. 316:43-48 (1996)). It has also been
suggested that Iectins such as conglutinin can
function as immunoglobulin-independent defense molecules due to complement-
mediated mechanisms. Thus,
conglutinin has been shown to be useful for purifying immune complexes ire
vitro and for removing circulating
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immune complexes from patients plasma in vivo (Lim et al., Biochem. Biophys.
Res. Commun. 218:260-266 {1996)).
We herein describe the identification and characterization of a novel
polypeptide having homology to the conglutinin
protein, designated herein as PR0702.
15. PR0703
Very-long-chain acyl-CoA synthetase ("VLCAS") is a long-chain fatty acid
transport protein which is active
in the cellular transport of long and very long chain fatty acids. [see for
example, Uchida et al., J iochem (Tokyo)
119(3):565-571 (1996) and Uchiyama et al., J Biol Chem 271(48):30360-30365
{/996). Given the biological
importance of fatty acid u~anspor< mechanisms, efforts are currently being
under taken to identify new, native proteins
which are involved in fatty acid transport. We describe herein the
identification of a novel polypeptide which has
homology to VLCAS, designated herein as PRO703.
16. Rg 0705
The glypicans are a family of glycosylphosphatidylinositol {GPI)-anchored
proteoglycans that, by virtue of
their cell surface localization and possession of heparin sulfate chains, may
regulate the responses of cells to
numerous heparin binding growth factors, cell adhesion molecules and
extracellular matrix components. Mutations
in one glypican protein cause of syndrome of human birth defects, suggesting
that the glypicans may play an
important role in development (Litwack et al., Dev. 211:72-87 (1998)). Also,
since the glypicans may interact
with the various extracellular matrices, they may also play important roles in
wound healing (McGrath et al., Pathol.
183:251-252 (1997)). Furthermore, since glypicans are expressed in neurons and
glioma cells, they may also play
an important role in the regulation of cell division and survival of cells of
the nervous system (Liang et al., J. Cell.
Biol_ 139:851-864 (1997)). It is evident, therefore, that the glypicans are an
extremely important family of
groteoglycans. There is, therefore, substantial interest in identifying novel
polypegtides having homology to members
of the glypican family. We herein describe the identification and
characterization of a novel polypeptide having
homology to K-glypican, designated herein as PR0705.
17. R 8
Aryl sulfatases. ate enzymes that exist in a number of different isoforms,
including aryl sulfatase A (ASA),
aryl sulfatase B (ASB) and aryl sulfatase C (ASC), and that function to
hydrolyze a variety of differem aromatic
sulfates. Aryl sti)fatases have been isolated from a variety of different
animal tissues and microbial sources and their
structures and functions have been extensively studied (see, e.g., Nichol and
Roy, J. Biochem. 55:643-651 (1964)).
ASA deficiency has been reported to be associated with metachromatic
leukodystrophy (MLD) (tiles et al., Prenat.
is n. 7(4):245-252 (/987) and Herska et al" ,Ami J. Med. Genet. 26(3):629-635
(1987)). Additionally, other
groups have reported that aryl sulfatases have been found in high levels in
natural killer cells of the immune system
and have hypothesized a possible role for these enzyrttes in NK cell-
rt>ediated cellular lysis (see, e.g., Zucker-Franklin
et al., Proc. ail. Acad. Sci. USA 80(22):6977-6981 (1983)). Given the obvious
physiological importance of the
aryl sulfatase enzymes, there is a substantial interest in identifying novel
aryl sulfatase homolog polypeptides. We
herein descn'be the identification and characterization of novel polypeptides
having homology to the aryl sulfatases,
wherein these novel polypeptides are herein designated PR0708 polypeptides.
CA 02421319 2003-03-25
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18. P_ RO, 20
Fibulin-1 is a cysteine-rich, calcium-bituiing. extracellular matrix (ECM)
component of basement membranes
and connective tissue elastic fibers and plasma protein, which has four
isoforms, A-D, derived from alternative
splicing. Fibulin-I is a modular glycoprotein with amino-terminal anaphlatoxin-
like modules followed by nine
. epidermal growth factor (EGF)-like modules and, depending on alternative
splicing, four possible carboxyl termini.
. 5 Fibulin 2 is a novel extracellular matrix protein frequently found in
close association with microfibrils containing
either fibronectin or fibrillin. There are multiple forms of fibulin-1 that
differ in their C-terminal regions that are
produced through the process of alternative splicing of their precursor RNA.
[see for example Tran et al., ati
'~I 15(7):479-493 (1997).]
Northern and Western blotting analysis of 16 cell lines established from
tumors formed in athymic mice and
malignant cell lines derived from patients indicate that low expression of
fibulin-1D plays a role in tumor formation
and invasion. [Qing et al., Oncogene, 18:2159-2168 (1997)]. Ovarian-cancer
cells are charactei'fzed by their ability
to invade freely the peritoneal cavity: It has been demonstrated that
estradiol stimulates the proliferation of estrogen-
receptor (ER)-positive ovarian-cancer cells, as well as expression of fibulin-
1. Studies on the effect of fibulin-i on
motility of the MDA-MB231 breast-cancer cell line, indicated inhibition
ofhaptotactic migration of MDA-MB231
cells, atxi the authors cotxluded that fibulin-1 can inhibit cancer cell
motility inn vitro and therefore has the potential
to inhibit tumor invasion. [Hayashido et al., lnt J Cancer , 75(4):654-658
(1998)]
Thus, fibulin, and molecules related thereto are of interest, particularly for
the use of preventing cancer.
Moreover, these molecules are generally of interest in the study of connective
tissue and attachmern molecules and
related mechanisms. Fibulin and related molecules are further described in
Adams, et al., J. Mgl. Biol.,
272(2):226-36 (1997); Kielty and Shuttleworth, Microse. Res. Tech., 38(4):413-
27 (1997); and Chil ..d 3CaL,. Surg,.
12(2Supp.):i31-5 (1997).
We herein describe the identification and characterization of novel
polypeptides having homology to fibulin;
designated herein as PR0320 polypeptides.
19. PRO. ?~
Oxidoreductases are that catalyze a reaction in which two molecules of a
compound interact so that
one molecule is oxidized and the other is reduced, with a molecule of water
entering the reaction. There are many
different types of oxidoreductase enzy~s that play very important
physiological roles in the mammalian organism.
Some of the most important oxidoreductases inchtde, for example, lyases,
lactases, cholesterol oxidases, and the like.
These enzymes play roles in such essential processes as digestion, signal
transduction, maintenance of ionic
homeostasis, and the like. As such, given that oxidoreductase enzymes find
various essential uses in the marturiafian
organism, there is a substantial interest in identifying novel oxidoreductase
enzyme homology. We herein describe
the identification and characterization of a novel poiypepdde having homology
to oxidoreductases, designated herein
as PR0324.
20. PR03~
Prostasin is a novel human seritte proteinase purified from human seminal
fluid. Imtrnmohistochemical
localization reveals that prostasin is present in epithelial cells and ducts
of the prostate gland. The cDNA for
prostasin has been cloned and characterized. Southern blot analysis, following
a reverse transcription polymerise
11
CA 02421319 2003-03-25
WO 9914b281 PCTlUS99l05028
chain reaction, indicates that prostasin mRNA is expressed in prostate, liver,
salivary gland, kidney, lung, pancreas,
colon, bronchus, renal proximal tubular cells, and prostate carcinoma LNCaP
cells. Cellular localization of prostasin
trtRNA was identified within epithelial cells of the human prostate gland by
in situ hybridization histochemistry. [See
for example, Yu et al., J Biol Chem. (1994) 269(29):18$43-18848, and Yu et
a1.,1 Biol Chem. (1994) 270(22):13483-
13489].
Thus, prostasin, and molecules related thereto are of interest, particularly
for the study, diagnosis and
treatment of medical conditions involving the prostate. Prostasin and related
molecules are further described in Yu
et al., Genomics (1996) 32(3):334-340. We herein describe the identification
and characterization of novel
polypeptides having homology to prostasin, designated herein as PR0351
polypeptides.
21. PR0352
Buryrophilin is a milk glycoprotein that constitutes more than 40% of the
total protein associated with the
fat globule tttembrane in marrunalian milk. Expression of butyrophilin mRNA
has been shown to correlate with the
onset of milk fat production toward the end pregnancy and is maintained
throughout lactation. Buryrophilin has been
identified in bovine, murine and human (see Taylor et al., Bioci~ir~,,
Biophys. Acta 1306:1-ll (1996), Ishii et al.,
Biochim. Biophys. Acts 1245:285-292 (1995), Mather et al., J.~ Sci. 76:3832-
3850 (1993) and Banghart et al.,
~. Biol. Chem. 2'13:4171-4179 (1998)) and is a type I transmembrane protein
that is incorporated into the fat globulin
membrane. It has been suggested that buryrophilin may play a role as the
principle scaffold for the assembly of a
complex with xanthine dehydrogenase/oxidase and other proteins that function
in the budding and release of milk-fat
globules from the apical surface during lactation (Banghart et al., supra).
Given that butyrophilin plays an obviously important role in mammalian milk
production, there is substantial
interest in identifying novel butyrophilin homologs. We herein describe the
identification and characterization of a
novel polypeptide having homology to buryrophilin, designated herein as
PR0352.
22. P,~t0381
The imrnuttophilins are a family of proteins that function as receptors for
immunosuppressant drugs, such
as cyclosporin A, FK506, and rapamycin. The immunophilins occur in two
separate classes, (1) the FK506-binding
proteins (FKBPs), which bind to FIC506 and rapamycin, and (2) the
cyclophiiins, which bind to cyclosporin A. With
regard to the FKSt?6-binding proteins, it has been reported that the
FK506lFKBP complex functions to inhibit the
activity of the serinelthreonine protein phosphatase 2B (calcineurin), thereby
providing immunosuppressant activity
(Gold, Mol. Neu,~biol. 15:285-306 (1997)). It has also been reported that the
FICBP immunophilins are found in
the mammalian nervous system and may be involved in axonal regeneration in the
cenual nervous system thr~gh
a mechanism that is independent of the process by which immunosuppression is
achieved (Gold, supra). Thus, there
is substantial interest in identifying novel polypeptides having homology to
the FKBP immunophilins. We herein
describe the identification and characterization of a novel polypepiide having
homology to an FKBP immunophiiin
protein, designated herein as PR0381.
23. PR 8
Mammalian cell membranes perform very important functions relating to the
structural integrity and activity
of various cells and tissues. Of particular interest in membrane physiology is
the study of transmembrane ion
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channels which act to directly control a variety of physiological,
pharmacological and cellular processes. Numerous
ion channels nave been identified including calcium (Ca), sodium (Na) and
potassium (K) channels, each of which
have been analyzed in detail to determine their roles in physiological
processes in vertebrate and insect cells.
One type of cell membrane-associated ion channel, the sodium channel, plays an
extremely important role
in a cell's ability to maintain ionic homeostasis as well as transmit
intraceilular~ and extracellular signals. Voltage-
gated sodium channels in brain neurons have been shown to be complexes of a
pore-forming alpha unit with smaller
beta-1 and beta-2 sulntnits (Isom et al., CSI 83:433-442 (1995)). Given the
obvious importance of sodium channels
in cellular homeostasis and other important physiological functions, there is
a significant interest in identifying novel
polypeptides having homology to sodium channel subunits. We herein describe
the identification and characterization
of a novel polypeptide having homology to the beta-2 subunit of the rat sodium
channel, designated herein as
PR0386.
24. PR054l1
Lecithin-cholesterol acyltransferase {"LCAT"), also known as
phosphatidylcholine-sterol acyltransferase is
a key enzyrtre in the intravascular metabolism of high density lipoproteins,
specifically in the process of cholesterol
metabolism. (see, for example, Brousseau et a1.,1. Lipid Res., 38(12):2537-
2547 (I997), Hill et aL, Biochem. J.,
294:879-884 (1993), and Drayna et al., Na r 327 (6123):632-634 (1987)]. Given
the medical importance of lipid
metabolism, efforts are currently being tinder taken to identify new, native
proteins which are involved in lipid
transport. We describe herein the identification of a novel polypeptide which
has homology to LCAT, designated
herein as PR0540.
25. PR06I5
Synaptogyrin is a synaptic vesicle protein that is uniformly disuibuted in the
nervous system. The cDNA
encoding synaptogyrin has been cloned and sequenced and the sequence predicts
a protein with a molecular mass of
25,900 D with four membrane-spanning domains. Synaptogyrin has been implicated
in membrar>e traffic to and from
the plasma membrane. Stenius et al., J. Cell. Biol. 131{fi-2):18U1-1809
(1995). In addition, a novel isoform of
synaptogyrin called cellugyrin exhibits sequence identity with synaptogyrin.
In rat tissues, cellugyrin and
synaptogyrins are expressed in mirror image patterns. Cellugyrin is
ubiquitously present in all tissues tested with the
lowest levels in brain tissue, whereas synaptogyrin protein is only detectable
in brain. In rat tissues, cellugyrin and
synaptogyrins are expressed in mirror image patterns. The synaptic vesicle
protein synaptogyrin may be a specialized
version of a ubiquitous protein, cellugyrin, with the two proteins sharing
structural similarity but differing in
localization. 'This finding supports the emerging concept of synaptic vesicles
as the simplified and specialized form
of a generic uafficking organelle. [lanz et al., J. Biol. Chem. 273(5):2851-
2857 {1998)] . The sequence for
cellugyrin derived from the Norway rat, Ranus norvegicus has been deposited in
the Genbank database on 23
December 1997, designated accession rntntber AF039085. See also, Janz et al.,
J. Biol. Chem. 273 (1998), in press.
Given the medical importance of synaptic transmission, efforts are currently
being under taken to identify
new, native proteins that may be pan of a simplified and specialized generic
trafficking organelle in the form of
synaptic vesicles. We describe herein the identification of a novel
polypeptide which has homology to synaptogyrin,
designated herein as PR0615.
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CA 02421319 2003-03-25
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26. PR0618
Enteropeptidase is a key enzyme in the intestinal digestion cascade
specifically cleaves the acidic propeptide
from trypsinogen to yield active trypsin. This cleavage initiates a cascade of
proteolytic reactions leading to the
activation of many pancreatic zymogens.
See, for example, Matsushima et al., J. 131o1. Chem. 269(31):19976-19982
(1994), Kitamoto et al., Proc: Nat. Acad.
~, 91(16):7588-7592 (1994). Enterokinase (enteropeptidase) is a related to
mammalian serine proteases involved
in digestion, coagulation, and fibrinolysis. LaVallie et al., J Biol Chem.,
268(31):23311-23317 (1993).
Given the medical importance of digestive processes, efforts are currently
being under taken to identify new,
native proteins that may be involved in digestion, coagulation; and
fibrinolysis. We describe herein the identification
of a novel polypeptide which has homology to enteropeptidase, designated
herein as PR0618.
27. RP 0719
Lipoprotein lipase is a key enzyme that mediates the hydrolysis of
triglycerides and phospholipids present
in circulating plasma lipoproteins (Dugi et al., J. Biol. Chem.. 270:25396-
25401 (1995)). Moreover, lipoprotein
lipase has been shown to mediate the uptake of lipoproteins into cells,
wherein cellular uptake of lipoproteins is
initiated by binding of lipoprotein lipase to cell surface proteoglycans and
to the low density lipoprotein (LDL)
receptor-related protein (Krapp et al., J. Lipid Res. 36:2362-2373 (1995)).
Thus, it is clear that lipoprotein lipase
plays an extremely important role in lipoprotein and cholesterol metabolism.
There is, therefore, substantial interest
in identifying novel polypeptides that share sequence homology and/or
biological activity with lipoprotein lipase. We
herein describe the idemificauon and characterization of a novel polypeptide
having sequence homology to lipoprotein
lipase H, designated heein as PR0719.
28. PR
The low density lipoprotein (LDL) receptor is a membrane-bound protein that
plays a key role in cholesterol
homeostasis, mediating cellular uptake of lipoprotein particles by high
affinity binding to its ligands, apolipoprotein
(apo) B-100 and apoE. The ligand-binding domain of the LDL receptor contains 7
cysteine-rich repeats of
approximately 40 amino acids, wherein each repeat contains 6 cysteines, which
form 3 intra-repeat disulfide bonds.
These unique structural features provide the LDL receptor with its ability to
specifically interact with apo B-100 and
apoE, thereby atlowing for transport of these lipoprotein particles across
cellular membranes and metabolism of their
components. Soluble fragments containing the extracetlular domain of the LDL
receptor have been shown to retain
the ability to interact with its specific lipoprotein ligands (Simmons et al.,
jL~3iol. C ;em. 272:25531-25536 {1997)).
Thus, it is clear that the LDL t~ecxptor is intimately involved in importam
physiological activities related to cholesterol
tnetabolism. As such, there is substantial interest in identifying novel LDL
receptor homolog proteins. We herein
describe the identification and characterization of a novel polygeptide having
homology to the human LDL receptor
protein, designated herein as PR0724..
29. ? 2
Expression of the human gene A4 is enriched in the colonic epithelium and is
transcriptionally activated on
differentiation of coIonic epithelial cells in vitro (OIiva et al. , arch.
Biochem. Bionhvs. 302:183-192 (1993) and Oliva
tt al., Am. J. Physiol. 272:C957-0965 (1997)). A4 cDNA contains an open
reading frame that predicts a polypeptide
14
CA 02421319 2003-03-25
WO 99146281 PCT/IIS99/05028
of approximately 17 kilodaltons in size. Hydropathy analysis of the A4 protein
revealed four putative membrane-
spanning alpha-helices. Immunocytochernical studies of cells expressiutg A4
protein indicated that expression is
localized to the endoplasmic, reticulum. The four membrane-spanning domains
and the biophysical characteristics
of the A4 protein suggest that it belongs to a family of integral membrane
proteins called proteoligids, some of which
multimerize to form ion channels. In fact, preliminary evidence has suggested
that A4 may itself multimerize and
take on the properties of an ion channel (OIiva et al., Arr~ J. Phvsiol.
272:C957-C965 (1997)). Given the importance
of ion channels in maintaining cellular homeostasis, there is a significant
interest in identifying novel polypeptides
having homology to lmown and putative ion channels. We herein describe the
identification and characterization of
a novel polypepdde having homology to the putative ion channel protein, A4,
designated herein as PR0772.
I0 38. ~R0852
Proteases are enzymatic proteins which are involved in a large number of very
important biological
processes in mammalian and non-mammalian orgatusms. Numerous different
protease enzymes from a variety of
different mammalian and non-mammalian organisms have been both identified and
characterized. The mammalian
protease enzymes play important roles in many different biological processes
including, for example, protein
digestion, activation, inactivation, or modulation of peptide hormone
activity, arid alteration of the physical properties
of proteins and enzymes.
In light of the important physiological roles played by protease enzymes,
efforts are currently being
undertaken by both industry and academia to idenfify new, native protease
homologs. Many of these efforts are
focused on the screening of mammalian recombinant DNA libraries to identify
the coding sequences for novel
secreted and membrane-bound receptor proteins. Examples of screening methods
and techniques are described in
the literature [see, for example, Klein et aL, Proc. Natl. Acad. Set., x:7108-
7113 (1996); U.S. Patent No.
5,536,637)]. We herein describe the identification of novel polypeptides
having homology to various protease
enzymes, designated herein as PR0852 polypeptides.
31. PR 853
Studies have reported that the redox state of the cell is an important
determinant of the fate of the cell.
Furthermore, reactive oxygen species have been reported to be cytotoxic,
causing inflammatory disease, including
tissue necrosis, organ failure, atherosclerosis, infertility, birth defects,
premature aging, mutations and malignancy.
Thus, the control of oxidation and reduction is important for a number of
reasons, including the control and
prevention of strokes, heart attacks, oxidative stress, hypertension and may
be associated with the development of
malignancies. The levels of antioxidant enzymes, such as reductases, which
catalyze the conversion of reactive
oxygen species to water have been simwn to be low in cancer cells. In
particular, malignant prostate epithelium may
have lowered expression of such antioxidant enzymes [Baker et ., s to
32(4):229-233 (I997)). In this regard,
reductases, are of interest. In addition, the transcription factors, NF-kappa
B and AP-1, are known to be regulated
by redox state and to affect the expression of a large variety of genes
thought to be involved in the pathogenesis of
AIDS, cancer, atherosclerosis and diabetic complications. Publications further
describing this subject matter include
et aL, Anticancer Res. fGreecel, 17:4599-4605 (1997), Kelsey, et al., [3r. Ji
Cancer, 76(7):852-4 (1997);
Friedrich acrd Weiss. 1. TheQr, 'o1., 187(4):529-00 (1997) and Pieulle, et
al., J. Bacteriol., 179(18):5684-92 (1997).
Given the physiological importance of redox reactions in tdvo, efforts are
currently being under talten to identify 'new,
CA 02421319 2003-03-25
WO 99!46281 PCT/EJS99/05028
native proteins which are involved in redox reactions. We describe herein the
identification of a novel prostate
specific polypeptide which has sequence similarity to reductase, designated
herein as PR0853.
32. PR 860
Neurofascin is a member of the Ll subgroup of the cellular adhesion molecule
("CAM") family of nervous
system adhesion molecules and is involved in cellular aggregation. CeII-cell
recognition and patterning of cell
contacts have a critical role in mediating reversible assembly of a wide
variety or transcellular complexes in the
nervous system. Cell interactions may be regulated through modulation of
anlryrin binding to neurofascin. See, for
example, Tuvia et al., Proc. Nat Acad. Sci., 94(24) 12957-12962 (1997).
Neurofascin has been described as a
member of the Ll subgroup of the immunoglobulin superfamily implicated in
neurite extension during embryonic
development for which numerous isoforms have been detected at various stages
of development. See also Hassel et
al., J. Biol. Chem., 272(45) 28742 28749 (1997), Grtlmet., Ce,~. Tissue Red
290(2) 423-428 (1997), Gatver et al.,
J. Cell. Biol., 137:703-7i4 (1997), and Lambent et al., J Neurosci., 17:7025-7-
36 (1997),.
Given the physiological importance of cellular adhesion rnoiecules and
development of the nervous system
in vivo, efforts are currently being under taken to identify new, native
proteins which are involved in regulation of
cellular interactions in the nervous system. We describe herein the
identification and characterization of a novel
polypeptide which has sequence similarity to neurofascin, designated herein as
PR0860.
33. ~,R_ 8046
The CMRF35 monoclonal antibody was used to identify a cell membrane antigen,
designated CMRF35,
which is present on the surf~e of moliochces; neutrophils, a proportion of
peripheral blood T and B lymphocytes and
lymphocytic cell lines. The CMRF35 cDNA encodes a novel integral membrane
glycoprotein member of the
immttnoglobulin (Ig) gene superfamily. The molecule comprises (a) a single
extracelluiar Ig variable domain
remarkably similar to the Fc receptor for polymeric IgA and IgM, (b) a
membrane-proximal domain containing a high
proportion of proIine, serine and threonine residues that was predicted to be
heavily O-glycosylated, (c) an unusual
transtnembrane anchor that contained a glutamic acid and a proline residue and
(d) a short cytoplasmic tail.
Transcripts encoding the CMRF35 protein have been detected in early mortocyric
cell lines, in peripheral blood T
cells and in some B Lymphoblastoid cell lines, confirming the results of
immunocytological staining. Jackson et al.,
Eur. J~. Imnn>ttol. 22(5):1157-1163 (1992). CMRF-35 molecules are
differentially expressed in hematopoietic cells,
and the expression of the antigen was shown to be markedly influenced by
stimulation with mitogens and cytokines.
See, for example, Clark et al:, Exp. Hematol. 25(8):759 (1997), Daish et al.,
ynol. 79(1):55-63 (1993), and
Clark et al., Tjssue Antigens 48:461 (1996).
Given the physiological importance of the immune system and antigens
associated with various immune
system cells, efforts are currently being under taken to identify new, native
proteins which are expressed on various
cells of the immune system. We describe herein the identification of a novel
polypeptide which has sequence
similarity to CMRF35, designated herein as PR0846.
34. gR0862
Lysozyme is a protein which is widely distributed in several human tissues and
secretions including milk,
tears and saliva. It has been demonstrated to hydrolyze Linkages between N-
aeetylglucosarnines. It has been
!b
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demonstrated to be an inhibitor of chemotaxis and of the production of toxic
oxygen free radicals and may also have
some role in the calcification process. As such, there is substantial interest
in identifying novel polypeptides having
homology to lysozyme. We describe herein the identification of a novel
polypeptide which has sequence similarity
to lysozyme.
S 35. PR08b4
Wnt-4 is a secreted glycoprotein which correlates with, and is required for,
kidney tubulogenesis. Mice
_, lacking Wnt-4 activity fail to form precubular cell aggregates; however,
other aspects of mesenchymal and ureterie
development are unaffected. Thus, Wnt-4 appears to act as an aucoinducer of
the mesenchyme to epithelial transition
that underlies nephron development. Stark et al., a a ;372(6507):679-683
(1994). In addition, members of the
Wnt gene family code for cysteine-rich, secreted proteins, which are
differentially expressed in the developing brain
and possibly act as intercellular signaling molecules. A Wnt gene, e.g., Wnt-1
is known to be essential for
specification of the midbrain cell fate. Yoshioka et al., Bj;~Birap~vs. Rel.
Co~un. 203(3):1581-1588 (1994).
Several member of the Wnt family of secreted factors are suongly implicated as
regulators of mammary cellular
growth and differentiation. Shimizu et al., Cell Growth Differ. 8(12) 1349-
1358. Wnt-4 is normally expressed in
early pregnancy. Wnt-4 may therefore be a local signal driving epithelial
branching in pregnancy. Edwards PA,
Biochem Soc Symp.63:21-34 (19981. See also, Lipschutz JH, Am. J. ICidnev Dis.
31(3):383-397, (1998). We
descxibe herein the identification and characterizaton of a novel polygeptide
which has sequence similarity to Wnt-4,
designated herein as PR0864.
36. PR0792
At least two cell-derived signals have been shown to be necessary for the
induction of immunoglobulin
isotype switching in B-cells. The first signal is given by either of the
soluble lymphokines, interleukin (IL~4 or IL-
13, which induce germline epsilon transcript expression, but this alone is
insufficient to trigger secretion of
immouttoglobulin E (IgE). The second signal is provided by a physical
interaction between B-cells and activated T-
cells, basophils and mast cells, and it has been shown that the CD40JCD40
ligand pairing is crucial for mediating
IgE synthesis. Additionally, amongst the numerous pairs of surface adhesion
molecules that are involved in IgE
synthesis, the CD231CD21 pair appears to play a key role in the generation of
IgE. CD23 is a protein that is
positively and negatively regulated by factors which increase or decrease IgE
production, respectively. Antibodies
to CD23 have been shown to inhibit IL-4-induced human IgE production in vitro
and to inhibit antigen-specific IgE
responses in a rat model, in an isotype selective manner (Bonnefoy et al.,
Eur. Respir. J. Suvnl. 22:63S-66S (1996)).
CD23 ittcetacu with CD21 on B-cells, preferentially driving IgE production.
Given that the CD23 protein plays an
extremely important role in the induction of a mammalian IgE response, there
is significant interest in identifying
novel polypeptides having homology to CD23. We herein describe the
identification and characterization of a novel
polypeptide having homology to CD23, designated herein as PR0792.
37.
Mindin and spottdin proteir~ are sxreted proteins that are sttucturaily
related to one another and which have
been identified in a variety of organisms. For example, Higashijima et al:,
Dev lo 192:211-227 (1997) have
reported the identification of spondin and mmdin expression in floor plate
cells in the 2ebrafish embryonic axis,
17
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thereby suggesting that mindin and spondin prtoteins play important roles in
embryonic development. This same
group has reported that mindin and spondin proteins function as extracellular
matrix proteins that have a high affinity
for the basal lamina. (Id.). It. has been reported that F-spondin is a
secreted protein that promotes neural adhesion
and neurite extension (Klar et al., Cell 69:95-110 (i992) and that M-spondin
is an extracellular matrix protein that
localizes to muscle attachment sites in Drosophila (IJmemiya et al., ev. Bi 1.
186:165-176 (1997)). 'Thus, there
is significant inteest in identifying novel polypeptides having homology to
the mindin and spondin proteins. We herein
describe the identification and characterization of a novel polypeptide having
homology to mindin2 and mindini,
designated herein as PR0866_
38. PR0871
Cyclophilins are a family of proteins that bind to cyclosporin A and possess
peptidyl-prolyl cis-trans
isomerase activity (Sherry et aL, Proc. Natl. Acad. Sci. USA 95:1758-1763
(1998)). In addition, cyclophilins are
secreted by activated cells and act in a cytokine-like manner, presumably via
signaling through a cell surface
cyclophilin receptor. Host cell-derived cyclophilin A has been shown to be
incorporated into HIV-1 virions and its
incorporation has been shown to be essential for viral infectiviry. Thus, one
or more the cyclophilins may be directly
associated with HIV-1 infectivity. Given the obvious importance of the
cyclophilin proteins, there is substantial
interest in identifying novel polypeptides which have sequence homology to one
or more of the cyclophilin proteins.
We herein describe the identification and characterization of a novel
polypeptide having homology to cyclophilin-like
protein CyP-60, designated herein as PR0871.
39. PRfJ873
Enzymatic proteins play important roles in the chemical reactions involved in
the digestion of foods, the
biosynthesis of macromolecules, the controlled release and utilization of
chemical energy, and other processes
necessary to sustain life. Enzymes have also been shown to play important
roles in combating various diseases and
disorders. For example, liver carboxylesterases have been reported to asslSt
in sensitizing human tumor cells to the
cancer prodrugs. Danks et al., report that stable expression of the cDNA
encoding a carboxylesterase in Rh30 human
rhabdomyosarcoma cells increased the sensitivity of the cells to the CPT-1I
cancer prodrug 8.1-fold. Cancer Res.
(1998) 58(1):20-22. The authors propose that this prodruglenzyme combination
could be exploited therapeutically
in a manner analogous to approaches currently under investigation with the
combinations of ganciclovirlherpes
simplex virus thymidine kinase and 5-fluorocytosine/cytosine deaminase. van
Pelt et al. demonsuated that a 55 kD
human liver carboxylesterase inhibits the invasion of Plasmodium falciparum
malaria sporozoites imo primary human
hepatocytes in culture. He t 1 (1997) 27(4):688-698.
Carboxylesterases have also been found to be of importance in the
dewxification of drugs, pesticides and
other xenobiotics. Purified htttnart liver carboxylesterases have been shown
to be involved in the metabolism of
various drugs including cocaine and heroin. Prindei et at. describe the
purification anti cloning of a broad substrate
specificity human liver carboxylesterase which catalyzes the hydrolysis of
cocaine and heroin and which may play
an important role in the degradation of these drugs in human tissues. J. 8iol.
Chem. (1997) 6:272(23):14769-14775.
Brienzinsld et al. describe a spectrophotometric competitive inhibition assay
used to identify drug or environmental
esters that are metabolized by carboxylesterases. jZrrn Iy~e b Dispos (1997)
25(9):1089-1096.
18
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In light of the irriportant physiological roles played by carboxylesterases,
efforts are being undertaken by
both industry and academia to identify new, native carboxylesterase homologs.
We herein describe the identification
and characterization of a novel polypeptide having homology to
carboxylesterase, designated herein as PR0873.
40. ~R094_0
CD33 is a cell-surface protein that is a member of the sialoadhesin family of
proteins that are capable of
mediating sialic-acid dependent binding with distinct speciftcities for both
the type of sialic acid and its linkage to
subterminal sugars. CD33 is specifically expressed in early myeloid and some
monocyte cell lineages and has been
shown to be strongly associated with various myeloid tumors including, for
example, acute non-lymphocytic leukemia
{ANLL). As such, CD33 has been suggested as a potential target for the
treatment of cancers associated with high
level expression of the protein. There is, therefore, significant interest in
the identification of novel polypeptides
having homology to CD33. In fact, one CD33 homolog (designated CD33L) has
already been identified and
described (see Takei et al., C~rtogenet. Cell Genet. 78:295-300 (1997)). We
herein describe the identification of
another novel polypeptide having homology to CD33, designated herein as
PR0940. The novel polypeptide described
herein also exhibits significant homology to the human OB binding proteins
designated HSU7I382'I and
HSU7I383_1 in the Dayhoff database (version 35.45 SwissProt 35).
41. P l
Cadherins are a large family of transmembrane proteins. Cadherins comprise a
family of calcium-dependent
glycoproteins that function in mediating cell-cell adhesion in virtually alt
solid tissues of multicellular organisms.
At least cadherins 1-13 as well as types B, E, EP, M, N, P and R have been
identified and characterized. Among
the functions cadherins are known for, with some exceptions, are that
cadherins participate in cell aggregation and
are associated with cell-cell adhesion sites. Recently, it has been reported
that while all cadherins share multiple
repeats of a cadherin specific motif believed to correspond to folding of
extracellular domains, members of the
cadherin superfamily have divergent strictures and, possibly, functions. In
particular it has been reported that
members of the cadherin superfamiiy are involved in signal transduction. See;
Suzuki, J. Cell Biochem., b1(4):531-
542 {1996). Cadtterins are further described in Tanihara et al., Dell Sci.,
10?(6}:1697-1704 (1994), Aberle et al.,
J. Cell Biochem., 61(4}:514-523 (1996) and Tanihara et al., Cell Adhes,
Commun., 2(I):15-26 (I994). We herein
describe the identification and characterization of a novel polypeptide having
homology to a cadherin protein,
designated herein as PR0941.
42. PRn944
Clostridium perfringens entemtoxin (CPE) is considered to be the virulence
factor responsible far causing
the symptoms of C. perfringens type A food poisoning and may also be involved
in other human and veterinary
illnesses (McClane, Toadcon. 34:1335-1343 (1996)). CPE carries out its adverse
cellular functions by binding to an
approximately 50 kD celt surface r~eptor protein designated the Clostridium
perfringens enterotoxin receptor (CPE-
R) to form an approxin~ely 90,000 kD complex on the surface of the cell. cDNAs
encoding the CPE-R protein have
been identified characterized in both human and mouse (Katahira et aL, J. Cell
Biol. 136:1239-1247 (1997) and
Kata>ztra et al., J. Biol. Chem. 272:26652-26658 (1997)). Since the CPE toxin
has been reported to cause a variety
of iihtesses in mammalian hosts arid those illnesses are initiated by binding
of the CPE toxin to the CPE-R, there is
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significant interest in identifying novel CPE-R homologs. We herein describe
the identification and characterization
of a novel polypeptide having homology to the CPE-R, designated herein as
PR0944.
43. PR 83
Membrane-bound proteins include not only cell-surface membrane-bound proteins,
but also proteins that are
found on the surface of intracellular vesicles. These vesicles are involved in
exocytosis, which is the fusion of _
secretory vesicles with the cellular plasma membrane, and have two main
functions. One is the discharge of the
vesicle contents into the extraceilular space, and the second is the
incorporation of new proteins and lipids into the
plasma membrane itself. Exocytosis can be either constitutive or regulated.
All eukaryotic cells exhibit constitutive
exocytosis, which is marked by the immediate fusion of the secretory vesicle
after formation. In contrast, regulated
exocytosis results in the accumulation of the secretory vesicles that fuse
with the plasma rrzembrane upon receipt of
an appropriate signal by vesicle-associated membrane proteins. Usually, this
signal is an increase in the cytosolic
free Ca2'' concentration. However, regulated exocytosis that is independent of
Cap' has been reported (see, e.g.
Fujita-Yoshigaki et al. J. Blot. Chem. (1996) 31:271(22):13130-13134).
Regulated exocytosis is crucial to many
specialized cells, including neurons (neurotransmitter release from synaptic
vesicles), adrenal chromaffm cells
(adrenaline secretion); pancreatic acinar cells (digestive enzyme secretion),
pancreatic ~i-cells (insulin secretion), mast
cells (histamine secretion), mammary cells (milk protein secretion), sperm
(enzyme secretion), egg cells (creation
of fertilization envelope) and adipocyces (insertion of glucose transporters
into the plasma membrane)
Disorders involving exocytosis are known. For example, inflammatory mediator
release from mast cells
leads to a variety of disorders, including asthma. Similarly, Chediak-Higashi
Syndrome (CHS) is a rare autosomal
recessive disease in which neutrophils, monocytes and lymphocytes contain
giant cytoplasmic granules. Accordingly,
the proteins involved in exocytosis are of paramount interest and efforts are
being undertaken by both industry and
academia to identify new, vesicle-associated proteins. For example, Skehel et
al, identified a 33-kilodalton membrane
protein in Aplysia, termed VAP-33, which is required for the exocytosis of
neurotransmitter. c'e ce (1995)
15:269(5230):1580-1583, and NeuronharmacoloQV (1995) 34(11}:1379-1385. Many
efforts are focused on the
screening of mammalian recombinant DNA libraries to identify the coding
sequences for novel vesicle- associated
membrane proteins. It is an object of the invention to provide proteins having
homology to the vesicle associated
protein, VAP-33, designated herein as PR0983.
44. PROlU.~'7
Proteases are enzymatic proteins which are involved in a large number of very
important biological
processes in mammalian and non-mammalian organisms. Numerous different
protease enzymes from a variety of
different mammalian and non-marritnalian organisms have been both identified
and characterized. The mammalian
protease enzymes play important roles in many different biological processes
including, for example, protein
digestion, activation, inactivation, or modulation of peptide hormone
activity, and alteration of the physical properties
of proteins and enzymes.
In light of the important physiological roles played by protease enzymes,
efforts are currently being
undertaken by both industry and academia to identify new, native protease
homologs. Many of these efforts are
focused on the screening of mattttnalian recombinant DNA libraries to identify
the coding sequences for novel
secreted proteins. Examples of screening methods and techniques are described
in the literature [see, for example,
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Klein et al., Proc. Natl. Acad" Sci., 9,x:7108-7113 (1996); U.S. Patent No.
5,536,637)]. We herein describe the
identification of novel polypeptides having homology to various protease
enzymes, designated herein as PR01057
polypeptides.
45. P 01 71
Thrombospondin-1 is a trimeric high molecular weight glycoprotein that is
released from platelet alpha-
granules in response to thrombin stimulation and that is also a transient
component of the extracellular matrix in
developing acid repairing tissues (Adams, Int. 1. B]o~hem. CeII Biol. 29:861-
865 (1997) and Qian et al., ro .
E~cp. Biol. Mec~ 212:199 207 (1996)). A variety of factors regulate
thrombospondin expression and the protein is
degraded by both extracellular and intracellular routes. Thrombospondin-1
functions as a cell adhesion molecule and
also modulates cell movement, cell proliferation, neurite outgrowth and
angiogenesis. As such, there is substantial
interest in identifying novel polypeptides having homology to thrombospondin.
We herein describe the identification
and characterization of a novel polypeptide having homology to thrombospondin,
designated herein as PR0107I.
46. R 072
Studies have reported that the redox state of the cell is an important
determinant of the fate of the cell.
Furthermore, reactive oxygen species have been reported to be cytotoxic,
causing inflammatory disease, including
tissue necrosis, organ failure, atherosclerosis, infertility, birth defects,
premature aging, mutations and malignancy.
Thus, the control of oxidation and reduction is important for a number of
reasons, including the control and
prevention of strokes, heart attacks, oxidative stress, hypertension and may
be associated with the development of
malignancies. The levels of antioxidant enzymes, such as reductases, which
catalyze the conversion of reactive
oxygen species to water have been shown to be low in cancer cells. In
particular, malignant piostate epithelium may
have lowered expression of such antioxidant enzymes [Baker et al., Prostate
32(4):229-233 (1997)]. In this regard,
reductases, are of interest. In addition, the transcription factors, NF-kappa
B and AP-1, are known to be regulated
by redox state and to affect the expression of a large variety of genes
thought to be involved in the pathogenesis of
AIDS, cancer, atherosclerosis and diabetic complications. Publications further
describing this subject matter include
Engrnatt et al., ticancer Res. ~, 17:4599-4605 (1997), Kelsey, et al., Br, J.
Cancer, 76(7):852-854 (1997);
Friedrich and Weiss, J. Theor,,Biol., 1f7(4):529-40 (1997) and Pieulle, et
al., ~, B~cteriol., 179(18):5684-92 (1997).
Given the physiological importance of redox~reactions in vivo, efforts are
currently being under taken to identify new,
native proteins which are involved in redox reactions. We describe herein the
identification of a novel polypeptide
which has sequence similarity to reductase enzymes, desiignated herein as
.PR01072.
47. 2 '
Protein disulfide isomerase is an enzymatic protein which is involved in the
promotion of correct refolding
of proteins through the establishment of correct disulfide bond formation.
Protein disulfide isomerase was initially
identified l~sed upon its ability to catalyze the renaturation of reduced
denatured RNAse (Goldberger et al., J. Biol.
Chem. 239:1406-1410 (1964) and Epstein et al., Cold Sprring Harbor Symp.
Quart. Biol. 28:439-449 (1963)).
Protein disulfide isomerase has been shown to be a resident enzyme of the
endoplasmic reticulum which is retained
in the endoplasmic reticulutn via a -KDEL or -H1DEL amino acid sequence at its
C-terminus.
Given the importance of disulfide bond-forming enzymes and their potential
uses in a number of different
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applications, for example in increasing the yield of correct refolding of
recombinantly produced proteins, efforts are
currently being undertaken by both industry and academia to identify new,
native proteins having homology to protein
disulfide isomerase. Many of these efforts are focused on the screening of
mammalian recombinant DNA libraries
to identify the coding sequences for novel protein disulfide isomerase
homologs. Examples of screening methods and
techniques are described in the literature [see, for example, Klein et al.,
Proc. Nat!. Acad. Sri., 93:7I08-71 I3 (1996);
U.S. Patent No. 5,536,637)]. We herein describe a novel polypeptide having
homology to protein disulfide
isomerase, designated herein as PR01075.
48. P 0181
In Drosophila, the dorsal-ventral polarity of the egg chamber depends on the
localization of the oocyte
nucleus and the gurken RNA to the dorsal-anterior corner of the oocyte. Gurken
protein presumably acts as a Iigand
for the drosophila EGF receptor (torpedo/DER) expressed in the somatic
follicle cells surrounding the oocyte.
Cornichon is a gene required in the gertnline for dorsal-ventral signaling
~(Roth et al., CeI 81:967-978 (1995)).
Cornichon, gurken and torpedo also function in an earlier signaling event that
establishes posterior follicle cell fates
and specifies the anterior-posterior polarity of the egg chamber. Mutations in
any or all of these genes prevent the
i5 formation of a correctly polarized microtubule cytoskeleton required for
proper localization of the anterior and
posterior determinants bicoid and oskar and for the asymmetric positioning of
the oocyte nucleus. Thus, it is clear
that the cornichon gene product plays an important role in early development.
We herein describe the identification
and characterization of a novel polypeptide having homology to the cornichon
protein, designated herein as PR0181.
49. PR
Efforts acre currently being undertaken to identify and characterize novel
uansmembrane proteins. We
herein describe the identification and characterization of a novel
transmembrane polypeptide, designated herein as
PRO195.
50. PR 5
Efforts arre currently being undertaken to identify and characterize novel
secreted proteins. We herein
descn'he the identification and characterization of a novel secreted
polypeptide, designated herein as PR0865.
51. ~R0827
VLA-2 is an cell-surface integrin protein that has been identified and
characterized in a number of
mammalian organisms, including both mouse and human. VLA-2 has been shown to
be a receptor on the surface
of cells for echovirus-1 (EV-1) which mediates infection of VLA-2-expressing
cells by EV-1 (Zhang et al.; ViroloQv
235(2)293-301 (1997) and Bergelson et al., '~ 255:1718-1720 (I992)). VLA-2 has
also been shown to mediate
the interaction of collagen with endothelium during in vitro vascular tube
formation (Jackson et al., ~el~,Biol.~int.
18(9):859-867 (1994)). Various other integrin proteins that share various
degrees of amino acid sequence homology
with VLA-2 have been identified and characterized in a variety of mammalian
organism. These integrins have been
reported to play important roles in a variety of differern physiological
functions. Therefore, there is significant
interest in identifying novel polypeptides having homology to o~ or more of
the integrin proteins. We herein
describe the identification and characterization of a novel polypeptide having
homology to VLA-2 integrin protein,
22
CA 02421319 2003-03-25
WO 9914b281 PCT/US99I05028
designated herein as PR0827.
52. P 114
Many important cytokine proteins have been identified and characterized and
shown to signal through
specific cell surface receptor complexes. For example, the class II cytokine
receptor family (CRF2) includes the
interferon receptors, the interleukin-ZO receptor and the tissue factor CRFB4
(Spencer et al., J. Exp. Med. 187:571-
578 (1998) and Kotenko et al., ~MBO J. 16:5894-5903 (1997)). Thus, the
multitude of biological activities exhibited
by the various cytoldne proteins is absolutely dependent upon the presence of
cytokine receptor proteins on the
surface of target cells. There is, therefore, a significant interest in
identifying and characterizing novel polypeptides
having homology to one or more of the cytokine receptor family. We herein
describe the identification and
characterization of a novel polypeptide having homology to cytotcine receptor
family-4 proteins; designated herein
as PR01117.
Irtoerferons {IFNs) encompass a large family of secreted proteins occurring in
vertebrates. Although they
were originally named for their antivirai activity, growing evidence supports
a critical role for IFNs in cell growth
and differentiation (laramillo et al., ,Cancer IrJvPS~t'gation 13(3):327-338
(1995)). IFNs belong to a class of negative
IS growth factors having the ability to inhibit the growth of a wide variety
of cells with both normal and transformed
phe~types. IFN therapy has been shown to be beneficial in the treatment of
human malignancies such as Karposi's
sarcoma, chronic myelogenous leukemia, non-Hodgkin's lymphoma, and hairy cell
leukemia as well as in the
treatment of infectious diseases such as hepatitis B (Gamliel et al., Scgnninp
Microseoov 2(1):485-492 (1988),
Einhorn et aL, MediOnco~,,, ~Tr ,P~rm~o.~r. 10:25 29 (1993), Ringenberg et
al., isso~i Medicine
85(1):21-26 (1988), Saracco et al., Journal of Gasttoenterologlr~nd ~Ie-
patoioev 10:668-673 (1995), Gonzalez-Mateos
et al., Hepato-Gastroe~erologv 42:893-899 (1995) and Malaguamera et al:,
Plgsr~n,~cotheranv 17(5):998-1005
(1997)).
Interferons can be classified into two major groups based upon their primary
sequence. Type I interferons,
IFN-a and 1FN-(3, are encoded by a superfamily of intronless genes consisting
of the IFN-ac gene family and a single
IFN-~ gene that are thought to have arisen from a common ancestral gene. Type
I interferons tray be produced by
most cell types. Type II IFN, or IFN-y, is restricted to lymphocytes (T cells
and natural ldIler cells) and is stimulated
by nonspecific T cell activators or specific antigens in vivo.
Although both type I and type ll IFNs produce similar antiviral and
anuproliferative effects, they act on
disti~t cell surface receptors, wherein the binding is get~rally species
specific (l.anger et al., Immttttol. Todav 9:393-
400 (1988}). Both IFN-ac and IFN-~i bind~competitively to the same high
affinity type I receptor, whereas IFN-y
binds to a distinct type ll receptor. The presence and number of IFN receptors
on the surface of a cell does not
ge~rally reflect the sensitivity of the cell to IFN, although it is clear that
the effects of the IFN protein is mediated
through binding to a cell surface interferon receptor. As such, the
identification and characterization of novel
interferon receptor proteins is of extreme interest.
We herein describe the identification anrl characterization of novel
interferon receptor polypeptides,
designated herein as "PR01114 interferon receptor" polypeptides. Thus, the
PR01114 polypeptides of the present
invention represents a novel cell surface interferon receptor
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53. 0237
Carbonic anhydrase is an enzymatic protein that which aids carbon dioxide
transport and release in the
mammalian blood system by catalyzing the synthesis (and the dehydration) of
carbonic acid from (and to) carbon
dioxide and water. Thus, the actions of carbonic anhydrase are essential for a
variety of important physiological
reactions in the mammal. As such, there is significant interest in the
identification and characterization of novel
polypeptides having homology to carbonic anhydrase. We herein describe the
identification and characterization of
a novel polypeptide having homology to carbonic anhydrase> designated herein
as PR0237.
54. PR 1
Numerous trypsin inhibitory proteins have been identified and characterized
(see, e.g., Yamakawa et al.,
l3iochim. Biophvs. Acta 1395:202-208 (1998) and Mizuki et al., Mammalian
Genome 3:274-280 (1992)). Trypsin
inhibitor proteins play important roles in a variety of different
physiological and biological pathways and are
specifically involved in such processes as the regulation of protein
degradation, digestion, and the like. Given the
important roles played by such enzymatic proteins, there is significant
interest in identifying and characterizing novel
polypeptides having homology to known trypsin inhibitor proteins. We herein
describe the identification and
characterisation of a novel poIygeptide having homology to a trypsin inhibitor
protein, designated herein as PR0541.
55, PR 7
Ixukocytes include monocytes, macrophages, basophils, and eosinophils and play
an important role in the
immune response. These cells are important in the mechanisms initiated by T
and/or B lymphocytes and secrete a
range of cytokines which recruit and activate other inflammatory ceps and
contribute to tissue destruction.
Thus, investigation of the regulatory processes by which leukocytes move to
their appropriate destination
and interact with other cells is critical. Currently, leukocytes are thought
to move from the blood to injured or
inflamed tissues by rolling along the endothelial cells of the blood vessel
wall. This movement is mediated by
transient interactions between selectins and their ligands. Next, the
leukocyte must move through the vessel wall and
into the tissues. This diapedesis and extravasation step involves cell
activation which promotes a more stable
leukocyte-endothelial cell interaction, again mediated by integrins and their
ligands.
Chemokines area large family of stntcturally related polypeptide cytokines.
These molecules stimulate
leukocyte movement and may explain leukocyte uatftcinng in different
inflammatory situations. Chemokines mediate
the expression of particular adhesion molecules on endothelial cells, and they
produce chemoattractants which activate
specific cell types. In addition, the chemokines stimulate proliferation and
regulate activation of specific cell types.
In both of these activities, chemokines demonstrate a high degree of target
cell specificity.
The chemoldne family is divided into two subfamilies based on whether two
amino terminal cysteine residues
are immediately adjacent (C-C) or separated by one amino acid (C X-C):
Chemoldrtes of the C-X-C family generally
activate neutrophils and fibroblasts while the C-C chemokines act on a more
diverse group of target cells including
monocytes/macrophages, basophils, eosinophils and T lymphocytes. The known
chemokines of both subfamilies are
synthesized by many diverse cell types as reviewed in Thomson A. (1994) The
Cytokine Handbook, 2 d Ed.
Academic Press, N.Y. Chemokines are also reviewed in Schall TJ (1994)
Chemotactic Cytokines: Targets for
Therapeutic Development. International Business Communications, Southborough
Mass: pp 180-270; and in Paul
WE (1993) Fundamental Immunology, 3rd Ed. Raven Press, N.Y. pp 822-826.
24
CA 02421319 2003-03-25
WO 99146281 PCTlUS99/05028
Known chemokines of the C-X-C subfamily include macrophage inflammatory
proteins alpha and beta
(MIP-I and MIP-2 ), interleukin-8 (IL-8), and growth regulated protein (GRO-
alpha and beta).
MIP-2 was first identified as a 6 kDa heparin binding protein secreted by the
mouse macrophage cell line
RAW 264.7 upon stimulation with lipopolysaccharide (LPS). MIP-2 is a member of
the C-X-C (or CXC) subfamily
of chemokines. Mouse MIP-2 is chemotactic for human ntutrophils and induces
local neutrophil infiltration when
injected into the foot pads of mice. Rat MIP-2 shows 86% amino acid homology
to the mouse MIP-2 and is
chemoractic for rat neutrophils but does not stimulate migration of rat
alveolar macrophages or human peripheral
blood eosinophils or lymphocytes. In addition, the rat MIP 2 has been shown to
stimulate proliferation of rat alveolar
epithelial cells but not fibrobtasts.
Current techniques for diagnosis of abnormalities in inflamed or diseased
issues mainly rely on observation
of clinical symptoms or serological analyses of body tissues or fluids for
hormones, polypeptides or various
metabolites. Problems exist with these diagnostic techniques. First, patients
may not manifest clinical symptoms at
early stages of disease. Second, serological tests do not always differentiate
between invasive diseases and genetic
syndromes. Thus, the identification of expressed chemokines is important to
the development of new diagnostic
techniques, effective therapies, and to aid in the understanding of molecular
pathogenesis.
To date, chemokines have been implicated in at least the following conditions:
psoriasis, inflammatory
bowel disease, renal disease, arthritis, immune-mediated aIopecia, stroke,
encephalitis, MS, hepatitis, and others.
In ion, non-ELR-containing chemokines have been implicated in the inhibition
of angiogenesis, thus indicating
that these chemokines have a rule in tumor vascularization and tutnorigenesis.
Therefore it is the object of this invention to identify polypeptides and
nucleic acids encoding the same which
have sequence identity and similarity with cytokine-induced neutrophil
chemaattractants, M1P-1, MIP-2, and other
related proteins. The efforts of this object are provided herein.
56. ; 80701
Beta neurexins and neuroligins are plasma membrane proteins that are displayed
on the neuronal cell surface.
Neuroligin 1 is ertric~d in synaptic plasma membranes and acts as a splice
site-specific ligand for beta neurexins as
descn'bed in Ichtchenko, et al., !'e , 81(3):435-443 (1995). The extracellular
sequence of neuroligin 1 is composed
of a catalytically inactive esterase domain homologous to
acetylcholinesterase. NeuroIigin 2 and 3 are similar in
structure and sequence to neuroligin 1. All neuroligins contain an N-terminal
hydrophobic sequence with the
cltatacteristics of a cleaved signal peptide followed by a large esterase
homology domain, a highly conserved single
traasmembrane region, and a short cytoplasmic domain. The three neuroligins
are alternatively spliced at the same
position and are expressed at high levels only in the brain. Tight binding of
the three neuroligins to beta neurexins
is observed oaly for beta neurexitts lacking an insert in splice site 4. Thus,
neuroligins constitute a multigene family
of brain-specific proteins with distinct isofotms that may have overlapping
functions in tttediating recognition
processes between neurons, see Ichtchenko, et al., J. Biol: Chem., 27I(5):2676-
2682 (1996). Moreover, neurexins
and neuroligins have been reported as functioning as adhesion molecules in a
Ca" dependent reaction that is regulated
by alternative splicing of beta neumacins, i.e., see Nguyen and Sudhof, J.
Bio~. Chem., 272(41):26032-26039 (1997).
Gives the foregoing, membrane bound proteins are of interest. More generally,
membrane-bound proteins
and receptors caa play an impoztaru role in the formation, differentiation and
maintenance of multicelhtlar organisms.
The fate of many individual cells, e.g., proliferation, migration,
differentiation, or interaction with other cells, is
CA 02421319 2003-03-25
WO 99/46281 PCT/US99/05028
typically governed by information received from other cells andlor the
immediate environment. This information is
often transmitted by secreted polypepfides {for instance, mitogenic factors,
survival factors, cytotoxic factors,
differentiation factors, neuropeptides, and hormones) which are, in turn,
received and interpreted by diverse cell
receptors or membrane-bound proteins. Such membrane-bound proteins and cell
receptors include, but are not limited
to, cytokine receptors, receptor kinases, receptor phosphatases, receptors
involved in cell-cell interactions, and
cellular adhesin molecules like selectins and integrins. For instance,
transduction of signals that regulate cell growth
and differentiation is regulated in part by phosphorylation of various
cellular proteins. Protein tyrosine kinases,
enzymes that catalyze that process, can also act as growth factor receptors.
Examples include fibroblast growth factor
receptor and nerve growth factor receptor.
Membrane-bound proteins and receptor molecules have various industrial
applications, inchtding as
pharmaceutical and diagnostic agents. Receptor immunoadhesins, for instance,
can be employed as dterapeutic agents
to block receptor-ligand interaction. The membrane-bound proteins can also be
employed for screening of potential
peptide or small molecule inhibitors of the relevant receptorlligand
interaction.
Efforts are being undertaken by both industry and academia to identify new,
native membrane-bound
receptor proteins, particularly those having sequence identity and/or
similarity with neuroligins 1, 2 and 3. Many
efforts are focused on the screening of mammalian recombinant DNA libraries to
identify the coding sequences for
novel secreted and membrane-bound receptor proteins. Examples of screening
methods and techniques are described
in the literature [see, for example, Klein et al., Proc. Nato, Acad. Sci.,
Q3_:7108-7113 (1996); U.S. Patent No.
5,536,037)]. The results of such efforts are provided herein.
57. PRO'704
VIP36 is localized to the Golgi apparatus and the cell surface, and belongs to
a family of legume Iectin
homologues in the animal secretory pathway that might be involved in the
trafficking of glycopmteins, glycolipids,
or both. It is fiu~ther believed that V1P36 binds to sugar residues of
glycosphingolipids andlor gycosylphosphatidyl-
inositol anchors and might provide a Link between the extracellularlluminal
face of glycolipid rafts and the cytoplasmic
protein segregation machinery. Further regarding VIP36, it is believed that
there is a signal at its C-terminus that
matches an internalization consensus sequence which confers its ability to
cycle between the plasma membrane and
Golgi. See, Fiedler, et al, EMBO J., 13(7):1729-1740 (1994); Fiedler and
Simons, . ci., 109(1):2?1-27b
(199; Itin, et al:, MB0 33., 14(10):2250-2256 (1995). ~ It is believed that
VIP36 is either the same as or very closely
related to the human GP36b protein. VIP36 andlor GP36b are of irnerest.
More generally, vesicular, cytoplasmic, extracellular and membrane-bound
proteins play important roles
in the formation, differentiation and maintenance of multicellular ozganisms.
The fate of many individual cells, e.g.,
proliferation, migration, differentiation, or interaction with other cells, is
typically governed by information received
from other cells andlor the immediate environment. This information is often
transmitted by secreted poiypeptides
(for instance, mitogenic factors, survival factors, cytatoxic factors,
differentiation factors, neuropeptides, and
hormones) which are, in rum, received and interpreted by diverse cell
receptors or membrane-bound proteins. These
secreted polypeptides or signaling molecules normally pass through the
cellular secretory pathway to reach their site
of action in the exuaceliular envirotttnent, usually at a membrane-bound
receptor protein.
Secreted proteins have various industrial applications, including use as
pharmaceuticals, diagnostics,
biasensors and bioreactors. In fact, most protein drugs available at present,
such as thrombolytic agents, interferons,
26
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WO 99/46281 PCTIUS99105028
interleukins, erythropoietins, colony stimulating factors, and various other
cytokines. are secretory proteins. Their
receptors, which are membrane-bound proteins, also have potential as
therapeutic or diagnostic agents. Receptor
immunoadhesins, for instance, can be employed as therapeutic agents to block
receptor-Iigand interaction.
Membrane-bound proteins can also be employed for screening of potential
peptide or small molecule inhibitors of
the relevant receptor/ligand interaction. Such membrane bound proteins and
cell receptors include, but are not limited
to, cytokine receptors, receptor kinases, receptor phosphatases, receptors
involved in cell-cell interactions, and
cellular adhesin molecules like selectins and integrins. Transduction of
signals that regulate cell growth and
differentiation is regulated in part by phosphorylation of various cellular
proteins. Protein tyrosine kinases, enzymes
that catalyze that process, can also act as growth factor receptors. Examples
include fibroblast growth factor receptor
and nerve growth factor receptor.
Efforts are being undertaken by both industry and academia to identify new,
native vesicular, cytoplasmic,
secreted and membrane-bound receptor proteins, particularly those having
sequence identity andlor similarity with
V1P36. Many efforts ate focused on the screening of matnmaiian recombinant
I~NA libraries to identify the coding
sequences for novel secreoed and membrane-bound receptor proteins. Examples of
screening methods and techniques
are described in the literature [see, for example, Klein et at., Proc.,~T~:l.
Acad. Ski"" 9:7108-7113 (1996); U.S.
Patent No. 5,536,637)].
58. PR0706
Arid phophatase proteins are secreted proteins which dephophorylate terminal
phosphate groups under acidic
pIi conditions. Acid phophatases contain a RI-IGXRXP amino acid sequence,
which is predicted to be mechanistically
sigt>ificant. Acid phosphatases may have important functions in the diagnosis
and treatment of human diseases. For
example, prostatic acid phosphatase is a secreted protein uniquely expressed
in prostatic tissue and prostate cancer.
The level of prostatic acid phosphatase is a potential prognostic factor for
local and biochemical control in prostate
cancer patients treated with radiotherapy, as described in LanlCford et al.,
int. J. Radiat. tZrtcol. Biol.~ P 3$
hJ'~. (2):
327-333 (1997). Research suggests that a cellular immune response to prostatic
acid phosphatase may mediate
destructive autoimmune prostatitis, and that xenogeneic forms of prostatic
acid phosphatase may prove useful for
im~notherapy of prostate cancer. See Fong et al.; J.J. Im unol. 169(7): 3113-
3I I7 (1997). Seminal prostatic acid
phospbatase levels correlate significantly with very low sperm levels
(oligospernua) in individuals over 35, see Singh
et aL, ,, ingapore Med. 1. 37(b): 598-599 (1996). Thus, prostatic acid
phosphatase has been implicated in a variety
of human diseases, and may have an important function in diagnosis and therapy
of these diseases. A series of
aminobenzylphosphatic acid compounds are highly potent inhibitors of prostatic
acid phosphatase, as described in
Beers et al., Bioorg_ Mcd. Ghern. 4(10): 1693-1701 (1996).
More generally, extracellnlar proteins play an important role in the
formation, differentiation and
maintenance of multicellular organisms. The fate of many individual cells,
e.g., proliferation, migration,
differentiation, or interaction with other cells, is typically governed by
information received from other cells and/or
the immediate environment. This information is often transmitted by secreted
poiypeptides (for instance, mitogenic
factors, survival factors, cywtoxic factors, differentiation factors,
neuropeptides, and hormones) which are, in turn,
received and interpreted by diverse cell receptors or membrane-bound proteins.
These secreted polypeptides or
signaling molecules normally pass through the cellular secretory pathway to
reach their site of action in the
extracelhilar envirantnent.
27
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WO 99/46281 PCT/US99/05028
Secreted proteins have various industrial applications, including
pharmaceuticals, diagnostics, biosensors
and bioreactors. Most protein drugs available at present, such as thrombolytic
agents, interferons, interleulcins,
erythropoietins, colony stimulating factors, and various other cytolcines, are
secretory proteins. Their receptors,
which are membrane proteins, also have potential as therapeutic or diagnostic
agents. Efforts are being undertaken
by both industry and academia to identify new, native secreted proteins,
particularly those having sequence identity
with prostate acid phosphatase precursor and lysosomaI acid phosphatase
precursor and in some cases, those having
identity with DNA found in fetal heart. Many efforts are focused on the
screening of mammalian recombinant DNA
libraries to identify the coding sequences for novel secreted proteins,
Examples of screening methods and techniques
are described in the literature [see, for example, Klein et al., Proc. Natl.
Acad. Sci., 93:7108-71 I3 (1996); U.S.
Patent No. 5,536,637)].
59. PRa707
Cadherins are a large family of transmembrane proteins. At least cadherins 1-
13 as well as types B, E, EP,
M, N, P and R have been characterized. Among the functions cadherins are known
for, with some exceptions,
cadherins participate in cell aggregation and are associated with cell-cell
adhesion sites. Cadherins are further
described in Tanihara, et al., J. Cell Sci., 107(6):1697-I704 (1994) and
Tanihara, et al., Cell Adhes. Cotnmun.,
2{1):15-26 (1994). Moreover, it has been reported that some members of the
cadherin superfamily are involved in
general cell-cell interaction processes including transduction. See,
Suz:uki,1. Cell Biochem., 61(4):531-542 (1996).
Therefore, novel members of the cadherin superfamily are of interest.
More generally, all novel proteins are of interest, including membrane-bound
proteins. Membrane-bound
pzoteins and receptors can play an important role in the formation,
differentiation and maintenance of muhicellular
organisms. The fate of many individual cells, e.g., proliferation, migration,
differentiation, or interaction with other
cells, is typically governed by information received from other cells and/or
the immediate environment. This
infonration is often transmitted by secreted polypeptides (for instance,
mitogenic factors, survival factors, cytotoxic
factors, differentiation factors, neuropeptides, and hormones) which are, in
turn, received and interpreted by diverse
cell receptors or membrane-bound proteins. Such membrane-bound proteins and
cell receptors include, but are not
limited to, cytokine receptors, receptor kinases, receptor phosphatases,
receptors involved in cell-cell interactions,
and cellular adhesin molecules like selectins and integrins. For instance,
transduction of signals that regulate cell
growth and differentiation is regulated in part by phosphorylation of various
cellular proteins. Protein tyrosine
kittases, enzymes that catalyze that process, can also act as growth factor
receptors. Examples include fibroblast
growth factor receptor and nerve growth factor receptor.
Membrane-bound proteins and receptor molecules have various industrial
applications, including as
pharmaceutical and diagnostic agents. Receptor immunoadhesins, for instance,
can be employed as therapeutic agents
to block receptor-ligand interaction. The membrane-bound proteins can also be
employed for screening of potential
peptide or small molecule inhibitors of the relevant receptorlligand
interaction.
Efforts are being ut~ertaken by both itulustry and academia to identify new,
native secreted and membrane-
bound receptor proteins, particularly membrane bound proteins having identity
with cadherins. The results of such
efforts are provided herein.
28
CA 02421319 2003-03-25
WO 99!46281 PC1'/US99105028
60. PR0322
Proteases are enzymatic proteins which are involved in a large number of very
important biological
processes in mamrnaIian and non-mammalian organisms. Numerous different
protease enzymes from a variety of
different marr>rnalian and non-marmnalian organisms have been both identified
and characterized, including the serine
proreases which exhibit specific activity toward various serine-containing
proteins. The mammalian protease enzymes
play important roles in biological processes such as, for example, protein
digestion, activation, inactivation, or
modulation of peptide hormone activity, and alteration of the physical
properties of proteins and enzymes.
Neuropsin is a novel serine protease whose mRNA is expressed in the central
nervous system. Mouse
neuropsin has been cloned, and studies have shown that it is involved in the
hippocampal plasticity. Neuropsin has
also been indicated as associated with extracellular matrix modifications and
cell migrations. See, generally, Chen,
et al., a osci., 7(2):5088-5097 (1995) and Chen, et al., J. H,;'tstochem.
Cvtochem., 46:313-320 (1998).
Efforts are being undertaken by both industry and academia to identify new,
native membrane-bound or
secreted proteins, particularly those having homology to neuropsin, serine
protease, neurosin and trypsinogen. Marry
efforts are focused on the screening of marnr;>alian recombinant DNA libraries
to identify the coding sequences for
novel secreted and membra~-bound receptor proteins. Examples of screening
methods and techniques are described
IS in the literature [see, for example, Klein et al., Proc. Nat!. Acad. Sci.,
Q~:7108-7113 (1996); U.S. Patent No.
5,S36,637)).
61. PR0526
Protein protein interactions include those involved with receptor and antigen
complexes and signaling
mechanisms. As more is known about the structural and functional mechanisms
underlying protein-protein
interactions, protein-protein interactions can be more easily manipulated to
regulate the particular result of the protein-
protein interaction. Thus, the underlying mechanisms of protein-protein
interactions are of interest to the scientific
and medical community.
All proteins containing Ieucine-rich repeats are thought to be involved in
protein-protein interactions.
heucine-rich repeats are short sequence motifs present in a ntunber of
proteins with diverse functions and cellular
locations. The crystal structure of ribonuclease inhibitor protein has
revealed that leucine-rich repeats correspond
to beta-alpha structural units. These units are arranged so that they form a
parallel beta-sheet with one surface
exposed to solvent, so that the protein acquires an unusual, nonglobuiar
shape. These two features have been
indicated as responsible for the protein-binding functions of proteins
containing Ieucine-rich repeats. See, Kobe and
Deisenhofer, TreO~ds Biochem. Sci., 19(1.0):415-421 (Oct. 1994).
A study has begirt reported on leucine-rich proteoglycans which serve as
tissue organizers, orienting and
ordering collagen fibrils during ontogeny and are involved in pathological
processes such as.wound healing, tissue
repair, and tumor stroma formation. Iozzo, R. V:, Crit. Rev: Bloc em.,~pj.
BioL, 32(2):141-174 (1997). Others
studies implicating Ieucine rich proteins in wound healing and tissue repair
are De La Salle, C., et al., Vouv. Rev:
Fr. Hematol: (), 37(4)215-222 (I995), reporting mutations in the leucine rich
motif in a complex associated
with the bleeding disorder Bernard-Sootier syndrome, Chlemetson, K. J.,
~~ombiHaemost: (Germany), 74(1):111-
i l6 (7uly 1995), reporting that platelets have Ieucine rich repeats and
Ruosiahti, E. L, et al., W09110727-A by La
Jo1)a Cancer Research Foutrdation reporting that decorin bimling to
transforming gmwth factor~i has involvement in
a treatment for cancer, wound healing and scarring. Related by function to
this group of proteins is the insulin like
29
CA 02421319 2003-03-25
WO 99/46281 PCT/US99l05028
growth factor (IGF), in that it is useful in wound-healing and associated
therapies concerned with re-growth of tissue,
such as connective tissue, skin and bone; in promoting body growth in humans
and animals; and in stimulating other
growth-related processes. The acid labile subunit (ALS) of IGF is also of
interest in that it increases the half life of
IGF and is pan of the IGF complex inin vivo. ALS is further described in Leong
and Baxter, Mol. Endocrinol.,
6(6):870-876 (1992); Baxter, J. Biol. Chem., 264(20}:11843-11848 (1989}: and
Khosravi, et al., J. Ciin. Er~docrinol.
M~tab., 82(12):3944-3951 (I997).
Another protein which has been reported to have leucine-rich repeats is the
SLIT protein which has been
reported to be useful in treating neuro-degenerative diseases such as
Alzheimer's disease, nerve damage such as in
Parkinson's disease, and for diagnosis of cancer, see, Artavanistsakonas, S,
and Rothberg, J. M., W09210518-A1
by Yale University. Also of interest is LIG-1, a membrane glycoprotein that is
expressed specifically in glial cells
in the mouse brain, and has Ieucine rich repeats and immunoglobulin-like
domains. Suzuki, et al., I. Bioli Chem.
(U.S.), 271(37}:22522 (1996). Other studies reporting on the biological
functions of proteins having ieucine rich
repeats include: Tayar, N., et al., Mol. Cell EndocririoL; (Ireland), 125(1-
2):65-70 (Dec. 1996) (gonadotropin
receptor involvement); Miura, Y., et al., ~lippon Rinsho (Japan), 54(7):1784-
1789 (July 1996) (apoptosis
involvement); Harris, P. C., et al., J. Am. Soc.,~Tephrol., 6(4):1125-1133
(Oct. 1995) (kidney disease involvement).
I S Efforts are therefore being undertaken by both industry and academia to
identify new proteins having Ieucine
rich repeats to better understand protein-protein interactions. Of particular
interest are those proteins having leucine
rich repeats and identity or similarity to known proteins having leucine rich
repeats such as ALS: Many efforts are
focused on the screening of mammalian recombinant DNA libraries to identify
the coding sequences for novel
secreted and membrane bound proteins having leucine rich repeats. Examples of
screening methods and techniques
are described in the literature [see, for example, HIein et al., )'roc. Natl.
Acad. Sci., x:7108-7113 (I996); U.S.
Patent No. 5.536,637)].
62. P~531
Cadherins are a large family of ttansmembrane proteins. Cadherins comprise a
family of calcium-dependent
glycoproteins that function in mediating cell-cell adhesion in virtually ail
solid tissues of multicellular organisms.
At least cadherins 1-13 as well as types B, E, EP, M, N, P and R have been
characterized. Among the functions
cadherins are known for, with some exceptions, cadherins participate in cell
aggregation and are associated with cell-
cell adhesion sites. Recently, it has been reported that while all cadherins
share multiple repeats of a cadherin
specific ~tif believed to correspond to folding of exuaceliular domains,
members of the cadherin superfamily have
divergent structures and, possibly, functions. In particular it has been
reported that members of the cadherin
superfatnily are involved in signal uansduction. See, Suzttla, ~. CeII
Biochem., 61(4):531-542 (1996). Cadherins
are further described in Tanihara, et al., ~ Cgll Sci., 107(6):1697-1704
(1994), Aberle, et al., J. Cep ioche .,
61(4):514-523 (1996) and Tanihara, et al., Cell Adhes. Corrmtun., 2(1):15-26
{1994).
Protocadherins are members of the cadherin superfamily which are highly
expressed in the brain. In some
3~ studies, protocadherins have shown cell adhesion activity. See, Sano, et
al., EMBO J., 12(6):2249-2256 (1993).
However, studies have also shown that some protocadherins, such as
protocadherin 3 (also referred to as Pcdb3 or
pc3), .do not show strong calcium dependent cell aggregation activity. See,
Sago, et al., Genomics, 29(3):631-640
(1995) for this study and further characteristics of Pcd63.
CA 02421319 2003-03-25
WO 99!46281 1'CTIUS99I05028
Therefore, novel members of the cadherin superfamily are of interest. More
generally, all membrane-bound
proteins and receptors are of interest: Such proteins can play an important
role in the formation, differentiation and
maintenance of multicellular organisms. The fate of many individual cells,
e.g., proliferation, migration,
differentiation, or interaction with other cells, is typically governed by
information received from other cells andlor
the immediate environment. This information is often transmitted by secreted
polypeptides (for instance, mitogenic
factors, survival factors, cytotoxic factors, differentiation factors,
neuropeptides, and hormones) which are, in turn,
ttceived and interpreted by diverse cell receptors or membrane-bound proteins.
Such membrane-bound proteins and
cell receptors irxlude, but are not limited to, cytokine receptors, receptor
kinases, receptor phosphatases, receptors
involved in cell-cell interactions, and cellular adhesin molecules like
selectins and integrins. For instance,
transduction of signals that regulate cell growth and differentiation is
regulated in part by phosphorylation of various
cellular proteins. Protein tyrosine kinases, enzymes that catalyze that
process, can also act as growth factoz
receptors. Examples include fibroblast growth factor receptor and nerve growth
factor receptor.
Membrane-bound proteins and receptor molecules have various industrial
applications, inchtding as
pharmaceutical and diagnostic agems. Receptor imttruttoadhesins, for instance,
can be employed as therapeutic agents
to block receptor-ligand interaction. The membrane-bound proteins can also be
employed for screening of potential
peptide or small molecule inhibitors of the relevant receptorlligand
interaction.
Efforts are therefore being undertaken by both industry and academia to
identify new, native membrane
bound proteins, particular those having sequence identity with protocadherins,
especially 3 and 4. Many efforts are
focused on the screening of mammalian recombinant DNA libraries to identify
the coding sequences for novel
membrane-bound proteins. Provided herein are the results of such efforts.
63. PR0534
Protein disulfide isomerase is an enzymatic protein which is involved in the
promotion of correct refolding
of proteins through the establishment of correct disulfide bond fot~tion.
Protein disulfide isomerase was initially
fled based upon its ability to catalyze the renaturation of reduced denatured
RNAse (Goldberger et al., J.J. Biol.
he . 239:1406-1410 (1964) and Epstein et al., Cold Siring arbor Syr~t p.
0~.~ant. Biol. 28:439-449 (1963)).
Protein disulfide isomerase has been shown to be a resident enzyme of the
endoplasmic reticulum which is retained
in the endoplasmic reticuhtm via a -ICDEL or -HDEL amino acid stquence at its
C-termitnts. Protein disulfide
isomerase and related proteins are further described in Laboissiere, tt al.,
J. Biol. Chem., 270(47:2800fi-28009
(1995); Jeettes, et al., ~, 193(2):151-156 (1997; Koivut~n, et al., en ' ~ ,
42(3):397-404 (1997); and Desilva,
et al., DNA CeI3'Biol,., 15(1):9-16 (1996). These studies indicate the
importance of the identification of protein
disulfide related proteins.
More generally, and also of interest are all novel membrane-bound proteins and
receptors. Such proteins
can play an important role in the formation, differentiation and maintenance
of multicellular organisms. The fate of
many individual cells, e.g., proliferation, migration, differentiation, or
interaction with other cells, is typically
governed by information received from other cells andlor the immediate
environment. This information is often
transmitted by secreted polypeptides (for instance, mitogenic factors,
survival factors, cytotoxic factors, differentiation
factors, neuropeptides, and hormones) which are, in turn, received and
imerpreted by diverse cell receptors or
rnembra~-bound proteins. Such membrane-boutxl proteins and .cell receptors
include, but are not limited to, cytokine
receptors, receptor kinases, receptor phosphatases, receptors involved in cell-
cell interactions, and cellular adhesin
31
CA 02421319 2003-03-25
WO 99146281 PCTlUS99l05028
molecules like selectins and integrins. For instance, transduction of signals
that regulate cell growth and
differentiation is regulated in pan by phosphorylation of various cellular
proteins. Protein tyrosine kinases, enzymes
that catalyze that process, can also act as growth factor receptors. Examples
include fibroblast growth factor receptor
and nerve growth factor receptor.
Membrane-bound proteins and receptor molecules have various industrial
applications, including as
pharmaceutical and diagnostic agents. Receptor itnmunoadhesins, for instance,
can be employed as therapeutic agents
to block receptor-ligand interaction. The membrane-bound proteins can also be
employed for screening of potential
peptide or small molecule inhibitors of the relevant receptorlligand
interaction.
Given the importance of membrane bound proteins, efforts are under way to
identity novel membrane bound
proteins. Moreover, given the importance of disulfide bond-forming enzymes and
their potential uses in a number
of different applications, for example in increasing the yield of correct
refolding of recombittantly produced proteins,
efforts are currently being undertaken by both itiaus>ry and academia to
identify new, native proteins having sequence
identity with protein disulfide isomerase, Many of these efforts are focused
on the screening of mammalian
recombinant DNA libraries to identify the coding sequences for novel protein
disulfide isomerase homologs. We
herein describe a novel polypeptide having sequence identity with protein
disulfide isomerase and the nucleic acids
encoding the same.
64. PR0697
Secreted frizzled related proteins (sFRPs) are related to the frizzled family
of transmembrane receptors.
The sFRPs are approximately 30 kDa in size, and each contains a putative
signal sequence, a frizzled-Like cysteine-
rich domain, and a conserved hydrophilic carboxy-terminal domain. It has been
reported that sFRPs may function
to modulate Wnt signaling, or function as ligands for certain receptors.
Ratmer, et al., PNAS USA, 94(7):2859-2863
(1997). Therefore, sFRPs and proteins having sequence identity andlar
similarity to sFRPs are of interest.
Another secreted protein of interest is any member of the family of secreted
apopwsis-related proteins
(SARPs). Expression of SARPs modifies the intracellular levels of beta-
catettin, suggesting that SARPs interfere with
the Wnt-frizzled proteins signaling pathway. Melkonyan, et al., pNAS USA,
94(25):13636-13641 (1997).
Therefore, SARPs and proteins having sequence identity andlor similarity w
SARPs are of interest.
In addition to sFRPs and SARPs, many extxacellular proteins are of interest.
Extracellular proteins play
an important role in the formation, differentiation and maintenance of
multicellular organisms. The fate of marry
individual cells, e.g., proliferation, migration, differentiation, or
interaction with other, cells, is typically governed
by information received from other cells andlor the immediate environment.
This information is often transmitted
by secreted polypegtides (for iactance, mitogenic factors, survival factors,
cytotoxic factors, differentiation factors,
neutopeptides, and hormones) which are, in txun, received and interpreted by
diverse cell receptors or membrane-
bound proteins. These secreted polypeptides or signaling molecules normally
pass through the cellular secretory
pathway to reach their site of action in the extracellular environment.
Secreted proteins have various industrial applications, including
pharmaceuticals, diagnostics, biosensors
land bioreactors. Most protein drugs available at present, such as
thrombolytic agents, interferons, interleukins,
erythropoietins, colotry stimulating factors, and various other cytokines, are
secretory proteins. Their receptors,
which are membrane proteins, also have potential as therapeutic or diagnostic
agents.
32
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WO 99/46281 PCTIUS99/05028
Efforts are being undertaken by both industry and academia to identify new,
native secreted proteins,
particularly those having sequence identity or similarity with sFRP-2 and SARP-
1. Many efforts are focused on the
screening of mattltnalian recombinant DNA libraries to identify the coding
sequences for novel secreted proteins.
Examples of screening methods and techniques are described in the literature
[see, for example, Klein et al., Proc.
Natl. Acad. Sci., 93:7108-7113 (1996); U.S. Patent No. 5,536,637)).
65. PR0717
Efforts are being undertaken by both industry and academia to identify new,
native transmembrane receptor
proteins. Many efforts are focused on the screening of mammalian recombinant
DNA libraries to identify the coding
sequences for novel receptor proteins. The results of such efforts are
provided herein.
66. PR 731
Cadherins are a large family of transmembrane proteins. Cadlierins comprise a
family of calcium-dependent
glycoproteins that function in mediating cell-cell adhesion in virtually all
solid tissues of multicellular organisms.
At least cadherins 1-13 as welt as types B, E, EP, M, N, P and R have been
characterized. Among the ftmctions
cadherins are known for, with some exceptions, cadherins participate in cell
aggregation and are associated with cell-
cell adhesion sites. Recently, it has been reported that while all cadherins
share multiple repeats of a cadherin
specific motif believed to correspond to folding of exuacellular domains,
members of the cadherin superfamily have
divergent structures and, possibly, functions, In particular it has been
reported that members of the cadherin
superfamily are involved in signal transduction. See, Suzuki,1. Cell Biochem.,
61(4):531-542 (1996). Cadherins
are further described in Tanihara, et al., J. Cell Sci., 107(6):1697-1704
(1994), Aberle, et al., J. Cell Biochem.,
61(4):514-523 (1996) and Tanihara, et al., Cell Adhes: Commun., 2(1):15-26
(1994).
Protocadherins are members of the cadherin superfamily which are highly
expressed in the brain. In some
studies, protocadherins have shown cell adhesion activity, See, Sano; et al.,
Elv~B_O-J., 12(6):2249-2256 (1993).
However, studies have also shown that some protocadherins, such as
protocadherin 3 (also referred to as Pcdh3 or
pc3), do not show strong calcium dependent cell aggregationactiviry. See,
Sago, et al., Genomics, 29(3):631-640
(1995) for this study and further characteristics of Pcdh3.
Therefore, novel members of the cadherin superfamily are of interest. More
generally, all membrane-bound
proteins and receptors are of interest: Such proteins can play an important
role in the formation, differentiation and
maintenance of multicellular organisms. The fate of many individual cells,
e.g., proliferation, migration,
differentiation, or interaction with other cells, is typically governed by
information received from other cells andlor
the immediate environment. This information is often transmitted by secreted
polypeptides (for instance, mitogenic
factors, survival factors, cytotoxic factors, differentiation factors,
neuropeptides, and hormones) which are; in turn,
received and interpreted by diverse cell receptors or membrane-bound proteins.
Such membrane-bound proteins and
cell receptors include, but are not limited to, cytokine receptors; receptor
kinases, receptor phosphatases, receptors
involved in cell-cell interactions, and cellular adhesin molecules like
selectins and integrins. For instance,
transduction of signals that regulate cell growth and differentiation is
regulated in part by phosphorylation of various
cellular proteins. Protein tyrosine kinases, enzymes that catalyze that
process, can also act as growth factor
receptors. Examples include fibroblast growth factor receptor and nerve growth
factor receptor.
33
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WO 99/46281 PCT/US99I05028
Membrane-bound proteins and receptor molecules have various industrial
applications, including as
pharmaceutical and diagnostic agents. Receptor immunoadhesins, for instance,
can be employed as therapeutic agents
to block receptor-ligand interaction. The membrane-bound proteins can also be
employed for screening of potential
peptide or small molecule inhibitors of the relevant receptor/ligand
interaction.
Efforts are therefore being undertaken by both industry and academia to
identify new, native membrane
botmd proteins, particular those having sequence identity with protocadherins,
especially 4, 68, 43, 42, 3 and 5.
Many efforts are focused on the screening of manttt>itliatt recombinant DNA
libraries to identify the coding sequences
for novel membrane-bound proteins. Provided herein are the results of such
efforts.
67. ' PR0218
Efforts are being undertaken by both industry and academia to identify new,
native membrane bound
proteins, particularly those having sequence identity with membrane regulator
proteins. Many efforts are focused
on the screening of matnmaiian recombinant DNA libraries to identify the
coding sequences for novel receptor
proteins.
68. P 8
The integrins comprise a supergene family of cell-surface glycoprotein
receptors that promote cellular
adhesion. Each cell has numerous receptors that define its cell adhesive
capabilities. Integrins are involved in a wide
variety of interaction between cells and other cells or matrix components. The
integrins are of particular importance
in regulating movement and function of immune system cells. The platelet
IIbIIIIA integrin complex is of particular
importance in regulating platelet aggregation. A member of the integrin
family, integrin ii-6, is expressed on epithelial
cells and modulates epithelial inflammation. Another integrin, leucocyte-
associated antigen-1 (LFA-1) is important
in the adhesion of lymphocytes during an immune response.
Of particular interest is H36-alpha 7, an integrity alpha chain that is
developmentally regulated during
myogenesis as described in Song, et al., J. Cell Biol., 117(3):643-657 (1992).
The expression pattern of the laminin-
binding alpha 7 beta 1 integrity is developmentally regulated in skeletal,
cardiac, and smooth muscle. Ziober, et al.,
1~V o1. Biol. yeil, 8(9):1723-1734 (1997). h has been reported that expression
of the alpha 7-Xl/X2 integrity is a novel
mechanism that regulates receptor affinity states in a cell-specific context
and may modulate integrity dependent events
during muscle development and repair. I_~. It has fiut?ter been reported that
laminins promote the locomotion of
skeletal myoblasts via the alpha 7 integrity receptor. In particular it was
reported that alpha 7 beta 1 receptor can
promote myoblast adhesion and motility on a resuicted number of laminin
isofotms and may be important in
myogenic precursor recruianent during regeneration and differentiation. Yao,
et al., J a '., 109(/3):3139-3150
(1996). Spliced variants of integrity alpha 7 are also described in Ixtutg, et
al., ,B'~oc~,em. Bio,~hys. Res. Cornmun.,
243(/):317-325 (1998) and Fornaro and Languino, Matrix Biol,, 16(4):185-I93
(1997). Moreover, it has been
reported that absence of integrity alpha 7 causes a form of muscular
dystrophy. Thus integrins, particularly those
related to integrity 7 and related molecules, are of interest.
In addition to the interest of integrins, more generally, all membrane-bound
proteins and receptors are of
interest since such proteins can play an important role in the formation,
differentiation and maintenance of
multicellular organisms. The fate of many individual cells, e.g.,
proliferation, migration, differentiation, or
interaction with other cells, is typically governed by information received
from other cells andlor the immediate
34
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WO 99/46281 PGT/US99105028
environment. This information is often transmitted by secreted polypeptides
(for instance, mitogenic factors, survival
factors, cytotoxic factors, differentiation factors, tieuropeptides, and
hormones) which are, in turn, received and
interpreted by diverse cell receptors or membrane-bound proteins. Such
membrane-bound proteins and cell receptors
include, but are not limited to, cytoktne receptors, receptor kinases,
receptor phosphatases, receptors involved in cell-
cell interactions, and cellular adhesin molecules like selectins and
integrins. For instance, transduction of signals that
regulate cell growth and differentiation is regulated in pan by
phosphorylation of various cellular proteins. Protein
tyrosine kinases, enzymes that catalyze that process, can also act as growth
factor receptors. Examples include
fibroblast growth factor receptor and nerve growth factor receptor.
Membrane-bound proteins and receptor molecules have various industrial
applications, including as
pham~aceutical and diagnostic agents. Receptor imtnunoadhesins, for instance,
can be employed as therapeutic agents
to block receptor-ligand interaction. The membrane-bound proteins can also be
employed for screening of potential
peptide or small molecule inhibitors of the relevant receptor/ligand
interaction.
Therefore, efforts are being undertaken by both industry and academia to
identify new, native receptor
proteins. Marry efforts are focused on the screening of mammalian recombinant
DNA libraries to identify the coding
sequences for novel receptor proteins. The results of such efforts,
particularly those focused on identifying new
IS polypeptides having sequence identity with integrins, are provided herein.
69. P~7
Testican is a multidomain testicular proteoglycan which is expressed in
numerous tissue types including,
but not limited to neuromuscular tissue, the brain and reproductive tissues.
Testican resembles modulators of cell
social behavior such as the regulation of cell shape, adhesion, migration and
proliferation. [Bonnet, F. et al., ~Biol.
Chem.; x(8):4373 {1996), Perin, J.P. et al., FAX (Switzerland); 70:191 (i994),
Alliel, P.M., et al, Eur. J.J.
~iochem., x(1):346 (1993), Charbonnier, F., et al., C. R. Seances Soc: Biol.
Fil. (France), x(1):127 (1997)].
Among other reasons, since testicau has been implicated in neuronal processes
and may be associated with the growth
of connective tissue, testican and related molecules are of interest.
More generally, all excracellular proteins are of interest. Exuacellular
proteins play an important role in
the formation, differentiation and maintenance of multicellular organisms. The
fate of many individual cells, e.g.,
proliferation, migration, differentiation, or interaction with other cells, is
typically governed by information received
from other cells and/or the immediate environment. This information is often
transmitted by secreted polypeptides
(for instance, mitogenic factors, survival factors; cytotoxic factors,
differentiation factors, neuropeptides, and
hormones) which are, in turn, received and ituetpreted by diverse cell
receptors or membrane-bound proteins: These
polypeptides or signaling molecules normally pass thmugh the celhtlar
secretory pathway to reach their site
of action in the extracellular enviromnent.
Secreted proteins have various industrial applications, including
pharmaceuticals, diagnostics, biosensors
and bioreactors. Most protein drugs available at present, such as thromboly6c
agents, interferons, interltukins,
erythropoietins, colony stimulating factors, and various other cytokines, are
secretory proteins. Their receptors,
which are membrane proteins, also have potential as therapeutic or diagnostic
agents. Efforts are being undertaken
by both industry and academia to identify new, native secreted proteins. Many
efforts are focused on the screening
of rttamtnaiian recombinant DNA libraries to identify the coding sequences for
novel secreted proteins. Examples
of screening methods and techniques are described in the literature [see, for
example, Klein et al., Proc. Natl Acad
CA 02421319 2003-03-25
WO 99/46281 PCT/US99/05028
ci., Q~:7108-7113 (1996); U.S. Patent No. 5,536,637)]. The results of such
efforts, particularly those focused on
identifying molecules having identity andlor similarity with testican are of
interest.
70. PR0733
T1/ST2 is a receptor-like molecule homologous to the type I interleukin-1
receptor, believed to be involved
in cell signaling. The Tl/ST2 receptor and/or putative ligands are further
described in Gayle, et al., J. Biol. them,,
271(10):5784-5789 (1996), Ktunar, et al., J. Biol. Chem., 270(46):27905-27913
(1995), and Mitcham, et al., . Bi I.
Chem., 271(10):5777-5783 (1996). These proteins; and proteins related thereto
are of interest.
More generally all membrane-bound proteins and receptors are of interest since
they can play an important
role in the formation, differentiation and maintenance of multicellular
organisms. The fate of many individual cells,
e.g., proliferation, migration, differentiation, or interaction with other
cells, is typically governed by information
received from other cells and/or the immediate environment. This information
is often transmitted by secreted
polypeptides (for instance, nvtogenic factors, survival factors, cytotoxic
factors, differentiation factors, neuropeptides,
and l~rmones) which are, in turn, received and interpreted by diverse cell
receptors or membrane bound proteins.
Such membrane-bound proteins and cell receptors include, but are not limited
to, cytokine receptors, receptor kinases,
receptor phosphatases, receptors involved in cell-cell interactions, and
cellular adhesin molecules like selectins and
integrins. For instance, transduction of signals that regulate cell growth and
differentiation is regulated in part by
phosphorylation of various cellular proteins. Protein tyrosine kinases,
enzymes that catalyze that process, can also
act as growth factor receptors. Examples include fibroblast growth factor
receptor and nerve growth factor receptor.
Membrane-bound proteins and receptor molecules have various industrial
applications, including as
pharmaceutical and diagnostic agents. Receptor immunoadhesins, for instance,
can be employed as therapeutic agents
to block receptor-ligand interaction. The membrane-bound proteins can also be
employed for screening of potential
peptide or small molecule inhibitors of the relevant receptor/ligand
interaction.
Efforts are being undertaken by both industry and academia to identify new,
native receptor proteins. Many
efforts are focused on the screening of mammalian recombinant DNA libraries to
identify the coding sequences for
novel receptor proteins. The results of such efforts ate provided herein.
71. PR0162
Pancreatitis-associated protein (PAP) is a secretory protein that is
overexpressed by the pancreas during
acute pancreatitis. Serum PAP concentrations have been shown to be abnormally
high in patients with acute
pancreatitis. Pezzilli et al., Am J Gastroenterol., 92(10):1887-1890 (1997).
PAP is syrnhesized by the pancreas due to pancreatic inflammation and has been
shown to be a good serum
marker for injury of the pancreas. In addition, sertun PAP levels appear to
strongly correlate with creatittine
clearance measurements. In patients with a pancreas-kidney transplantation,
PAP may prove to be a useful biological
and histologicai marker of pancreatic graft rejection. Van der Pij1 et al.,
]f'r~nSplantation. 63(?):995-1003 (1997).
Further, PAP has been shown to be useful in screening neonates for cystic
fibrosis. In fact, PAP may discriminate
cystic fibrosis t~onates with better specificity than the current
immunoreactive trypsis assay. Iovatuta et al., C-R.
tlcad. Aci. III, 317(6):561-564.
Secreted proteins such as PAP have various industrial applications, including
pharmaceuticals, diagnostics,
biosensors and bioreactors. Most protein drugs available at present, such as
thrombolytic agents, interferons,
36
CA 02421319 2003-03-25
WO 99/46281 PCT/US99IOS028
interleulans, erythropoietins, colony stimulating factors, and various other
cytokines, are secretory proteins. Their
receptors, which are membrane proteins, also have potential as therapeutic or
diagnostic agents.
Efforts are being undertaken by both industry and academia to identify new,
native secreted proteins. Many
efforts are focused an the screening of marntnalian recombinant DNA libraries
to identify the coding sequences for
novel secreted proteins. Examples of screening methods and techniques are
described in the literature jsee, for
S exatt~ple, Klein et al., Proc. Natl. Acad.Sci., Q~:7108-7113 (1996); U.S.
Patent No. 5,536,63'n]. The results of such
efforts are presented herein.
7Z. P 7
Anti-neoplastic urinary protein (ANUP) was identified as the major protein
present in a fraction of human
urine which exhibits antiproliferative activity against human tumor cell lines
without affecting the growth of several
normal diploid cell lines or tumor cells of mouse or hamster origin. Sloane et
al., Biochem. 1., 234(2):355-362
(1986).
ANUP is a unique cytokine that has beenfound in human granulocytes. The N-
terminal amino acid
sequence has been shown to be unique. A synthetic peptide corresponding to the
first nine residues, with Cys at
positions 4 and 7, was found to be an anti-tumor agent in vitro. Ridge and
Sloane, Cvtolcine, 8(1):1-5 (1996).
Secreted proteins such as ANUP have various industrial applications, including
pharmaceuticals, diagnostics,
biosensors and bioreactors. Most protein drugs available at present, such as
thrombolytic agents, interferons,
interleukins, erythropoietins, colony stimulating factors, and various other
cytokines, are secretory proteins. Their
receptors, which are membrane proteins, also have potential as therapeutic or
diagnostic agents. Efforts are being
undertaken by both industry atxl academia to identify new, native secreted
proteins. Many efforts are focused on the
screening of mammalian recombinant DNA libraries to identify the coding
sequences for novel secreted proteins.
Examples of screening methods and techniques are described in the literature
[see, for example, Klein et al., ro
~fatl. Aead. Sci., ,Q3:7108-7113 (1996); U.S. Patent No. 5,536,b37)].
73. PR01008
Diekkopf 1 (dkk-1) is a member of a family of secreted proteins and functions
in head induction. Dkk 1
is an inducer of Spemarut organizer in amphibian embryos, Glinka, et al.,
Nature, 391 (6665):357-362 (1998). Dkk 1
is a potent arnagottist of Wnt signalling, suggesting that dkk genes encode a
family of secreted Wnt inhibitors. Thus,
dkk-1 family members and related molecules are of interest.
More generally, all .extracellular proteins are of interest since they can
play an important role in the
formation, differentiation and maintenance of mttlticellular organisms. The
fate of many individual cells, e.g.,
proliferation, migration, differentiation, or irneraction with other cells, is
typically governed by information received
from other cells andlor the immediate environment. This information is often
transmitted by secreted polypeptides
(for instance, mitogenic factors, survival factors, cytotoxic factors,
differentiation factors, neuropeptides, and
horrn4nes) which are, in nun; received and inoe<preted by diverse cell
receptors or membrane-bound proteins: These
secreted polypeptides or signaling molecules normally pass through the
cellular secretory pathway to reach their site
of action in the extracellutar emrimmaent.
Secreted proteins have various industrial applications, including
pharmaceuticals, diagnostics, biosensors
and bioreactors. Most protein drugs available at present, such as thromboiytic
agents, interferons, interleukins,
37
CA 02421319 2003-03-25
WO 99/46281 PCTNS99/05028
erythropoietins, colony stimulating factors, and various other cytokines, are
secretory proteins. Their receptors,
which are membrane proteins, also have potential as therapeutic or diagnostic
agents.
Efforts are being undertaken by both industry and academia to identify new,
native secreted proteins,
particularly those related to dkk-I. Many efforts are focused on the screening
of mammalian recombinant DNA
Hbraries to identify the coding sequences for novel secreted proteins.
Examples of screening methods and techniques
S are described in the literature (see, for example, Klein et al., Proc.~Vatl.
Acad. Sci., 93:7108-7113 (1996); U.S.
Patent No. 5,536,637)]. The results of such efforts to identify.molecules
related to dkk-1 are provided herein.
74. PR01012
Protein disulfide isomerase is an enzymatic protein which is involved in the
promotion of correct refolding
of proteins through the establishment of correct disulfide bond formation.
Protein disulfide isomerase was initially
identified based upon its ability to catalyze the renaturation of reduced
denatured RNAse (Goldberger et al., J.J. Biol.
C_hem. 239:1406-1410 (1964) and Egstein et al., Cold Spring Harbor Symp.
Ouant. Biol. 28:439-449 (1963)).
Protein disulfic~ isomerase has been shown to be a resident enzyme of the
endoplasmic reticulum which is retained
in the endoplasmic reticulum via a -KDEL or -HDEL amino acid sequence at its C-
terminus. Protein disulfide
IS isomerase atui related proteins are further described in Laboissiere, et
al., J. Biol. Chem., 270(4?:28006-28009
(1995); 3eenes, et al., a e, 193(2):151-156 (1997; Koivunen, et al., enomics,
42(3):397-404 (1997); and Desilva,
et al., DNA Cel~~iol., 15(1):9-16 (1996). These studies indicate the
importance of the identification of protein
disulfide related proteins.
More generally, the identification of all extracellular and membrane-bound
proteins is of interest since they
play important roles in the formation, differentiation and maintenance of
multicellular organisms. The fate of many
individual cells, e.g., proliferation, migration, differentiation, or
interaction with other cells, is typically governed
by information received from other cells and/or the immediate environment.
This information is often iransmitted~
by secreted polypeptides (for instance, mitogenic factors, survival factors,,
cytotoxic factors, differentiation factors,
neuropeptides, and hormones) which are, in turn, received and interpreted by
diverse cell receptors or membrane-
bound proteins. These secreted polypeptides or signaling molecules normally
pass through the cellular secretory
pathway to reach their site of action in the extracellular environment,
usually at a membrane-bound receptor protein.
Secreted proteins have various industrial applications, including use as
pharmaceuticals, diagnostics,
biosensors and bioreactors. In fact, most protein drugs available at present,
such as thrombolytic agents; interferons,
interleuktns, erythropoietins, colony stimulating factors, and various other
cytokines, are secretory proteins. Their
receptors, which are membrane-bound proteins, also have potential as
therapeutic or diagnostic agents. Receptor
immunoadhesins, for instance, can be employed as therapeutic agents to block
receptor-ligand interaction.
Membrane-bound proteins can also be employed for screening of potential
peptide or small molecule inhibitors of
the relevam receptor/Iigand interaction. Such membrane-bound proteins and cell
receptors include, but are not limited
to, cytokine receptors, receptor kinases, receptor phosphatases, receptors
involved in cell-cell interactions, and
cellular adhesin molecules like selectins and integrins. Transduction of
signals that regulate cell growth and
differentiation is regulated in part by phc~phorylation of various cellular
proteins. Protein tyrosine kinases, enzymes
that catalyze that process, can also act as growth factor receptors. Examples
include fibroblast growth factor receptor
and nerve growth factoit receptor.
38
CA 02421319 2003-03-25
WO 99!4628! PCTIUS99105028
Of particular interest are cellular proteins having endoplasmic reticulum (ER)
retention signals. These
proteins are retained in the cell arid fut>ction closely with endoplasmic
reticulum in protein production. Such proteins
have been 'described previously, l.c., see Shorrosh and Dixon, Pl ., 2(1):51-
58 (1992).
Efforts are being undertaken by both industry and academia to identify new,
native secreted and membrane-
bound receptor proteins, and in particular, cellular proteins having ER
retension signals. Many efforts' are focused
on the screening of mammalian recombinant DNA libraries to identify the coding
sequences for novel secreted and
membrane-bound receptor proteins. Examples of screening methods and techniques
are described in the literature
[see, for example, Klein et al., Proc. Nat!. Acad. Sci., X3_:7108-7113 (1996);
U.S. Patent No. 5,536,63']. The
results of such efforts, particularly the identification of novel polypeptides
and nucleic acids encoding the same, which
have sequence identity and similarity to protein disulfide isomerase are
presented herein.
75. PR01914
Oxygen free radicals and antioxidants appear to play an important role in the
central nervous system after
cerebral ischemia and reperfusion. Moreover, cardiac injury, related to
ischaemia and reperfusion has been reported
to be caused by the action of free radicals. Additionally, studies have
reported that the redox state of the cell is a
IS pivotal determinant of the face of the cells. Furthermore, reactive oxygen
species have been reported to be eytotoxic,
causing inflammatory disease, including tissue necrosis, organ failure,
atheroselerosis, infertility, birth defects,
ptetttature aging, mutations and malignancy. Thus, the control of oxidation
and reduction is important for a number
of reasons including for control and prevention of strokes, heart attacks,
oxidative stress and hypertension. In this
regard, reductases, and particularly, oxidoreductases, are of interest.
Publications further describing this subject
matter include Kelsey, et al., Br. 1. Cancer, 76(7):852-4 (1997); Friedrich
and Weiss, 1. Theor. Biol., 187(4):529-40
{199?) and Pieulle, et a!.,1. Bacceriol., 179(18):5684-92 (1997).
In addition to reductases in particular, novel polypeptides are generally of
interest. Extracellutar proteins
play an important role in the formation, differentiation and maintenance of
multicellular organisms. The fate of many
individual cells, e.g., proliferation, migration, differentiation, or
interaction with other cells, is typically governed
by information received from other cells audlor the immediate environment.
This information is often transmitted
by secreted polypeptides (for instance, mitogenic factors, survival factors,
cytotoxic factors, differentiation factors,
neuropeptides, and hormones) which are, in turn,, received and interpreted by
diverse cell receptors or membrane-
bound proteins. These secreted polypeptides or signaling molecules normally
pass through the cellular secretory
pathway to reach their site of action in the extracellular environment.
Secreted groteins have various industrial applications; including
pharmaceuticals, diagnostics, biosensors
and bioreactors. Mast protein drugs available at present, such as thromboiytic
agents, interferons, interleukins,
erythropoietins, colony stimulating factors, and various other cytokines, are
secretory proteins. Their receptors,
which are membrane proteins, also have potential as therapeutic or diagnostic
agents. Efforts are being undertaken
by both industry and academia to identify new, native secreted proteins. Many
efforts are focused on the screening
of mamatalian recombinant DNA libraries to identify the coding sequences for
novel secreted proteins. Examples
of screening methods and techniques are described in the literature [see, for
example, Klein et al., ~c. Nat!, g~~aa.
,iii, Q~:7108-7113 (1996); U.S. Patent No. 5,536,637)]. The results of such
efforts, particularly those identifying
polypeptides having sequet~e identity with redtxtases, and the nucleic acids
encoding the same, are presented herein.
39
CA 02421319 2003-03-25
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76. PR01017
Enzymatic proteins play important roles in the chemical reactions involved in
the digestion of foods, the
biosynthesis of macromolecules, the controlled release and utilization of
chemical energy. and other processes
necessary to sustain life. Sulfouansferases are enzymes which transfer sulfate
from a sulfate donor to acceptor
substrates, particularly those containing terminal glucoronic acid. The HNK-1
carbohydrate epitope is expressed
on several neural adhesion glycoproteins and a glycolipid, and is involved in
cell interactions. The
glucuronyltransferase and sulfotransferase are considered to be the key
enzymes in the biosynthesis of this epitope
because the rest of the structure occurs often in glycoconjugates. HNK-1
sulfouansfererase is further described in
Bakker, H., et al., J. BioI. trhem., 272(47):29942-29946 (1997).
In addition to HNK-1 sulfouansfererase, and novel proteins related thereto,
all novel proteins ate of
IO interest. Extracellular and membrane-bound proteins play important roles in
the formation, differentiation and
maintenance of multicellular organisms. The fate of many individual cells,
e.g., proliferation, migration,
differentiation, or interaction with other cells, is typically governed by
information received from other cells and/or
the immediate environment. This information is often transmitted by secreted
polypeptides (for instance, mitogenic
factors, survival factors, cytotoxic factors, differentiation factors,
neuropeptides, and hormones) which are, in turn,
received and interpreted by diverse cell receptors or membrane-bound proteins.
These secreted polypeptides or
signaling molecules normally pass through the cellular secretory pathway to
reach their site of action in the
extracellular environment, usually at a membrane-bound receptor protein.
Secreted proteins have various industrial applications, including use as
pharmaceuticals, diagnostics,
biosensors and bioreaetors. 1n fact, most protein drugs available at present,
such as thrombolytic agents, interferons,
interleukins, erythropoietins, colony stimulating factors, and various other
cytakines, are secretory proteins. Their
receptors, which are membrane-bound proteins, also have potential as
therapeutic or diagnostic agents. Receptor
immunoadliesins, for instance, can be employed as therapeutic agents to block
receptor-ligand interaction.
Membrane-bound proteins can also be employed for screening of potential
peptide or small molecule inhibitors of
the relevant receptor/ligand interaction. Such membrane-bound proteins and
cell receptors include, but are not limited
to, cytokine receptors, receptor kinases, receptor phosphatases, receptors
involved in cell-cell interactions, and
cellular adhesin molecules like selectitts and integrins. Transduction of
signals that regulate cell growth and
differentiation is regulated in part by phosphorylation of various cellular
proteins. Protein tyrosine kinases, enzymes
that catalyze that process, can also act as growth factor receptors.
Exatriples include fibroblast growth factor receptor
and nerve growth factor recepmr.
Efforts are being undertaken by both industry and academia to identify new,
native secreted and membrane-
bound receptor proteins, particularly those having sequence identity with HNK-
1 sulfotransferase. Many efforts are
focused on the screening of mammalian recombinant DNA libraries to idenrify
the coding sequences for novel
secreted and membrane-bound receptor proteins. Examples of screening rrtethods
and techniques are described in
the literature [see, for example, Klein et at., Proc. Nab. Acad. Sci., Q~:7108-
7113 (1996); U.S. Patent No.
5,536,637)j. The results of such efforts are provided herein.
77. 74
Enzymatic proteins play important roles in the chemical reactions involved in
the digestion of foods, the
biosynthesis of macromolecules, the controlled release and utilization of
chemical energy, and other processes
CA 02421319 2003-03-25
WO 99/46281 PCT/US99I05028
necessary to sustain life. Glucose dehydrogettase functions in the oxidation
of glucose to gluconate to generate
metabolically useful energy. The regulation of the PQQ-linked glucose
dehydrogenase in different organisms is
reviewed in Neijssel, et al.,. ~ntonie Van Leeuwenhoek, 56{1):51-61 (1989).
Glucose dehydrogenase functions as
an auxiliary energy generating mechanism, because it is maximally synthesized
under conditions of energy stress.
In addition to molecules related to glucose dehydrogettase, all novel proteins
are of interest. Extracellular
and membrane-bound proteins play important roles in the formation,
differentiation and maintenance of multicellular
organisms. The fate of many individual cells, e.g., proliferation, migration,
differentiation, or interaction with other
cells, is typically governed by information received from other cells andlor
the immediate environment. This
information is often uansmitted by secreted polypeptides (for instance,
mitogenic factors; survival factors, cytotoxic
factors, differentiation factors, neuropeptides, and hormones) which are, in
turn, received and interpreted by diverse
cell receptors or membrane-bound proteins. These secreted polypeptides or
signaling molecules normally pass
through the cellular secretory pathway to reach their site of action in the
extracellular environment, usually at a
membrane-bound receptor protein.
Secreted proteins have various industrial applications, including use as
pharmaceuticals, diagnostics,
biosensors and bioreactors. In fact, most protein drugs available at present,
such as thrombolytic agents, interferons,
interleukuts, erythropoietins, colony stimulating factors, and various other
cytokines, are secretory proteins. Their
receptors, which are membrane-bound proteins, also have potential as
therapeutic or diagnostic agents. Receptor
immunoadhesins, for instance, can be empioyed as therapeutic agents to block
receptor-ligand interaction.
Membrane-bound proteins can also be employed for screening of potential
peptide or small molecule inhibitors of
the relevant receptor/ligand interaction. Such membrane-bound proteit~ and
cell receptors include, but are not limited
to, cytokine receptors, receptor kinases, receptor phosphatases, receptors
involved in cell-cell interactions, and -
cellular adhesin molecules like selectins and integrins. Transduction of
signals that regulate cell growth and
differetroation is regulated in part by phosphorylation of various celiular
proteins. Protein tyrosine kinases, enzymes
that catalyze that process, can also act as growth factor receptors. Examples
include fibroblast growth factor receptor
and nerve growth factor receptor:
Efforts are being undertaken by both industry and academia to identify new,
native secreted and membrane-
bound receptor proteins, and particularly cellular proteins and those related
to dehydrogenase or oxidoreductase.
Many efforts are focused on the screening of mammalian recombinant DNA
libraries to identify the coding sequences
for novel secreted and membrane-bound receptor proteins. Examples of screening
methods and techniques are
described in the literature [see, for example; Klein et al., Proc.,_Natl.
Acad. Sci., Q~:7108-7113 (1996); U.S. Patent
No. 5,536,637)]. The results of such efforts are presented herein.
?8. PR01031
It has been reported that the cytokine interletzlatt 17 (IL17) stimulates
epithelial, endothelial, and fibroblastic
cells to secrete cytokines such as IL-6, IL-8, and granulocyte-colony-
stimulating factor, as well as prostaglandin E2.
Moreover, it has been shown that when cultured in the presence of IL-17,
fibroblasts could sustain proliferation of
CD34+ preferential maturation into neutrophils. Thus it has been suggested
that IL-17 constitutes an early initiator
of the T cell-dependent inflammatory reaction andlor an element of the
cytokine network that bridges the immune
system to hematopoiesis. See, Yao, et al., J.J. Im ni unol.; 155(12):5483-5486
(1995); Fossiez, et al., Ji];~~C .~ Med.,
1830:2593-2603 (1996); Ket»dy, et ai., I. Inter~rr. on C'wtokine Res.,
16(8):611-617 (1996). Thus, proteins related
41
CA 02421319 2003-03-25
WO 99/46281 PCT/U599/05028
to IL-17 are of interest.
More generally, all novel proteins are of interest. Extracellular proteins
play an important role in the
formation, differentiation and maintenance of multicellular organisms. The
fate of many individual cells, e.g.,
proIiferatian, migration, differentiation, ar interaction with other cells, is
typically governed by information received
from other cells andlor the immediate environment. This information is often
transmitted by secreted polypeptides
(for instance, mitogenic factors, survival factors, cytotoxic factors,
differentiation factors, neuropeptides, and
hormones) which are, in turn, received and interpreted by diverse cell
receptors or membrane-bound proteins. These
secreted polypeptides or signaling molecules normally pass through the
cellular secretory pathway to reach their site
of action in the extracellular environment.
Secreted proteins have various indusuial applications, including
pharmaceuticals, diagnostics, biosensors
and bioreactors. Most protein drugs available at present, such as thrombolytic
agents; interferons, interleukins,
erythropoietins, colony stimulating factors, and various other cytokines, are
secretory proteins. Their receptors,
which are membrane proteins, also have potential as therapeutic or diagnostic
agents.
Efforts are being undertaken by both industry and academia to identify new,
native secreted proteins,
particularly those related to IL-17. Many efforts are focused on the screening
of matntnalian recombinant DNA
libraries to identify the coding sequences for hovel secreted proteins.
Examples of screening methods and techniques
ate described in the literature [see, for example, Klein et al., Proc. Natl.
Acad. Sci., x:7108-7113 (1996); U.S.
Patent No. 5,536,637)]. The results of such efforts are presented herein.
79. PR0938
Protein disulfide isomerase is an enzymatic protein which is involved in the
promotion of correct refolding
of proteins through the establishment of correct disulfide bond formation:
Protein disulfide isomerase was initially
identified based upon its ability to catalyze the renaturation of reduced
denatured RNAse (Goldberger et at., J t 1.
Chem. ~Q:1406-1410 (1964) and Epstein et al., ,old Spring Harbor Sym"p Ouant
Bioi. .8,:439-449 (1963)).
Protein disulfide isomerase has been shown to be a resident enzyme of the
endoplasmic reticulum which is retained
23 in the endoplasmic reticulum via a -KDEL or -HDEL amino acid sequence at
its C-terminus. Protein disulfide
isomerase and related proteins are further described in Laboissiere, et al., J-
Bio . C,~em., 0 47 :28006-28009
(i995); Jeenes, et al., Ger e_, 193(21:151-156 (1997); Koivunen, et al., eno '
, x:397-404 (1997); Desilva, et
at., DNA Cell Bio~., x:9-16 (1996); Freedman, et aI. T~e_xtds in B~chem. Sci.
,~,Q:331-336 (1994}; Bulleid, N.J.
Advances i~ Prot. Chem. 44:125-50 (1993); and Noiva, R., Prot. Exp. and
Purification ~:1-13 (1994}. These studies
indicate the importance of the identification of protein disulfide related
proteins.
More generally; and also of interest are all novel membrane-bound proteins and
receptors. Such proteins
can play an implant role in the formation, differentiation and maintenance of
multicellular organisms. The fate of
many individual cells, e.g:, proliferation; migration, differentiation, or
interaction with other cells, is typically
governed by information received from other cells and/or the immediate
environment. This information is often
transmitted by secreted polypeptides (for ittstattce, rnitogettic factors,
survival factors, cytotoxic factors, differentiation
factors, neuropeptides, and hormones) which are, in mrn, received and
interpreted by diverse cell receptors or
r~nobrane-bound proteins. Such membrane-bound proteins and ceD receptors
include, but are not limited to, cytokine
receptors, receptor kinases, receptor phosphatases, recepwrs involved in cell-
cell interactions, and cellular adhesin
rnolecules like selectins and integrins. For instance, aansduction of signals
that regulate cell growth and
42
CA 02421319 2003-03-25
WO 99/46281 PCTIUS99/05028
differentiation is regulated in part by phosphorylation of various cellular
proteins. Protein tyrosine kinases, enrymes
that catalyze that process, can also act as growth factor receptors. Examples
include ftbroblast growth factor receptor
and nerve growth factor receptor.
Membrane-bound proteins and receptor molecules have various industrial
applications, including as
pharmaceutical and diagnostic agents. Receptor intnnunoadhesins, far instance,
can be employed as therapeutic agents
to block receptor-ligand interaction. The membrane-bound proteins can also be
employed for screening of potential
peptide or small molecule inhibitors of the relevant receptor/ligand
interaction.
Given the importance of membrane bound proteins, efforts are under way to
identity novel membrane bound
proteins. Moreover, given the importance of disulfide bond-forming enzymes and
their potential uses in a number
of different applications, for example in increasing the yield of correct
refolding of recombinantly produced proteins,
efforts are currently being undertaken by both industry and academia to
identify new, native proteins having sequence
identity with protein disulfide isomerase. Many of these efforts are focused
on the screening of mammalian
recombinant DNA libraries to identify the coding sequences for novel protein
disulfide isomerase homologs.
We herein describe the identification and characterization of a novel
polypeptide having homology to protein
disulfide isomerase.
IS
80. PROIO,~
The low density lipoprotein (LDL) receptor is a membrane-bout protein that
plays a key role in cholesterol
ltorneostacis, mediating cellular uptake of lipoprotein particles by high
affinity binding to its ligands, apolipoprotein
(apo) B-100 and apoE. The ligand-bring domain of the LDL receptor contains 7
cysteine-rich repeats of
approximately 40 amino acids, wherein each repeat contains 6 cys;eines, which
form 3 infra-repeat disulfide bonds.
These unitNe s~cairal features provide the LDL receptor with its ability to
specifically interact with apo B-100 and
apoE, thereby allowing for transport of these lipoprotein particles across
cellular membranes and metabolism of their
componenic. Soluble fragments containing the extracellular domain of the LDL
receptor have been shown to retain
the ability to interact with its specific lipoprotein ligands (Simmons et al.,
,fi. Biol. Chem. 272:25531-25536 (1997)).
LDL receptors are further descn'bed in Javitt, ~5., 9(13):1378-1381 (1995) and
Herz and Willnow,
Acad. Sci., 737:14-19 (1994). Thus, proteins having sequence identity with LDL
receptors are of interest. .
More generally, ali membrane-bound proteins and receptors can play an
important role in the formation,
differentiation and maintenance of multicelhtlar organisms. The fate of many
individual cells, e.g., proliferation,
migration, differentiation, or interaction with other cells, is typically
governed by information received from other
cells atndlor the immediate environrr~rtt. This information is often
uansmitted by secreted polypeptides (for instance,
mitogeruc factors, survival factors, cytotoxic factors; differentiation
factors; txuropeptides; and hormones) which are,
in rum, received and interprtted by diverse cell receptors or membrane-bound
proteins. Such membrane-bound
proteitLS and cell receptors inchtde, but are not limited to; cytolatie
receptors, receptor kinases, receptor phosphatases,
trceptors imrolved in cell-cell interactions, and cellular adhesin molecules
like selectins and integrins. For instance,
transduction of that regulate cell growth and differentiation is regulated in
part by phosphorylation of various
cellular proteins. Protein tyrosine kinases, enzymes that catalyze that
process, can also act as growth factor
receptors. Examples include fibroblast growth factor receptor and nerve growth
factor receptor. 'Of particular
interest are membrane bound proteins that have type II transmembrane domains.
43
CA 02421319 2003-03-25
WO 99/46281 PCT/US99/85028
Membrane-bound proteins and receptor molecules have various indusuial
applications, including as
pharmaceutical and diagnostic agents. Receptor imrnunoadhesins, far instance,
can be employed as therapeutic agents
to block receptor-ligand interaction. The membrane-bound proteins can also be
employed for screening of potential
peptide or small molecule inhibitors of the relevant receptorlligand
interaction.
Efforts are thus being undertaken by both industry and academia to identify
new, native proteins, particularly
membrane bound proteins including type II transmembrane bound proteins. Many
efforts are focused on the
screening of mammalian recombinant DNA libraries to identify the coding
sequences for novel receptor proteins.
The results of such efforts are provided herein.
81: P_-801083
Of particular interest are membrane bound proteins that belong to the seven
uansmembrane (7TM) receptor
superfamily. Hxamples of these receptors include G-protein coupled receptors
such as ion receptors. Another
example of a TTM receptor superfamily member is described in Osterhoff, et
al., DNA CeIl,~iol., 16(4):3'79-389
(1997).
Membrane-bound proteins and receptor molecules have various industrial
applications, including as
pharmaceutical and diagnostic agents. Receptor immunoadhesins, for instance,
can be employed as therapeutic agents
to block receptor-ligand interaction. The membrane-bound proteins can also be
employed for screening of potential
peptide or small molecule inhibitors of the relevant receptorJligand
interaction.
Efforts are being undertaken by both industry atxi academia to identify new,
native receptor proteins. Many
efforts are focused on the screening of mammalian recombinant DNA libraries to
identify the coding sequences fox
novel receptor proteins. The results of such efforts are presented herein.
82. PRO
Polypeptides involved in survival, proliferation and/or differentiation of
cells are of interest. Polypeptides
known to be involved in the survival, proliferation and/or differentiation of
cells include VEGF and members of the
bone morphogenetic protein family. Therefore, novel polypeptides which are
related to either VEGF or the bone
morphogenetic protein are of interest.
The heparin-binding endothelial cell-growth factor, VEGF, was identified and
purified from media
conditioned by bovine pituitary follicular or foliiculo-stellate cells over
several years ago. See Ferrara et al.,
Biophys. Res. Comm. 161, 851 (1989). VEGF is a naturally occurring compound
that is produced in follicular or
folliculo-stellate cells (FC), a morphologically well characterized population
of granular cells. The FC are stellate
cells that send cytoplasmic processes between secretory cells. .
VEGF is expressed in a variety of tissues as multiple homodimeric forms (121,
165, 189 and 206 amino
acids per monomer) resulting from alternative RNA splicing. VEGF,,1 is a
soluble mitogen that does not bind
heparin; the longer forms of VEGF bind heparin with progressively higher
affinity. The heparin binding forms of
VEGF can be cleaved in the carboxy terminus by plasmin to release (a)
diffusible forms) of VEGF. Amino acid
sequencing of the carboxy terrninal peptide identified after plasmin cleavage
is Arg,~a Ala,l,. Amino terminal "core"
per, VEGF (1-110) isolated as a homoditner, binds neutralizing monoclonal
antibodies (4.6.1 and 2E3) and soluble
forms of FMS-like tyrosine kinase (FLT 1), kinase domain region (KDR) and
fetal liver kinase (FLK) receptors with
similar affinity compared to the intact VEGF,~ homodimer.
44
CA 02421319 2003-03-25
WO 99146281 ~ PCT/US99/05028
As noted, VEGF contains two domains that are responsible respectively for
binding to the KDR and FLT-1
receptors. These receptors exist only on endothelial (vascular) cells. As
cells become depleted in oxygen, because
of trauma and the Like, VEGF production increases in such cells which then
bind to the respective receptors in order
to signal ultimate biological effect. The signal then increases vascular
permeability and the cells divide and expand
to form new vascular pathways - vasculogenesis and angiagenesis.
Thus, VEGF is useful for treating conditions in which a selected action on the
vascular endothelial cells;
in the absence of excessive tissue growth, is important, for example, diabetic
ulcers and vascular injuries resulting
from trauma such as subcutaneous wounds. Being a vascular (artery and venus)
endothelial cell growth factor, VEGF
restores cells that are damaged, a process referred to as vasculogenesis, and
stimulates the formulation of new
vessels, a process referred to as angiogenesis.
VEGF would also find use in the restoration of vasculature after a myocardial
infarct, as well as other uses
that can be deduced. In this regard, inhibitors of VEGF are sometimes
desirable, particularly to mitigate processes
such as angiogenesis and vasculogenesis in cancerous cells.
Regarding the bone morphogenetic protein family, members of this family have
been reported as being
involved in the differentiation of cartilage and the promotion of
vascuiarization and osteoinduction in preformed
hydroxyapatite. Zou, et al., Genes Dev. (U.S.), 11(i7):2191 (1997); L,evine,
et al., Ann. last. Surø., 39(2):I58.
(1997). A number of related bane morphogenetic proteins have been identified,
all members of the bone
morphogenetic protein (BMP) family. Bone marphogenetic native and mutant
proteins; nucleic acids encoding
therefor, related compounds including receptors, host cells and uses are
further described in at Least: U.S. Patent Nos.
5,670,33$; 5,454,419; 5,661,007; 5,637,480; 5,631,142; 5,166,058; 5,620,867;
5,543,394; 4,877,864; 5,013,649;
55,106,748; and 5,399,677. Of particular interest are proteins having hamology
with bone morphogenetic protein
I, a procollagen C-proteinase that plays key roles in regulating matrix
deposition.
The present imrention is predicated upon research intended to identify novel
polypeptides which are related
to VEGF and the BMP family, and in particular, palypeptides which have a role
in the survival, proliferation and/or
differentiation of cells. While the novel polypeptides are not expected to
have biological activity identical to the
known polypeptides to which they have homology, the known polypeptide
biological activities can be used to
detern~ the relative biological activities of the novel polypeptides. In
particular, the novel polypeptides described
herein can be used in assays which are intended to determine the ability of a
polypeptide to induce survival,
proliferation or differentiation of cells. In turn, the results of these
assays can be used accordingly, for diagnostic
and therapeutic purposes. The results of such research is the subject of the
present invention.
83. PR0285 and PR0286
The cloning of the Toll gene of Drosoptula, a maternal effect gene that plays
a central role in ttte
establishment of the embryonic dorsal-ventral pattern, has been reported by
Hashimoto et at., ~e 5,~, 269-279
(1988). The Drosophila Toll gene encodes an integral membrane protein with an
extracytoplasmic domain of 803
amino acids and a cytoplasrrric domain of 269 amino acids. The exaacytoplasmic
domain has a potential membrane-
spanning segment, and contains multiple copies of a leucine-rich segment, a
structural motif found in many
transmembrane proteins. The Toll protein controls dorsal-ventral patterning in
Drosoptula embryos and activates
the transcription factor Dorsal upon binding to its ligand Spatzle. (Morisato
and Anderson, eI 76, 677-688 (1994).)
In adult Drosopluta, the TollIDorsai signaling pathway participates in the
anti-fungal imunme response. (ixnaitre et
CA 02421319 2003-03-25
WO 99!46281 PCT/US99/05028
al., Ce~l_ $6, 973-983 (1996).)
A human homologue of the Drosophila Toll protein has been described by
Medzhitov et al., attire 388,
394-397 (1997). This human Toll, just as Drosophila Toll, is a type I
transmembrane protein, with an extracellular
domain consisting of 21 tandemly repeated leucine-rich motifs (leucine-rich
region - LRR), separated by a non-LRR
region, and a cytoplasmic domain homologous to the cytoplasmic domain of the
human interleukin-1 (IL-1) receptor.
A constitutively active mutant of the human Toll uansfected into human cell
lines was shown to be able to induce
the activation of NF-xB and the ezpression of NF-tcB-controlled genes for the
inflammatory cytokines IL-I, IL-6 and
IL-8, as well as the expression of the constimulatory molecule B7.1; which is
required for the activation of native
T cells. It has been suggested that Toll functions in vertebrates as a non-
clonal receptor of the immune system, which
can induce signals for activating both an innate and an adaptive immune
response in vertebrates. The human Toll
gene reported by Medzhitov et al., supra was most strongly expressed in spleen
and peripheral blood leukocytes
(PBL), and the authors suggested that its expression in other tissues may be
due to the presence of macrophages and
dendritic cells, in which it could act as an early-warning system for
infection. The public GenBank database contains
the following Toll sequences: Toll! (DNAXII HSU88540-1, which is identical
with the random sequenced full-length
cDNA IfHUMRSC78b-1); TolI2 (DNAXIi HSU88878-1); To113 (DNAXII HSU88879-1); and
To114 {DNAXII
HSU88880-1, which is identical with the DNA sequence reported by Medzhitov et
al., supra}. A partial ToII
sequence (1'0115) is available from GenBank under DNAX~ HSU88881-1.
Further human homologues of the Drosophila Toll protein, designated as Toll-
Like receptors (huTLRsl-5)
were recently cloned and shown to mirror the topographic structure of the
Drosophila counterpart (Rock et al., Proc.
Nal,[_,~cad. Sci. USA 95, 588-593 [1998]}. Overexpression of a constitutively
active mutant of one human TLR
(Toll protein homologue - Medzhitov et al., supra; TLR4 - Rock et al., supra)
leads to the activation of NF-x8 and
induction of the inflammatory cytokines and constimulatory molecules.
Medzhitov et al., supra.
84. PR0213-1, PR01330 and PR014A9
Cancer is characterized by the increase in the number of abnormal, or
neoplastic, cells derived from a
normal tissue which proliferate to fotrn a tumor mass, the invasion of
adjacent tissues by these neoplastic tumor cells,
and the generation of malignant cells which eventually spread via the blood or
lymphatic system to regional lymph
nodes and to distant sites (metastasis). In a cancerous state a cell
proliferates under conditions in which normal cells
would not grow. Cancer manifests itself in a wide variety of forms,
characterized by different degrees of
invasiveness and aggressiveness.
Alteration of gene expression is. intimately related to the uncontrolled cell
growth and de-differentiation
which are a common feature of all cancers. The genomes of certain well studied
tumors have been found to show
decreased expression of recessive genes, usually referred to as tumor
suppression genes, which would normally
function to prevern malignant cell growth, and/or overexpression of certain
dominant genes; such as oricogenes, that
as to pror~ce malignant growth. Each of these genetic changes appears to be
responsible for importing sotxzc of the
traits that, in aggregate, represent the full neoplastic phenotype (Hunter,
Cell CSI, l I29 [1991]; Bishop, Cell 64, 235-
248 [1991]).
A well known trtiecchanism of gene (e.g. otxogene) overexpression in cancer
cells is gene amplification. This
is a process where in the chromosome of the ancestral cell multiple copies of
a particular gene are produced. The
process involves unscheduled replication of the region of chromosome
comprising the gene, followed by
46
CA 02421319 2003-03-25
WO 99146281 ~ PCT/US99105028
recombination of the replicated segments back into the chromosome (Alitalo et
aL, Adv. Cancer Res. 47, 235-281
[I986]). It is believed that the overexpression of the gene parallels gene
amplification, i.e. is proportionate to the
number of copies made.
Proto-oncogenes that encode growth factors and growth factor receptors have
been identified to play
important roles in the pathogenesis of various human malignancies, including
breast cancer. For example, it has been
found that the human ErbB2 gene (erbB2, also known as her2, or c-erbB-2),
which encodes a 185-kd transmembrane
glycoprotein receptor (p185HER2; HER2) related to the epidermal. growth factor
receptor (EGFR), is overexpressed
in about ZS% to 30% of human breast cancer (Slamon et al., Science 235:177-I82
[i987J; Slamonet al., Science
244:707-712 [1989]).
It has been reported that gene amplification of a protooncogene is an event
typically involved in the more
malignant forms of cancer, and could act as a predictor of clinical outcome
(Schwab et al., Genes Chromosomes
Cancer l, 181-193 [1990]; Alitalo et al., supra). Thus, erbB2 overexpression
is commonly regarded as a predictor
of a poor prognosis, especially in patients with primary disease that involves
axillary lymph nodes (Slamon et al.,
j1987] and [1989], supra; Ravdin and Chamness, Gene 159:19-27 [1995]; and
Hynes and Stem, Biochem Biophys
Acta 1198: 165-184 [1994]), and has been linked to sensitivity andlor
resistance to hormone therapy and
chemotherapeutic regimens, including CMF (cyclophosphatnide, methotrexate, and
fluoruracil) and anthracyclines
(Baselga et al., Oncology 1 I (3 Suppl 1): 43-48 [1997]). However, despite the
association of erbB2 overexpression
with poor prognosis, the odds of HER2-positive patients responding clinically
to treatment with taxanes were greater
than three times those of HER2-negative patients (Ibid). A recombinant
humanized anti-ErbB2 (anti-HER2)
raonoclonal antibody (a humanized version of the marine anti-ErbB2 antibody
4D5, referred to as rhuMAb HER2
ZO or Hereeptin 70) has been clinically active in patients'with ErbB2-
overexpressing metastatic breast cancers that had
received extensive prior anticancer therapy: ($aselga et al.; J. Clin. Oncol.
14;737-744 [1996]).
Tt~ protein Notch and its homologues are key regulatory receptors in
determining the cell fate in various
development processes. The protein Notch-4, also known as int-3 oncogene, was
originally identified as a frequent
uarget in mouse matnmary tumor virus (MMVS). Notch-4 is believed to be a
transgene which affects the
differentiation capacity of stem cells and Ieads to neoplastic proliferation
in epithelial cells. Shirayoshi et al., Genes
Cells Z(3): 2I3-224 (1997). During embryogenesis, the expression of Notch-4
was detected in endothelial cells of
blood vessels forming tissues such as the dorsal aorta, intersegmental
vessels, yolk sac vessels, cephalic vessels,
heart, vessels in branchial arches, and capillary plexuses. Notch-4 expression
in these tissues was also associated
with flk 1, the major regulatory gene of vascttlogenesis and angiogenesis.
Notch-4 is also upreguiated in vitro during
the dilfation of endothelial.stem cell. The endothelial cell specific
expression pattern of Notch-4, as well as its
s~cwray similarity to Notch suggest that Notch-4. is an endothelial cell
specific homologue of Notch and that it may
play a role in vaculogenesis and angiogenesis.
8~. PR029$
Efforts are being undertaken by both industry arid academia to identify new,
native receptor proteins. Many
efforts are focused an the screening of rmamma3ian recombinant DNA libraries
to identify the coding sequences for
novel receptor proteins. We herein describe the identification and
characterization of novel transmembrane
polypeptides, designated herein as PR0298 polypeptides.
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CA 02421319 2003-03-25
WO 99/4b281 PCT/US99/05028
86. ~R0337
Neuronal development in higher vertebrates is characterized by processes that
must successfully navigate
distinct cellular environrnent.en route to their synaptic targets. The result
is a functionally precise formation of neural
circuits. The precision is believed to result form mechanisms that regulate
growth cone pathfmding and target
recognition, followed by latter refinement and remodeling of such projections
by events that require neuronal activity,
Goodman and Shatz, Cell/Neuron [Suppl.] 72(10): 77-98 (1993). It is further
evident that different neurons extend
nerve fibers that are biochemically distinct and rely on specific guidance
cues provided by cell-cell, cell-matrix, and
chemotrophic interactions to reach their appropriate synaptic targets, Goodman
et al., supra.
One particular tz~ans by which diversity of the neuror>al cell surface may be
generated is through differential
expression of cell surface proteins referred to as cell adhesion molecules
(CAMS). Neuronally expressed CAMS have
been implicated in diverse developmental processes, including migration of
neurons along radial glial cells, providing
permissive or repulsive substrates for neurite extension, and in promoting the
selective fasciculation of axons in
projectional pathways. lessel, Neuron 1: 3-13 (1988); Edelinan and Crossin,
Annu. Rev. Biochem. 60: 1S5-190
(1991). Interactions between CAMs present on the growth cone membrane and
molecules on opposing cell
membranes or in the extracellular matrix are thought to provide the specific
guidance cues that direct nerve fiber
outgrowth along appropriate projectional pathways. Such interactions are
likely to result in the activation of various
second messenger systems within the growth cone that regulate neurite
outgrowth. Doherry and Walsh, Curr. Opin
Neurobiol. 2: S95-601 (1992}.
In higher vertebrates, most neural CAMS have been found to be members of three
major structural families
of proteins: the irnegrins; the cadherins, and the immunoglobulin gene
superfamily (igSF}. Jessel, supra.; Takeichi,
Anrru. Rev. Biochem. 59: 237 2S2 (1990); Reichardt and Tomaselli, Annu. Rev.
Neurosci. 14: 531-S70 (1991). Cell
adhesion molecules of the IgSF (or Ig-CAMs), in particular, constitute a large
family of proteins frequently implicated
in neural cell interactions and nerve fiber outgrowth during development,
Salzer and Colman, Dev. Neurosci. 11:
37?-390 (1989); Briimmendorf and Rathjen, J. Neurochem. 6I: 1207-1219 (1993).
However, the majority of
mammalian Ig-CAMs appear to be too widely expressed to specify navigational
pathways or synaptic targets
suggesting ihat other CAMS, yet to be identified, have role in these more
selective interactions of neurons.
Many of the known neural Ig-CAMs have been found to be attached to the plasma
membrane via a
glycosylphosphatidylinositol (GPI) anchor. Additionally, many studies have
implicated GPI-anchored proteins in
providing specific guidance cues during the outgrowth on neurons in specific
pathways. In studies of the grasshopper
nervous system, treatment of embryos with phosphatidylinositol-specific
phopholipase C tPIPLC), which selectively
removes GPI-anchored proteins from the surfaces of cells, resulted in
misdirection and faulty navigation among
subsets of pioneering growth cones, as well as inhibited migratory patterns of
a subset of early neurons, Chang et
al., Devel. l I4: 507-519 (1992). The projection of retinal fibers w the optic
rectum appears to depend, in part, on
a 33 kDa GPI-anchored protein, however, the precise nature of this protein is
unknown. Stahl et al., Neuron 5: 735-
743 (1990).
3S The expression of various GPI-anchored proteins has been characterized
amongst the different populations
of primary rat neurons amongst dorsal root ganglion, sympathetic neurons of
the cervical ganglion, sympathetic
neurons of the superior cervical ganglion, and cerebellar granule neurons.
Rosen et al., J. Cell Biol. 117: 617-b27
(1992). In contrast to the similar pattern of total membrane protein
expression by these different types of neurons,
stribt~g differences were observed in the expression of GPI-anchored proteins
between these neurons. Recently, a
48
CA 02421319 2003-03-25
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65 kDa protein band known as neurotrimin was discovered and found to be
differentially expressed by primary
neurons (Rosen et al., supra), and restricted to the nervous system and found
to be the most abundant and earliest
expressed of the GPI-anchored species in the CNS. Struyk et aL, J.
Neuroscience IS(3): ZI41-2156 (1995). The
discovery of neurotrimin has further lead to the identification of a family of
IgSF members, each containing three Ig-
Iike domains that share significant amino acididentity, rtow ternted IgLON.
Sttvyk et al., supra; Pimenta et al., Gene
170(2): 189-95 (1996).
Additional members of the IgLON subfamily include opiate binding cell adhesion
molecule (OBCAM),
Schofield et al" EMBO J. 8: 489-495 (1989); limbic associated membrane protein
(LAMP), Pimenta et al:, supra;
CEPU-I; GPSS, Wilson et al., J. Cell Sci. 109: 3129-3138 (1996); Eur. J.
Neurosci. 9(2): 334-41 (1997); and
AvGp50, Hancox et al., Brain Res. Mol. Brain Res. 44(2): 273-85 (1997).
While the expression of neurotrimin appears to be widespread, it does appear
to correlated with the
development of several neural circuits. For example, between E18 and P10,
neurotimin mRNA expression within
the forebrain is maintained at high levels in neurons of the developing
thalamus, cortical subplate, and cortex,
particularly laminae V and VI (with less intense expression in II, II, and IV;
and minimal expression in lamina I).
Cortical subplate neurons may provide an early; temporary scaffold for the
ingrowing thalamic afferents en route to
their final synaptic targets in the cortex. Atlendoerfer and Shatz, Anrnt.
Rev. Neurosci. 17: 185-218 (1994).
Conversely, subplate neurons have been suggested to be required for cortical
neurons from layer V to select VI to
grow into the thalamus, and neurons from layer V to select their targets in
the colIiculus, pons, and spinal cord
(McConnell et al., J. Neurosci. 14: 1892-1907 (1994). The high level
expression of neurotrimin in many of these
projections suggests that it could be involved in their development.
In the hindbrain, high levels of neurotrimin message expression were observed
within the pontine nucleus
and by the internal granule cells and Purkinje cells of the cerebellum. The
pontine nucleus received afferent input
from a variety of sources including corticopontirie fibers of layer V, and is
a major source of afferent input, via mossy
fibers, to the granule cells which, in turn, are a major source of afferent
input via parallel fibers to Purkinje cells.
]Patsy and Chan-Palsy, The cerebellar cortex: cytology and organization. New
York: Springer (1974]. High level
expression of neurotrimin these neurons again suggests potential involvement
in the establishment of these circuits.
Neurotrimin also exhibits a graded expression pattern in the early postnatal
striatum. Increased neurotrimin
expression is found overlying the dorsolateral striatum of the rat, while
lesser hybridization intensity is seen overlying
the ventromedial striatum. Struyk et aL, supra. This region of higher
neurottimitt hybridization intensity does not
cotrespo~i to a cytoarchitecturally differentiable region, rather it
corresponds to the primary area of afferent input
from layer VI of the contratateral sensorirnotor cortex (Gerfen, Nature 3I1:
461-464 (/984); Donoghue and
Herkenham, Brain Res. 365: 397-403 (1986)). The venuomedial striatum, by
contrast, receives the majority of its
afferent input from the perirhinal and association: cortex. It is noteworthy
that a complementary graded pattern of
LAMP expression, has been observed within the striatium, with highest
expression in venuomedial regions, and
lowest expression dorsolaterally. Levitt, Science 223: 299-301 (1985);,
Chesselet et al., Neuroscience 40: 725-733
(1991).
87. PR0403
Type II transmembrane proteins, also known as single pass ttansmembrane
proteins have an N-terminal
portion lodged in the cytoplasm while the C-terminal portion is exposed to the
extracelhttar domain.
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CA 02421319 2003-03-25
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Endothelia is a family of vasoconstrictor peptides about which much activity
has been focused to better
understand its basic pharmacological, biochemical and molecular biological
features, including the presence and
structure of isopeptides and their genes (endothelia-I, -2 and u3), regulation
of gene expression, intracellular
processing, specific endothelia converting enzymes (ECE), receptor subtypes
(ET-A and ET-B), intracellular signal
transduction following receptor activation, etc.
The endothelia (ET) family of peptides have potent vascular, cardiac and renal
actions which may be of
pathophysiological importance in many human disease states. . ET-1 is
expressed as an inactive 212 amino acid
prepropeptide. The prepropeptide is first cleaved at Arg52-Cys53 and Arg92-
AIa93 and then the carboxy terminal
Lys91 and Arg92 are trimmed from the protein to generate the propeptide big ET-
1.
Endothelia is generated from inactive intermediates, the big endothelins, by a
unique processing event
catalyzed by the zinc metalloprotease, endothelia converting enzyme (ECE). ECE
was recently cloned, and its
structure was shown to be a single pass transmembrane protein with a short
intracellular N-terminal and a long
extracellular C-terminal that contains the catalytic domain and numerous N-
glycosylation sites. ECEs cleave the
endothelia propeptide betw~n Trp73 and Va174 producing the active peptide, ET,
which appears to function as a
Local xather than a circulating hormone (Rubanyi, G.M. & Polokoff, M.A.,
Pharmachological Reviews 46: 325-415
(1994): Thus ECE activity is a potential site of regulation of endothelia
production and a possible target for
therapeutic intervention in the endothelia system. By blocking ECE activity,
it is possible stop the production of ET-I
by inhibiting the conversion of the relatively inactive precursor, big ET-1,
to the physiologically active form.
Endothelins may play roles in the pathophysiology of a number of disease
states including: 1) cardiovascular
diseases (vasospasm, hypertension, myocardial ischemia; reperfusion injury and
acute myochardial infarction, stroke
(cerebral ischemia), congestive heart failure, shock, atherosclerosis,
vascular chickening); 2) kidney disease (acute
and chronic renal failure, glomeruionephritis, cirrhosis); 3) lung disease
(bronchial asthma, pulmonary hypertension);
4) gastrointestinal disorders (gastric ulcer, inflammatory bowel diseases); 5)
reproductive disorders (premature labor,
dysmenorhea, preeclampsia) and 6) carcinogenesis. Rubanyi & Polokoff, supra.
SSIMMARY OF THE INVENTION
1. PRA
Applicants have identified a cDNA clone that encodes a novel polypeptide,
wherein the polypeptide is
designated in the present application as "PR0213".
In one embodimern, the invention provides an isolated nucleic acid molecule
comprising DNA encoding a
PR0213 polypeptide. In one aspect, the isolated nucleic acid comprises DNA
encoding the PR0213 polypeptide
having amino acid residues 1 to 293 of Figure 2 (SEQ ID N0:2), or is
complementary to such encoding nucleic acid
sequence, and remains siably bound to it under at least moderate, and
optionally, under high stringency conditions.
In another embodiment, the invention provides isolated PR0213 polypeptide. In
particular, the invention
provides isolated native sequence PRO213 polypeptide, which in one embodiment,
includes an amino acid sequence
comprising residues 1 to 295 of Figure 2 (SEQ ID N0:2).
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2. PR0274
Applicants have identified a cDNA clone that encodes a novel polypeptide,
wherein the polypeptide is
designated, in the present application as " PR0274".
In one embodiment, the invention provides an isolated nucleic acid molecule
comprising DNA encoding a
PR0274 polypeptide. In one aspect, the isolated nucleic acid comprises DNA
encoding the PR0274 polypeptide
having amino acid residues 1 to 492 of Figure 4 (SEQ ID N0:7), 'or is
complementary to such encoding nucleic acid
sequence, and remains stably bound to it under at least moderate, and
optionally, under high stringency conditions.
The isolated nucleic acid sequence may comprise the cDNA insert of the
DNA.39987-1184 vector deposited on April
21, 1998 as ATCC 209786 which includes the rntcleotide sequence encoding
fRO274.
In another embodiment, the invention provides isolated PR0274 polypeptide. In
particular, the invention
provides isolated native sequence PR02?4 polypepude, which in one embodiment,
includes an amino acid sequence
comprising residues 1 to 492 of Figure 4 (SEQ ID N0:7). An additional
embodiment of the present invention is
directed to an isolated extracellular domain of a PR0274 polypeptide.
Optionally, the PRO274 polypeptide is
obtained or is obtainable by expressing the polypeptide encoded by the cDNA
insert of the DNA39987-1184 vector
deposited on April 21, 1998 as ATCC 209786.
IS In another embodiment, the invention provides three expressed sequence tags
(EST) comprising the
nucleotide sequences of SEQ ID N0:8 (herein designated as DNAI?873), SEQ ID
N0:9 (herein designated as
DNA36I57) and SEQ ID NO:10 (herein designated as DNA28929) (see Figure S 7,
respactively).
3. PR030Q
Applicants have identified a cDNA clone that encodes a novel polypeptide,
wherein the polypeptide is
designated in the present application as "PR0300"
In one embodiment; the invention provides an isolated nucleic acid molecule
comprising DNA encoding a
PR0300 polypeptide. In one aspect, the isolated nucleic acid comprises DNA
encoding the PR0300 polypeptide
having amino acid residues I to 457 of Figure 9 (SEQ II) N0:19), or is
complementary to such encoding nucleic acid
ZS sequence, and remains stably bound to it under at least moderate, and
optionally, under high stringency conditions.
Tlbe isolated nucleic acid sequence may comprise the cDNA insert of the
DNA4062S-I 189 vector deposited on April
21; 1998 as ATCC 209788 which includes the nucleotide sequence encoding
PR0300.
In another embodiment, the invention provides isolated PR0300 polypeptide. In
particular, the invention
provides isolated native sequence PR0300 polypeptide, which in one embodiment,
includes an amino acid sequence
comprising residues 1 to 4S7 of Figure 9 (SEQ ID N0:19). An additional
embodiment of the present invention is
directed to an isolated extracellular domain of a PR0300 polypeptide.
Optionally, the PR0300 polypeptide is
obtained or is obtainable by expressing the polypeptide encoded by the cDNA
insert of the DNA4062S-I 189 vector
deposited on April 21, 1998 as ATCC 20978$.
3S 4.
Applicants have identified a cDNA clone that encodes a novel transmembrane
polypeptide, wherein the
polypepiide is designated in the present application as °PR0284".
in one embodiment, the invention provides an isolated nucleic acid molecule
comprising DNA encoding a
PR0284 polypeptide. In one aspect, the isolated nucleic acid comprises DNA
encoding the PR0284 polypeptide
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having amino acid residues 1 to 285 of Figure 11 (SEQ ID N0:28), or is
complementary to such encoding nucleic
acid sequence, and remains stably bound to it under at least moderate, and
optionally, under high stringency
conditions. In other aspects, the isolated nucleic acid comprises DNA encoding
the PR0284 polypeptide having
amino acid residues about 25 to 285 of Figure 11 (SEQ ID N0:28) or I or about
25 to X of Figure 11 (SEQ ID
N0:28), where X is any amino acid from 71 to 80 of Figure 11 (SEQ 1D N0:28),
or is complementary to such
encoding nucleic acid sequence, and remains stably bound to it under at least
moderate, and optionally, under high
stringency conditions. The isolated nucleic acid sequence may comprise the
cDNA insert of the DNA23318-1211
vector deposited on April 21, 1998 as ATCC 209787 which includes the
nucleotide sequence encoding PR0284.
In another embodiment, the invention provides isolated PR0284 polypeptide. In
particular, the invention
provides isolated native sequence PR0284 polypeptide, which in one embodiment,
includes an amino acid sequence
comprising residues 1 to 285 of Figure I I {SEQ ID N0:28). Additional
embodiments of the present invention are
directed to isolated PR0284 polypeptides comprising amino acids about 25 to
285 of Figure 11 (SEQ ID N0:28) or
1 or about 25 to X of Figure 11 (SEQ ID N0:28), where X is any amino acid from
71 to 80 of Figure I 1 (SEQ 1D
N0:28}. Optionally, the PR0284 polypeptide is obtained or is obtainable by
expressing the polypeptide encoded by
the cDNA insert of the DNA23318-I21 I vector deposited on April 21, 1998 as
ATCC 209787.
In another embodiment, the invention provides an expressed sequence tag (EST)
designated herein as
DNA12982 which comprises the nucleotide sequence of SEQ ID N0:29.
In another embodiment, the invention provides an expressed sequence tag (EST)
designated herein as
DNA15886 which comprises the nucleotide sequence of SEQ ID N0:30.
5. PR0296
Applicants have identified a cDNA clone that encodes a novel polypeptide
having homology to the sarcoma-
amplified protein SAS, wherein the polypeptide is designated in the present
application as "PR0296".
In one embodiment, the invention provides an isolated nucleic acid molecule
comprising DNA encoding a
PR0296 polypeptide. In one aspect, the isolated nucleic acid comprises DNA
encoding the PR0296 polypeptide
having amino acid residues 1 to 204 of Figure 15 (SEQ ID N0:36}, or is
complementary to such encoding nucleic
acid sequence, and remains stably bound to it under at least moderate, and
optionally, under high stringency
conditions. In other aspects, the isolated nucleic acid comprises DNA encoding
the PR029b polypeptide having
amino acid residues about 35 to 204 of Figure 15 (SEQ ID NO:36) or amino acid
1 or about 35 to X of Figure 15
(SEQ 1D N0:36), where X is any amino acid from 42 to 51 of Figure 15 (SEQ ID
N0:36), or is complementary to
such encoding nucleic acid sequence, and remains stably bound to it under at
least moderate, and optionally, under
high stringency conditions. The isolated nucleic acid sequence may comprise
the cDNA insert of the DNA39979-
12I3 vector deposited on April 21, 1998 as ATCC 209789 which inchtdes the
nucleotide sequence encoding PR0296.
In another embodi~nt, the invention provides isolated PR0296 polypeptide. In
particular, the invention
provides isolated native sequence PR0296 polypeptide, which in one embodiment,
includes an amino acid sequence
comprising residues 1 to 204 of Figure 15 (SEQ ID NO:36). Additional
embodiments of the present invention are
directed to PR0296 polypeptides comprising amino acids about 35 to 204 of
Figure 15 (SEQ ID N0:36) or amino
acid 1 or about 35 to X of Figure 15 (SEQ ID NO:36), where X is any amino acid
from 42 to 51 of Figure 15 (SEQ
ID N0:36). Optionally, the PRO296 polypeptide is obtained or is obtainable by
expressing the polypeptide encoded
by the cDNA insert of the DNA39979-1213 vector deposited on April 21, 1998 as
ATCC 209789.
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In another embodiment, the invention provides an expressed sequence tag (EST)
designated herein as
DNA23020 comprising the rntcieotide sequence of SEQ ID N0:37.
In another embodiment, the invention provides an expressed sequence tag (EST)
designated herein as
DNA21971 comprising the nucleotide sequence of SEQ ID N0:38.
In another embodiment, the invention provides an expressed sequence tag (ES'17
designated herein as
DNA29037 comprising the nucleotide sequence of SEQ ID N0:39.
6. P]It0329
Applicants have identified a cDNA clone that encodes a novel polypeptide
having homology to a high affinity
immunoglobulin F~ receptor, wherein the polypeptide is designated in the
present application as "PR0329" .
In one embodiment, the invention provides an isolated nucleic acid molecule
comprising DNA encoding a
PR0329 polypeptide. In one aspect, the isolated nucleic acid comprises DNA
encoding the PR0329 polypeptide
having amino acid residues 1 to 359 of Figure 20 (SEQ ID N0:45), or is
complementary to such encoding nucleic
acid sequence, and remains stably bound to it under at least moderate, and
optionally, under high stringency
conditions. The isolated nucleic acid sequence may comprise the cDNA insert of
the DNA40594-1233 vector
deposited on February 5, 1998 as ATCC 209617 which includes the nucleotide
sequence encoding PR0329.
In another embodiment; the invention provides isolated PR0329 polypeptide. In
particular, the invention
provides isolated native sequence PR0329 polypeptide, which in one embodiment,
includes an amino acid sequence
comprising residues I to 359 of Figure 20 (SEQ ID N0:45). Optionally, the
PR0329 polypeptide is obtained or is
obtainable by expressing the polypeptide encoded by the cDNA insert of the
DNA40594-1233 vector deposited on
February 5, 1998 as ATCC 209617.
7. PR0362
Applicants have identified a cDNA clone that encodes a novel polypeptide
having hotrwlogy to A33 antigen
and HCAR metnbrane~bo~.uid protein, wherein the polypeptide is designated in
the present application as "PR0362".
1n one embodiment, the invention provides an isolated nucleic acid molecule
comprising DNA encoding a
PR0362 polypeptide. In one aspect, the isolated nucleic acid comprises DNA
encoding the PR0362 polypeptide
having amino acid residues 1 to 321 of Figure 22 (SEQ ID N0:52), or is
complementary to such encoding nucleic
acid sequence, and remains stably bound to it under at least moderate, and
optionally, under high stringency
conditions. In another aspect, the isolated nucleic acid comprises DNA
encoding the PR0362 polypeptide having
amino acid residues 1 to X of Figure 22 (SEQ ID N0:52) when X is any amino
acid from amino acid 271 to 280,
or is compn~ry to such encoding nt~leic acid sequence, and. remains stably
bound to it under at least moderate,
and o~ionally, utzder high stringency conditions. The isolated nucleic acid
sequence may comprise the cDNA insert
of the DNA45416-1251 vector deposited on February 5 ~ 1998 as ATCC 209620
which includes: the nucleotide
sequence encoding PR0362.
In another embodiment, the invention provides isolated PR0362 polypeptide. In
particular, the invention
provides isolated native sequence PR0362 polypeptide, which in one embodiment,
includes an amino acid sequence
comQrising residues 1 to 321 of Figure 22 (SEQ Ih N0:52). An additional
embodiment of the present invention is
directed to as isolated extracellular domain of a PR0362 poiypeptide
comprising amino acids 1 to X of the amino
acid sequence shown in Figure 23 (SEQ ID NO:52), wherein X is any amino acid
from amino acid 271 to 280:
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Optionally, the PR0362 polypeptide is obtained or is obtainable by expressing
the polypeptide encoded by the cDNA
insert of the DNA45416-1251 vector deposited on February 5, 1998 as ATCC
209620:
8. PR0363
Applicants have identified a cDNA clone that encodes a novel polypeptide
having homology to the cell
surface receptor protein HCAR, wherein the polypeptide is designated in the
present application as "PR0363".
In one embodiment, the invention provides an isolated nucleic acid molecule
comprising DNA encoding a
PR0363 polypeptide. In one aspect, the isolated nucleic acid comprises DNA
encoding the PR0363 polypeptide
having amino acid residues 1 to 373 of Figure 24 (SEQ ID NO:$9), or is
complementary to such encoding nucleic
acid sequence, and remains stably bound to it under at least moderate, and
optionally, under high stringency
conditions. In another aspect, the isolated nucleic acid comprises DNA
encoding a PR0363 extracellular domain
poIypeptide having amino acid residues 1 to X of Figure 24 (SEQ ID N0:59)
where X is any amino acid from amino
acid 216 to amino acid 225, or is complementary to such encoding nucleic acid
sequence, and remains stably bound
to it tinder at least moderate, and optionally, under high stringency
conditions. The isolated nucleic acid sequence
may comprise the cDNA insert of the DNA45419-1252 vector deposited on February
5, 1998 as ATCC 209616
which includes the nucleotide sequence encoding PR0363.
In another embodiment, the invention provides isolated PR0363 polypeptide. In
particular, the invention
provides isolated native sequence PRO363 polypeptide, which in one embodiment,
includes an amino acid sequence
comprising residues 1 to 373 of Figure 24 (SEQ ID N0:59). An additional
embodiment of the present invention is
directed to an isolated exuacellular domain of a PR0363 polypeptide, wherein
that extracellular domain may
comprise amino acids 1 to X of the sequence shown in Figure 24 (SEQ ID N0:59),
where X is any amino acid from
amino acid 216 to 225. Optionally, the PR0363 polypeptide is obtained or is
obtainable by expressing the
polypeptide encoded by the cDNA insert of the DNA45419-1252 vector deposited
on February 5, 1998 as ATCC
209616.
9. PR
Applicants have identified a cDNA clone that encodes a novel poIypeptide
having homology to tumor
necrosis factor receptor, wherein the polypeptide is designated in the present
application as "PR0868".
In one embodiment, the invention provides an isolated nucleic acid molecule
comprising DNA encoding a
PR0868 polypeptide. In one aspect, the isolated nucleic acid comprises DNA
encoding the PR0868 polypeptide
having amino acid residues l to 655 of Figure 26 (SEQ ID N0:64), or is
complementary to such encoding nucleic
acid sequence, and remains stably bound to it under at least moderate, and
optionally, under high stringency
conditions. In another aspect, the isolated nucleic acid comprises DNA
encoding the PR0868 polypeptide having
amino acid residues 1 to X of Figure 26 (SEQ ID N0:64), where X is any amino
acid from amino acid 343 to 352
of the sequence shown in Figure 26 (SEQ ID N0:64), oz is complementary to such
encoding nucleic acid sequence,
and remains stably bound to it under at least moderate, and optionally, under
high suiagency conditions. In yet
another aspect, the isolated nucleic acid comprises DNA encoding the PR0868
polypeptide having amino acid
residues X to 655 of Figure 26 (SEQ ID N0:64), where X is arty amino acid from
amino acid 371 to 380 of the
sequence shown in Figure 26 (SEQ ID N0:64), or is complementary to such
encoding nucleic acid sequence, and
remains stably bound to it under at least moderate, and optionally, under high
stringency conditions. The isolated
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rntcleic acid sequence may comprise the cDNA insert of the DNAS2594-1270
vector deposited on March 17, 1998
as ATCC 209679 which includes the nucleotide sequence encoding PR0868.
In another embodiment, the invention provides isolated PR0868 potypeptide. In
particular; the invention
provides isolated native sequence PR0868 poiypeptide, which in one embodiment,
includes an amino acid sequence
compzising residues I to 655 of Figure 26 (SEQ ID N0:64). In another aspect,
the isolated PR0868 polypeptide
S comprises amino acid zesidues 1 to X of Figure 26 ($EQ ID N0:64), where X is
any amino acid from amino acid
343 to 352 of the sequence shown in Figure 26 (SEQ ID NO:b4). In yet another
aspect, the PR0868 poIypeptide
comprises amino acid residues X to 6S5 of Figure 26 (SEQ ID N0:64), where X is
any amino acid from amino acid
371 to 380 of the sequence shown in Figure 26 (SEQ ID N0:64). Optionally, the
PR0868 polypeptide is obtained
or is obtainable by expressing the polypeptide encoded by the cDNA insert of
the DNA52594-1270 vector deposited
on March 17, 1998 as ATCC 209679.
10. PR0382
Applicants have identified a cDNA clone that encodes a novel polypeptide
having homology to serine
proteases, wherein the polypeptide is designated in the present application as
"PR0382".
In one embodiment, the invention provides an isolated nucleic acid molecule
comprising DNA encoding a
PR0382 polypeptide. In one aspect, the isolated nucleic acid comprises DNA
encoding the PRO382 polypeptide
having amino acid residues 1 to 453 of Figure 28 (SEQ ID N0:69), or is
complementary to such encoding nucleic
acid sequence, and remains stably bound to it under at least moderate, and
optionally, under high stringency
conditions. The isolated nucleic acid sequence may comprise the cDNA insert of
the DNA45234-1277 vector
deposited on March 5, 1998 as ATCC 209654 which includes the nucleotide
sequence encoding PR0382.
In another embodiment, the invention provides isolated PR0382 poIypeptide. In
particular, the invention
provides isolated native sequence PR0382 polypeptide, which in one embodiment,
includes an amino acid sequence
comprising residues 1 to 453 of Figure 28 (SEQ ID N0:69). An additional
embodimem of the present invention is
directed to an isolated extracellular domain of a PR0382 polypeptide, with or
without the signal peptide. Optionally,
the PR0382 polypeptide is obtained or is obtainable by expressing the
polypeptide encoded by the cDNA insert of
the DNA45234-1277 vector deposited on March 5; 1998 as ATCC 209654.
11. PROS
Applicants have identified a cDNA clone that encodes a novel polypeptide
having homology to meltrin,
wherein the polypeptide is designated in the present application as "PR054S".
In one embodiment, the invention provides an isolated.nucleic acid molecule
comprising DNA encoding a
PR0545 polypeptide. In one aspect, the isolated nucleic acid comprises DNA
encoding the PR0545 polypeptide
having amino acid residues 1 to 735 of Figure 30 (SEQ ID N0:74), or is
complementary to such encoding nucleic
acid sequence, and remains stably bound to it under at least moderate, and
optionally, under high stringency
conditions, The isolated nucleic acid sequence may comprise the cDNA insert of
the vector deposited on March 5,
1998 as ATCC 209655 which includes the nucleotide seguence encoding PR0545.
In another embodiment, the invention provides isolated PR0545 polypeptide. In
particular, the invention
pm'v~es isolated Native sequence PR0545 polypeptide, which in o~ embodiment,
includes an amino acid sequence
uprising residues 1 to 735 of Figure 30 (SEQ ID N0:74). An additional
embodiment of the present invention is
CA 02421319 2003-03-25
WO 99!46281 PCT/US99/05028
directed to an isolated extracellular domain of a PRO545 polypeptide.
Optionally, the PR0545 polypeptide is
obtained or is obtainable by expressing the polypeptide encoded by the cDNA
insert of the vector deposited on March
5, 1998 as ATCC 209655.
In another embodiment, the invention provides an expressed sequence tag (EST)
designated herein as
DNA13217 comprising the nucleotide sequence of SEQ ID N0:75 (Figure 31).
I2. P~061?
Applicants have identified a cDNA clone that encodes a novel polypeptide
having homology to CD24,
wherein the polypeptide is designated in the present application as "PR06i7".
In one embodiment, the invention provides an isolated nucleic acid molecule
comprising DNA encoding a
PR0617 polypeptide. In one aspect, the isolated nucleic acid comprises DNA
encodutg the PR0617 polypeptide
having amino acid residues 1 to 67 of Figure 33 (SEQ ID N0:85), or is
complementary to such encoding nucleic acid
sequence, and remains stably bound to it under at least moderate, and
optionally, under high stringency conditions.
The isolated nucleic acid sequence may comprise the cDNA insert of the
DNA48309-1280 vector deposited on March
5, 1998 as ATCC 209656 which includes the nucleotide sequence encoding PR0617.
In another embodiment, the invention provides isolated PR0617 polypeptide. In
particular, the invention
provides isolated native sequence PR06I7 polypeptide, which in one embodiment,
includes an amino acid sequence
comprising residues 1 to 67 of Figure 33 (SEQ ID N0:85). Optionally, the
PR0617 polypeptide is obtained or is
obtainable by expressing the polypeptide encoded by the cDNA insert of the
DNA48309-1280 vector deposited on
March 5, 1998 as ATCC 209656.
I3. PR070o
Applicants have identified a cDNA clone that encodes a novel polypeptide
having sequence similarity to
protein disulfide isomerase, wherein the polypeptide is designated in the
present application as "PR0700" .
In one embodiment, the invemion provides an isolated nucleic acid molecule
comprising DNA encoding a
PR0700 polypeptide. In one aspect, the isolated nucleic acid comprises DNA
encoding the PR0700 polypeptide
having amino acid residues 1 to 432 of Figure 35 (SEQ ID N0:90), or is
complementary to such encoding trucleic
acid sequence, and remains stably bound to it under at least moderate, and
optionally, under high stringency
conditions. In another aspect, the isolated nucleic acid comprises DNA
encoding the PR0700 polypeptide having
amino acid residues from about 34 to 432 of Figure 35 (SEQ ID NO:90), or is
complementary to such encoding
ntxleic acid sequerbce, and remains stably bound to it under at least
moderate, and optionally, under high stringency
conditiwns. The isolated tntcleic acid sequence may comprise the cDNA insert
of the vector deposited on March 3I,
1998 as ATCC 209?21 which includes the nucleotide sequence etxoding PR0700.
In another embodiment, the invention provides isolated PR0700 polypeptide. In
particular, the invention
provides isolated native sequence PR0700 polypeptide, which in one embodiment,
includes an amino acid sequence
comprising residues 1 to 432 of Figure 35 (SEQ ID N0:90). In another
embodiment, the invention provides an
isolated PR0700 polypeptide absent the signal sequence, which includes an
amino acid sequence comprising residues
fra~at about 34 to 432 of Figure 35 (SEQ B? N0:90}. Optionally, the PR0700
polypeptide is obtained or is obtainable
by expressing the poiypeptide encoded by the cDNA insert of the vector
deposited on March 31, 1998 as ATCC
209721.
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I4. R 0
Applicants have identified a cDNA clone that encodes a novel polypeptide
having homology to conglutinin,
wherein the polypeptide is designated in the present application-as "PR0702".
In one embodiment, the invention provides an isolated nucleic acid molecule
comprising DNA encoding a
PR0702 polypeptide. In one aspect, the isolated nucleic acid comprises DNA
encoding the PR0702 polypeptide
having amino acid residues I to 277 of Figure 37 (SEQ ID N0:97), or is
complementary to such encoding nucleic
acid sequence, and remains stably bound to it under at least moderate, and
optionally, under high stringency
conditions. In another aspect, the isolated nucleic acid comprises DNA
encoding the PR0702 polypeptide having
amino acid residues 26 to 277 of Figure 37 (SEQ ID N0:97), or is complementary
to such encoding nucleic acid
sequence, and remains stably bound to it under at least moderate, and
optionally, under high stringency conditions.
The isolated nucleic acid sequence may comprise the cDNA insert of the
DNA50980-1286 vector deposited on March
31, 1998 as ATCC 209717 which includes the nucleotide sequence encoding
PR0702.
1n another embodiment, the invention provides isolated PR0702 polypeptide. In
particular, the invention
provides isolated native sequence PR0702 polypeptide, which in one embodiment,
includes an amino acid sequence
comprising residues 1 to 277 of Figure 37 (SEQ ID N0:97), An additional
embodiment of the present invention is
directed to an isolated PR0702 polypeptide comprising amino acid residues 26
to 277 of Figure 37 (SEQ ID N0:97).
Optionally, the PR0702 polypeptide is obtained or is obtainable by expressing
the polypeptide encoded by the cDNA
insert of the DNA50980-1286 vector deposited on March 3i, 1998 as ATCC 209717.
15. PRO'103
Applicants have identified a cDNA clone that encodes a novel polypeptide
having sequence similarity to
VLCAS, wherein the polypeptide is designated in tire present application as
"PR0703".
In one embodiment, the invention provides an isolated nucleic acid molecule
comprising DNA encoding a
PR0703 polypeptide. In one aspect, the isolated nucleic acid comprises DNA
encoding the PRO?03 polypeptide
having amino acid residues 1 to 730 of Figure 39 (SEQ ID N0:102), or is
complementary to such encoding nucleic
acid sequence, and remains stably bound to it under at least moderate, and
optionally, under high stringency
conditions. In another aspect, the isolates! nucleic acid comprises DNA
encoding the PR0703 polypeptide having
amino acid residues from about 43 to 730 of Figure 39 (SEQ ID NO:102), or is
complementary to such encoding
nucleic acid sequence, and remains stably bound to ~it under at least
moderate, and optionally, under high stringency
conditions. The isolated nucleic acid sequence may comprise the cDNA insert of
the DNA50913-1287 vector
deposited on March 31, 1998 as ATCC 209716 which includes the nucleotide
sequence encoding PR0703.
In another embodiment, the invention provides isolated PRO703 polypeptide. In
particular, the invention
provides isolated native sequence PR0703 polypeptide, which in one embodiment,
includes an amino acid sequence
comprising residues l to 730 of Figure 39 (SEQ ID NO:I02}. In another
embodiment, the invention provides an
isolated PR0703 polypeptide absent the signal sequence, which includes an
amino acid sequence comprising residues
from about 43 to 730 of Figure 30 (SEQ ID NO:I02). Optionally, the PR0730
polypeptide is obtained or is
obtainable by expressing the polypeptide encoded by the cDNA insert of the
DNA50913-1287 vector deposited on
March 31, 1998 as ATCC 209716.
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16. PR0705
Applicants have identified a cDNA clone that encodes a novel poiypeptide
having homology to K-glypican,
wherein the polypeptide is.designated in the present application as "PR0705".
In one embodiment, the invention provides an isolated nucleic acid molecule
comprising DNA encoding a
PR0705 polypeptide. In one aspect, the isolated nucleic acid comprises DNA
encoding the PR0705 polypeptide
S having amino acid residues 1 to 555 of Figure 41 (SEQ ID NO:109), or is
complementary to such encoding nucleic
acid sequence, and remains stably bound to it under at least moderate, and
optionally, under high stringency
conditions. In another aspect, the isolated nucleic acid comprises DNA
encoding the PR0705 polypeptide having
amino acid residues about 24 to 555 of Figure 41 (SEQ ID N0:109), or is
complementary to such encoding nucleic
acid sequence, and remains stably bound to it under at least moderate, and
optionally, under high stringency
conditions. The isolated nucleic acid sequence may comprise the cDNA insert of
the DNA50914-1289 vector
deposited on March 31, 1998 as ATCC 209722 which includes the nucleotide
sequence encoding PR0705.
In another embodiment, the invention provides isolated PR0705 polypeptide. In
particular, the invention
provides isolated native sequence PR0705 polypeptide, which in one embodiment,
includes an amino acid sequence
comprising residues 1 to 555 of Figure 41 {SEQ ID N0:109). An additional
embodiment of the present invention
is directed to an isolated PR0705 polypeptide comprising amino acid residues
about 24 to 555 of Figure 41 (SEQ
ID N0:109). Optionally, the PR0705 polypeptide is obtained or is obtainable by
expressing the polypeptide encoded
by the cDNA insert of the DNA50914-1289 vector deposited on March 31, 1998 as
ATCC 209722.
17. PR0708
Applicants have identified a cDNA clone that encodes a novel polypeptide
having homology to the aryl
sulfatases, wherein the polypeptide is designated in the present application
as "PRO708".
In one embodiment, the invention provides an isolated nucleic acid molecule
comprising DNA encoding a
PR0708 polypeptide. In one aspect, the isolated nucleic acid comprises DNA
encoding the PR0708 polypeptide
having amino acid residues 1 to 515 of Figure 43 (SEQ ID NO:I14), or is
complementary to such encoding nucleic
acid sequence, and remains stably bound to it under at least moderate, and
optionally, under high stringency
conditions. The isolated nucleic acid sequence may comprise the cDNA insert of
the DNA48296-1292 vector
deposited on March 11, 1998 as ATCC 209668 which includes the nucleotide
sequence encoding PR0708.
In another embodiment, the invention provides isolated PRO?08 polypeptide. In
particular, the invention
provides isolated native sequence PR0708 polypeptide, which in one embodiment,
includes an amino acid sequence
comprising residues 1 to 5I5 of Figure 43 (SEQ ID N0:114). Another embodiment
is directed to a PR0708
polypeptide comprising residues 38-515 of the amino acid .sequence shown in
Figure 43 (SEQ ID N0:114).
Optionally, the PR0708 polypeptide is obtained or is obtainable by expressing
the polypeptide encoded by the cDNA
insert of the DNA48296-1292 vector deposited on March 11, 1998 as ATCC 209668.
I8. ~R0320
Applicants have identified a cDNA clone that encodes a novel polypeptide
having homology to fibulin,
wherein the polypeptide is designated in the present application as "PR0320".
In one embodiment, the invention provides an isolated nucleic acid molecule
comprising DNA encoding a
PR0320 polypeptide. In one aspect, the isolated nucleic acid comprises DNA
encoding the PR0320 polypeptide
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having amino acid t~esidues I to 338 of Figure 45 (SEQ ID N0:119), or is
complementary to such encoding nucleic
acid sequence, and remains stably bound to it under at least moderate, and
optionally, under high stringency
conditions. The isolated nucleic acid sequence rnay comprise the cDNA insert
of the vector deposited on March 1 I,
1998 as ATCC 209670 which includes the nucleotide sequence encoding PR0320.
In another embodiment, the invention provides isolated PR0320 polypeptide. In
particular, the invention
provides isolated native sequence PR0320 polypeptide, which in one embodiment,
includes an amino acid sequence
comprising residues I to 338 of Figure 45 (SEQ ID N0:119). Optionally, the
PR0320 polypeptide is obtained or
is obtainable by expressing the polypeptide encoded by the cDNA insert of the
vector deposited on March 11, 1998
as ATCC 209670.
19. PR 3 4
Applicants have identified a cDNA clone that encodes a novel polypeptide
having homology to
oxidoreductases, wherein the polypeptide is designated in the present
application as "PR0324".
In one embodiment, the invention provides an isolated ztucleic acid molecule
comprising DNA encoding a
PR0324 polypeptide. In one aspect, the isolated nucleic acid comprises DNA
encoding the PR0324 polypeptide
having amitw acid residues 1 to 289 of Figure 47 (SEQ ID N0:124), or is
complementary to such encoding nucleic
acid sequence, and remains stably bound to it under at least moderate, and
optionally, under high stringency
conditions. In other aspects, the isolated nucleic acid comprises DNA encoding
the PR0324 polypeptide having
amino acid residues 1 or about 32 to X of Figure 47 (SEQ ID N0:124), where X
is any amino acid from 131 to 140,
or is complementary to such encoding nucleic acid sequence, and remains stably
bound to it under at least moderate,
and optionally, under high stringency conditions. The isolated nucleic acid
sequence may comprise the cDNA insert
of the DNA36343-1310 vector deposited on March 30, 1998 as ATCC 209718 which
includes the nucleotide sequence
erncoding PR0324.
1n another embodiment, the invention provides isolated PR0324 polypeptide. In
particular, the invention
provides isolated native sequence PR0324 poiypeptide, which in one embodiment,
inchtdes an amino acid sequence
comprising residues 1 to 289 of Figure 47 (SEQ ID N0:124). The invention also
provides isolated PR0324
polypeptide comprising residues I or about 32 to X of Figure 47 (SEQ ID
N0:124}, wherein X is any amino acid
from about 131-140. Optionally, the PR0324 polypeptide is obtained or is
obtainable by expressing the polypeptide
encoded by the cDNA insert.of the DNA36343-1310 vector deposited on March 30,
1998 as ATCC 209718.
20. pR0351
Applicants have identified a cDNA clone that encodes a novel polypeptide
having sequence similarity to
prostasin, wherein the polypeptide is designated in the present application as
"PR0351".
In one en>bodiment, the invention provides an isolated nucleic acid molecule
comprising DNA et~oding a
PR0351 polypeptide. In one aspect, the isolated nucleic acid comprises DNA
encoding the PR0351 polypeptide
having amino acid residues 1 to 571 of Figure 49 (SEQ ID N0:132), or is
complementary to such encoding nucleic
acid sequence, and remains stably bound to it under at least moderate, and
optionally, under high stringency
conditions. In another aspect, the isolated nucleic acid comprises DNA
encoding the PRO351 polypeptide having
amino acid residues about 16 to 571 of Figure 49 (SEQ ID N0:132), or is
complementary to such encoding nucleic
acid sequence, and remains stably bound to it under at least moderate, and
optionally, under high stringency
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conditions. The isolated nucleic acid sequence may comprise the cDNA insert of
the DNA40571-1315 vector
deposited on April 21, 1998 as ATCC 209784 which includes the nucleotide
sequence encoding PR0351.
In another embodiment, the invention provides isolated PR0351 polypeptide. In
particular, the invention
provides isolated native sequence PR0351 polypeptide, which in one embodiment,
includes an amino acid sequence
comprising residues 1 to 571 of Figure 49 (SEQ ID N0:132). In another
embodiment, the invention provides an
isolated PR0351 polypeptide absent the signal sequence, which includes an
amino acid sequence comprising residues
from about I6 to 5?I of Figure 49 (SEQ ID N0:132), Optionally, the PR0351
polypeptide is obtained or is
obtainable by expressing the polypeptide encoded by the cDNA insert of the
DNA40571-1315 vector deposited on
April 21, 1998 as ATCC 209784.
21. PR0352
Applicants have identified a cDNA clone that encodes a novel polypeptide
having homology to butyrophilin,
wherein the polypeptide is designated in.the present application as "PR0352".
In one embodiment, the invention provides an isolated nucleic acid molecule
comprising DNA encoding a
PR0352 polypeptide. In one aspect, the isolated nucleic acid comprises DNA
encoding the PR0352 polypeptide
having atnitto acid residues 1 to 316 of Figure 51 (SEQ ID NO:137), or is
complementary to such encoding nucleic
acid sequence, and remains stably bound to it under at least moderate, and
optionally, under high stringency
conditions. in other aspects, the isolated nucleic acid comprises DNA encoding
the PR0352 polypeptide having
amino acid residues of about 29 to 316 of Figure 51 (SEQ ID N0:137), or 1 or
about 29 to X of Figure 51, where
X is any amino acid from 246 to 255, or is complementary to such encoding
nucleic acid sequence, and remains
stably bound to it under at least moderate, and optionally, under high
stringency conditions. The isolated nucleic acid
sequence may comprise the cDNA insert of the DNA41386-1316 vector deposited on
March 26, 1998 as ATCC
209703 which includes the nucleotide sequence encoding PR0352.
In another embodiment, the invention provides isolated PR0352 polypeptide. In
particular, the invention
provides isolated native sequence PR0352 polypeptide, which in one embodiment,
includes an amino acid sequence
comprising residues 1 to 316 of Figure 51 (SEQ ID N0:137). In other
embodiments, the invention provides isolated
PR0352 polypeptide comprising residues about 29 to 316 of Figure 51 (SEQ ID
NO:I37) and 1 or about 29 to X of
Figure 51 (SEQ ID NO:I37), wherein X is any amino acid from 246 to 255.
Optionally, the PR0352 polypeptide
is obtained or is obtainable by expressing the polypeptide encoded by the cDNA
insert of the DNA41386-1316 vector
deposited on March 26, 1998 as ATCC 209703.
22. PR0381
Applies have identified a cDNA clone that encodes a novel poiypeptide having
homology to immunophilin
proteins, wherein the polypeptide is designated in the present application as
"PR0381".
In one embodiment, the invention provides an isolated nucleic acid molecule
comprising DNA encoding a
PR0381 polypeptide. In one aspect, the isolated nucleic acid comprises DNA
encoding the PR0381 polypeptide
having amino acid residues 1 to 211 of Figure 53 (SEQ ID N0:145), or is
complementary to such encoding nucleic
acid sequence, and remains stably bound to it under at least moderate, and
optionally, under high stringency
conditions. In another aspect, the isolated nucleic acid comprises DNA
encoding the PR0381 polypeptide having
amino acid residues about 21 to 211 of Figure 53 (SEQ ID N0:145), or is
complementary to such encoding nucleic
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acid sequence, and remains stably bound to it under at /east moderate, and
optionally, under high stringency
conditions. The isolated nucleic acid sequence may comprise the cDNA insert of
the DNA44194-1317 vector
deposited on April 28, 1998 as ATCC 209808 which includes the nucleotide
sequence encoding PR0381.
In another embodiment, the invention provides isolated PR0381 polypeptide. In
particular; the invention
provides isolated native sequence PRO381 poiypeptide, which in one embodiment,
includes an amino acid sequence
comprising residues 1 to 211 of Figure 53 (SEQ ID N0:145). Another embodiment
is directed to a PR0381
plypeptide comprising amino acids about 21 to 211 of Figure 53 (SEQ ID
N0:145). Optionally; the PR0381
polypeptide is obtained or is obtainable by expressing the polypeptide encoded
by the cDNA insert of the DNA44194-
1317 vector deposited on April 28, 1998 as ATCC 209808.
23. P~0386
Applicants have identified a cDNA clone that encodes a novel polypeptide
having homology to the beta-2
subunit of a soditun channel, wherein the polypeptide is designated in the
present application as "PR0386".
In one embodiment, the invention provides an isolated nucleic acid molecule
comprising DNA encoding a
PR0386 polypeptide. In one aspect, the isolated nucleic acid comprises DNA
encoding the PR0386 polypeptide
having amino acid residues 1 to 215 of Figure 55 (SEQ ID NO:150), or is
complementary to such encoding nucleic
acid sequence, and remains stably bound to it under at least moderate, and
optionally, under high stringency
conditions. In another aspect, the isolated nucleic acid comprises DNA
encoding the PR0386 polypeptide having
amino acid residues about 21 to 215 of Figure 55 (SEQ ID NO:150) or 1 or about
21 to X, where X is any amino
acid from 156 to 165 of Figure 55 (SEQ ID N0:150), or is complementary to such
encoding nucleic acid sequence,
and remains stably bound to itunder at least moderate, and optionally, under
high stringency conditions. The isolated
nucleic acid sequence may comprise the eDNA insert of the DNA45415-1318 vector
deposited on April 28, 1998 as
ATCC 209810 which includes the nucleotide sequence encoding PR0386.
In another embodiment, the invention provides isolated PR0386 polypeptide. In
particular, the invention
provides isolated native sequence PR0386 polypeptide, which in one embodiment,
includes an amino acid sequence
comprising residues 1 to 215 of Figure 55 (SEQ ID N0:150). Other embodiments
of the present invention are
directed to PR0386 polypeptides comprising amino acids about 21 to 215 of
Figure 55 (SEQ ID NO:150) and 1 or
about 21 to X of Figure 55 (SEQ ID N0:150), wherein X is airy amino acid from
156 to 165 of Figure 55 (SEQ ID
NO:150). Optionally, the PR0386 polypeptide is obtained or is obtainable by
expressing the polypeptide encoded
by the cDNA insert of the DNA45415-1318 vector deposited on April 28, 1998 as
ATCC 209810.
In attori~r etttbodiment, the invention provides an expressed sequence tag
(EST) comprising the nucleotide
sequence of SEQ ID NO:151 which corrsponds to an EST designated herein as
DNA23350.
In another embodiment, the invention provides an expressed sequence tag (EST)
comprising the nucleotide
sequence of SEQ ID NO: 352 which corrsgonds to an EST designated herein as
DNA23S36.
24, pR05d~U_
Applicants have identified a cDNA clone that encodes a novel polypeptide
having sequence similarity to
LCAT, wherein the polypeptide is designated in the present application as
°PR0540".
In one embodiment, the invention provides an isolated nucleic acid molecule
comprising DNA encoding a
PR0540 polypeptide. In one aspect, the isolated nucleic acid comprises DNA
eroding the PR054U polypeptide
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having amino acid residues 1 to 412 of Figure 59 (SEQ ID N0:157), or is
complementary to such encoding nucleic
acid sequence, and remains stably bound to it under at least moderate, and
optionally, under high stringency
conditions. In another aspect, the isolated nucleic acid comprises DNA
encoding the PR0540 polypeptide having
amino acid residues about 29 to 412 of Figure 59 (SEQ 1D N0:157), or is
complementary to such encoding nucleic
acid sequence, and remains stably bound to it under at least moderate, and
optionally, under high stringency
conditions. The isolated nucleic acid sequence may comprise the cDNA insert of
the DNA44189-1322 vector
deposited on March 26, 1998 as ATCC 209699 which includes the nucleotide
sequence encoding PR0540.
In another embodiment, the invention provides isolated PR0540 polypeptide. In
particular, the.invention
provides isolated native sequence PR0540 polypeptide, which in one embodiment,
includes an amino acid sequence
comprising residues I to 412 of Figure 59 (SEQ ID N0:157). The invention also
provides isolated PR0540
polypeptide, which in one embodiment, includes an amino acid sequence
comprising residues about 29 to 412 of
Figure 59 (S$Q ID N0:157}. Optionally, the PR0540 polypeptide is obtained or
is obtainable by expressing the
poiypeptide encoded by the cDNA insert of the DNA44i89-1322 vector deposited
on March 26, 1998 as ATCC
209699.
25. PR0615
Applicants have identified a cDNA clone that encodes a novel polypepdde having
sequence similarity to
synaptogyrin, wherein the polypeptide is designated in the present application
as "PR0615".
In one embodiment, the invention provides an isolated nucleic acid molecule
comprising DNA et~oding a
PR0615 polypeptide. In one aspect, the isolated nucleic acid comprises DNA
encoding the PR0615 polypeptide
having amino acid residues 1 to 224 of Figure 61 (SEQ ID N0:162), or is
complementary to such encoding nucleic
acid sequence, and remains stably bound to it under at least moderate, and
optionally, under high stringency
conditions. In another aspect, the isolated nucleic acid comprises DNA
encoding the PR0615 poiypeptide having
amino acid residues X to 224 of Figure 61 (SEQ ID N0:162), where X is any
amino acid from 157 to 166, or is
complementary to such encoding nucleic acid sequence, and remains stably bound
to it under at least moderate, and
optionally, under high stringency conditions. The isolated nucleic acid
sequence may comprise the cDNA insert of
the DNA48304-1323 vector deposited on April 28, 1998 as ATCC 209811 which
includes the nucleotide sequence
encoding PR0615.
In another embodiment, the invention provides isolated PR0615 polypeptide. In
particular, the invention
provides isolated native sequence PR0615 polypeptide, which in one embodiment,
includes an amino acid sequence
comprising residues 1 to 224 of Figure 61 (SEQ ID N0:162). An additional
embodiment of the present invention
is directed to an isolated exuacellular domain of a PR0615 polypeptide which
comprises amino acid residues X to
224 of Figure 61 (SfiQ ID N0:162), where X is any amino acid from 157 to 166
of Figure 61 (SEQ ID N0:162).
Optionally, the PROtilS polypeptide is obtained or is obtainable by ezpressing
the polypeptide encoded by the cDNA
insert of the DNA48304-1323 vector deposited on April 28, 1998 as ATCC 209$11.
26. PR0618
Applicants have identified a cDNA clone that encodes a novel polypeptide
having sequence similarity to
enteropeptidase, wherein the poIypeptide is designated in the present
application as "PR0618".
In one embodiment, the invention provides an isolated nucieic acid molecuie
comprising DNA encoding a
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PR06I8 polypeptide. In one aspect, the isolated nucleic acid comprises D'NA
encoding the PRO618 polypeptide
having amino acid residues 1 to 802 of Figure 63 (SEQ ID NO:169}, or is
complementary to such encoding nucleic
acid sequence, and remains stably bound to it under at least moderate, and
optionally, under high stringency
conditions. In another aspect, the isolated nucleic acid comprises DNA
encoding an isolated extracellular domain
of a PR0618 polypeptide having amino acid residues X to 802 of Figure 63 (SEQ
ID N0:169), where X is any amino
acid from 63 to 72 of Figure 63 {SEQ ID N0:169), or is complementary to such
encoding nucleic acid sequence, and
remains stably bound to it under at least moderate, and optionally, under high
stringency conditions. The isolated
nucleic acid sequence rnay comprise the cDNA insert: of the DNA49152-1324
vector deposited on April 28, 1998 as
ATCC 209813 which includes the nucleotide sequence encoding PR0618.
In another embodiment, the invention provides isolated PR0618 poiypeptide. In
particular, the invention
provides isolated native sequence PR0618 polypeptide, which in one embodiment,
includes an amino acid sequence
comprising residues 1 to 802 of Figure 63 (SEQ ID N0:169). An additional
embodiment of the present invention
is directed to an isolated extracellular domain of a PR061~8 polypeptide
comprising amino acid X to 802 where X
is any amino acid from 63 to 72 of Figure 63 (SEQ ID N0:169). Optionally, the
PR0618 polypeptide is obtained
or is obtainable by expressing the polypeptide encoded by the cDNA insert of
the DNA49152-1324 vector deposited
on April 28, 1998 as ATCC 209813.
In another emboditnetu, the invention provides an expressed sequence tag (EST)
comprising the nucleotide
sequence of SEQ ID N0:170, designated herein as DNA35597 (see Figure 64).
27. PR0719
Applicarns have identified a cDNA clone that encodes a novel palypeptide
having homology to lipoprotein
lipase H, wherein the polypeptide is designated in the present application as
"PR0719".
In one embodiment, the invention provides an isolated nucleic acid molecule
comprising DNA encoding a
PR0719 polypeptide. In one aspect, the isolated nucleic acid comprises DNA
encoding the PR0719 polypeptide
having amino acid residues 1 to 354 of Figure 66 (SEQ ID N0:178), or is
complementary to such encoding nucleic
acid sequence, and remains stably bound to it under at least moderate, and
optionally, under high stringency
conditions. In another aspect, the isolated nucleic acid comprises DNA
encoding the PR0719 polypeptide having
amino acid residues about 17 to 354 of Figure 66 (SEQ ID N0:178), or as
complementary to such encoding nucleic
acid sequence, and remains stably bound to it under at least moderate, and
optionally, under high stringency
conditions. The isolated nucleic acid sequence may comprise the cl:fNA insert
of the DNA49646-I32? vector
deposited on March 26, 1998 as ATCC 209705 which includes the nucleotide
sequence encoding PR0719.
In another embodiment, the irrvention provides isolated PR0719 polypeptide. 1n
particular, the invention
provides isolated native sequence PR0719 polypeptide, which in one embodiment,
includes an amino acid sequence
comprising residues 1 to 354 of Figure 66 (SEQ ID N0:178). In another
embodiment, the invention provides isolated
PR0719 polypeptide which comprises residues about 17 to 354 of Figure 66 (SEQ
ID N0:178). Optionally, the
3S PR07i9 polypeptide is obtained or is obtainable by expressing the
polypeptide encoded by the eDNA insert of the
DNA49646-1327 vector deposited on March 26, 1998 as ATCC 209705.
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28. RO!724
Applicants have identified a cDNA clone that encodes a novel polypeptide
having homology to the LDL
receptor, wherein the polypeptide is designated in the present application as
"PR0724".
In one embodiment, the invention provides an isolated nucleic acid molecule
comprising DNA encoding a
PR0724 polypeptide, In one aspect, the isolated nucleic acid comprises DNA
encoding the PR0724 polypeptide
having amino acid residues 1 to 7I3 of Figure 68 (SEQ ID NO:I83), or is
complementary to such encoding nucleic
acid sequence, and remains stably bound to it under at least moderate, and
optionally, under high stringency
conditions. In another aspect, the isolated nucleic acid comprises DNA
encoding a soluble PR0724 polypeptide
having amino acid residues 1 to X of Figure 68 (SEQ ID NO: I83) where X is any
amino acid from amino acid' 437
to 446, or is complementary to such encoding nucleic acid sequence, and
remains stably bound to it under at least
10. moderate, and optionally, under high stringency conditions. The above two
polypeptides may either possess or not
possess the signal peptide. The isolated nucleic acid sequence may comprise
the cDNA insert of the DNA49631-1328
vector deposited on April 28, 1998 as ATCC 209806 which includes the
nucleotide sequence encoding PR0724.
In another embodiment, the invention provides isolated PR0724 polypeptide. In
particular, the invention
provides isolated native sequence PR0724 polypeptide, which in one embodiment,
includes an amino acid sequence
comprising residues 1 to 713 of Figure 68 (SEQ ID N0:183). In another
embodiment, the invention provides isolated
soluble PR0724 polypeptide. In particular, the invention provides isolated
soluble PR0724 polypeptide, which in
otte embodiment, inchides an amino acid sequence comprising residues 1 to X of
Figure 68 (SEQ ID N0:183), where
X is any amino acid from 437 to 446 of the sequence shown in Figure 68 (SEQ ID
NO:183). Optionally, the PR0724
polypeptide is obtaita:d or is obtainable by expressing the polypeptide
encoded by the cDNA insert of the DNA49631
1328 vector deposited on April 28, 1998 as ATCC 209806:
29. PR0772
Applicants have identified a cDNA clone that encodes a novel polypeptide
having homology to A4 protein,
wherein the poiypeptide is designated in the present application as "PR0772" .
1n one embodiment, the invention provides an isolated nucleic acid molecule
comprising DNA encoding a
PR0772 polypeptide. In one aspect, the isolated nucleic acid comprises DNA
encoding the PRO772 polypeptide
having amino acid residues 1 to 152 of Figure 70 (SEQ ID N0:190), or is
complementary to such encoding nucleic
acid sequence, and remains, stably bound to it under at least moderate, and
optionally, under high stringency
conditions. In other aspects, the isolated nucleic acid comprises DNA encoding
the,PR0772 polypeptide having
amino acid residues 1 to X of Figure 70 (SEQ ID N0:190), where X is any amino
acid from 21 to 30 of Figure 70
(SEQ )D N0:190), or is complementary to such encoding nucleic acid sequence,
and remains stably bound to it under
at least moderate, and optionally, under high stringency conditions. The
isolated nucleic acid sequence may comprise
the cDNA insert of the DNA49645-1347 vector deposited on April 28, 1998 as
ATCC 209809 which includes the
nucleotide sequence etxoding PR0772.
In another embodiment, the invention provides isolated PR07?2 polypeptide. ht
particular, the invention
provides isolated native sequence PR0772 polypeptide, which in one embodiment,
includes an amino acid sequence
comprising residues 1 to 152 of Figure 70 (SEQ ID N0:190). Additional
embodiments of the present invention are
directed to PRO772 polypeptides comprising amino acids l to X of Figure 70
(SEQ 1D NO:I90), where X is any
amino acid from 21 to 30 of Figure 70 (SEQ ID N0:190). Optionally, the PR0772
polypeptide is obtained or is
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obtainable by expressing the polypeptide encoded by the cDNA insert of the
DNA49645-1347 vector deposited on
April 28, 1998 as ATCC 209809.
In another embodiment, the invention provides an expressed sequence tag (EST)
designated herein as
DNA43509 comprising the nucleotide sequence of SEQ ID N0:191 (Figure 71).
30. PR 8
Applicants have identified a cDNA clone that encodes a novel polypeptide
having homology to various
protease enzymes, wherein the polypeptide is designated in the present
application as "PRO852".
In one embodiment, the invention provides an isolated nucleic acid molecule
comprising DNA encoding a
PR0852 polypeptide. In one aspect, the isolated nucleic acid comprises DIVA
encoding the PR0852 polypeptide
having amino acid residues 1 to 518 of Figure 73 (SEQ ID NO:I96), or is
complementary to such encoding nucleic
acid sequence, and remains stably bound to it under at least moderate, and
optionally, under high stringency
conditions. In other aspects, the isolated nucleic acid comprises DNA encoding
the PR0852 polypeptide having
amino acid residues about 21 to 518 of Figure 73 (SEQ 1D N0:196) or 1 or about
21 to X of Figure 73 (SEQ ID
N0:196) where X is any amino acid from amino acid 461 to amino acid 470 of
Figure 73 (SEQ ID NO:196), or is
complementary to such encoding nucleic acid sequence, and remains stably bound
to it under at least moderate, and
optionally, under high stringency conditions. The isolated nucleic acid
sequence may comprise the cDNA insert of
the DNA45493-1349 vector deposited on April 28, 1998 as ATCC 209805 which
includes the nucleotide sequence
encoding PR0852.
In another embodiment, the invention provides isolated PR0852 polypeptide. In
particular; the invention
provides isolated native sequence PR0852 polypeptide, which in one embodiment,
includes an amino acid sequence
comprising residues 1 to 518 of Figure 73 (SEQ ID N0:196). In other
embodiments, the PR0852 comprises amino
acids about 21 to amino acid 518 of Figure 73 (SEQ ID N0:196) or amino acids 1
or about 21 to X of Figure 73
(SEQ ID N0:196), where X is any amino acid from amino acid 461 to amino acid
470 of Figure 73 (SEQ ID
N0:196). Optionally, the PR0852 polypeptide is obtained or is obtainable by
expressing the poIypeptide encoded
by the cDNA insert of the DNA45493-1349 vectordeposited on April 28" 1998 as
ATGC 209805.
31. PR0853
Applicants have identified a cDNA clone that encodes a novel polypeptide
having sequence similarity to
reductase, wherein the polypeptide is designated in the present application as
"PRO$S3".
In one embodiment, the invention provides an isolated nucleic acid molecule
comprising DNA encoding a
PR0853 polypeptide. In otne aspect, the isolated nucleic acid comprises DNA
encoding the PR0853 polypeptide
having amino acid residues 1 to 377 of Figure 75 (SEQ ID N0:206), or is
complementary to such encoding nucleic
acid sequence, and remains stably bound to it under at least moderate, and
oprionally, under high stringency
conditions. In another aspect, the isolated nucleic acid comprises DNA
encoding the PR0853 polypeptide having
amino acid residues about 17 to 377 of Figure 75 (SEQ ID N0:206), or is
complementary to such encoding nucleic
acid sequence, and remains stably bound to it under at least moderate, and
optionally, under high stringency
conditions. The isolated nucleic acid seguence may comprise the cDNA insert of
the DNA48227-1350 vector
deposited on April 28, 1998 as ATGC 209812 which includes the nucleotide
sequence encoding PR0853.
In another embodimern, the invention provides isolated PR0853 polypeptide. In
particular, the invention
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provides isolated native sequence PR0853 polypeptide, which in one embodiment,
includes an amino acid sequence
comprising residues 1 to 377 of Figure 75 (SEQ ID N0:206). In another
embodiment, the invention provides an
isolated PR0853 golypeptide, absent the signal sequence, which includes an
amino acid sequence comprising residues
from about 17 to 377 of Figure 75 (SEQ ID N0:206). Optionally, the PR0853
polypeptide is obtained or is
obtainable by expressing the polypeptide encoded by the cDNA insert of the
DNA48227-1350 vector deposited on
April 28, 1998 as ATCC 209812.
32. PR0860
Applicants have identified a cDNA clone that encodes a novel polypeptide
having sequence similarity to
neurofascin, wherein the polypeptide is designated in the present application
as "PR0860".
In one embodiment, the invention provides an isolated nucleic acid molecule
comprising DNA encoding a
PR0860 polypeptide. In one aspect, the isolated nucleic acid comprises DNA
encoding the PR0860 polypeptide
having amino acid residues I to 985 of Figure 77 (SEQ ID NO:211), or is
complementary to such encoding nucleic
acid sequence, and remains stably bound to it under at least moderate, and
optionally, under high stringency
conditions. In another aspect, the isolated nucleic acid comprises DNA
encoding the PR0850 polypeptide having
amino acid residues 1 to X of Figure 77 (SEQ ID N0:2I1), where X is any amino
acid from 443-452 of Figure 77
(SEQ ID N0:211), or is complementary to such encoding nucleic acid sequence,
and remains stably bound to it under
at least moderate, and optionally, under high stringency conditions. The
isolated nucleic acid sequence may comprise
the cDNA insert of the DNA41404-1352 vector deposited on May 6, 1998 as ATCC
209844 which includes the
rrucleotide sequence encoding PR0860.
In another embodiment, the invention provides isolated PR0860 polypeptide. In
particular, the invention
provides isolated native sequence PR0860 polypeptide, which in one embodiment,
includes an amino acid sequence
comprising residues 1 to 985 of Figure 77 (SEQ ID N0:211). In another
embodiment, the invention provides an
isolated PR0860 polypeptide which includes an amino acid sequence comprising
residues 1 to X of Figure 77 (SEQ
ID N0:211), where X is any amino acid residue from 443 to 452 of Figure 77
(SEQ ID N0:211). Optionally, the
PR0860 polypeptide is obtained or is obtainable by expressing the polypeptide
encoded by the cDNA insert of the
DNA41404-1352 vector deposited on May 6, 1998 as ATCC 209844.
33. ~R0846
Applicants have identified a cDNA clone that encodes a novel polypeptide
having sequence similarity to
CMRF35, wherein the polypeptide is designated in the present application as
"PR0846".
1n one embodiment, the invention provides an isolated nucleic acid molecule
comprising DNA encoding a
PR0846 polypeptide. In one aspect, the isolated nucleic acid comprises DNA
encoding the PR0846 polypeptide
having amino acid residues 1 to 332 of Figure 79 (SEQ ID N0:216), or is
complementary to such encoding nucleic
acid sequence, and remains stably bound to it under at least moderate, and
optionally, under high stringency
conditions. In other aspects, the isolated nucleic acid comprises DNA encoding
the PR0846 polypeptide having
amhto acid residues about 18 to 332 of I igute 79 (SEQ m N0:216) or 1 or about
18 to X of SEQ ID N0:216, where
X is auy amino acid from 243 to 252 of Figure 79 (SEQ ID N0:216); or is
complementary to such encoding nucleic
acid sequence, and remains stably bound to it under at least moderate, and
optionally, under high stringency
conditions. The isolated nucleic acid sequence may comprise the cDNA insert of
the DNA44196-1353 vector
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deposited on May 6, 1998 as ATCC 209847 which includes the nucleotide sequence
encoding PR0846.
in another embodiment, the invention provides isolated PR0846 polypeptide. In
particular, the invention
provides isolated native sequence PR0846 polypeptide, which in one embodiment,
includes an amino acid sequence
comprising residues 1 to 332 of Figure ?9 (SEQ ID N0:216). In ocher
embodiments, the invention provides an
isolated PR0846 polypeptide absem the signal sequence, which includes an amino
acid sequence comprising residues
from about 18 to 332 of Figure 79 (SEQ ID N0:216). ~rdditionaI embodiments of
the present invention are directed
to an isolated PR0846 polypeptide comprising amino acid 1 or about 18 to X of
Figure 79 (SEQ ID N0:216), where
X is any amino acid from 243 to 252 of Figure 79 (SEQ ID N0:216). <~tionally,
the PR0846 polypeptide is
obtained or is obtainable by expressing the polypeptide encoded by the cDNA
insert of the DNA44196-1353 vector
deposited on May 6, 1998 as ATCC 209847.
34. _P_R0862
Applicants have identified a cDNA clone that encodes a novel polypeptide
having sequence similarity to
lysozyme, wherein the polypeptide is designated in the present application as
"PR0862" .
In one embodiment, the invention provides an isolated nucleic acid molecule
comprising DNA encoding a
PR0862 poiypeptide. in one aspect, the isolated nucleic acid comprises DNA
encoding the PR0862 polypeptide
having amino acid residues 1 to 146 of Figure 81 (SEQ ID N0:221), or is
complementary to such encoding nucleic
acid sequence, and remains stably bound to it under at least moderate" and
optionally, under high stringency
conditions. in another aspect, the isolated nucleic acid comprises DNA
encoding the PR0862 polypeptide having
amino acid residues about 19 to 146 of Figure 81 {SEQ ID N0:22i), or is
complementary to such encoding nucleic
acid sequence, and remains stably bound to it under at least moderate, and
optionally; under high stringency
conditions, The isolated nucleic acid sequence may comprise the eDNA insert of
the DNA5218?-1354 vector
deposited on May 6, 1998 as ATCC 209845 which includes the nucleotide sequence
encoding PR0862.
In another embodiment, the invention provides isolated PR0862 polypeptide. In
particular, the invention
provides isolated native sequence PR0862 polypeptide, which in one embodiment,
includes an amino acid sequence
comprising residues 1 to 146 of Figure 81 (SEQ ID N0:22I). in another
embodiment, the invention provides an
isolated PR0862 polype~de absent the signal sequence, which includes an amino
acid sequence comprising residues
from about 19 to 146 of Figure 81 (SEQ ID N0:221 ). Optionally; the PR0862
polypeptide is obtained or is
obtainable by expressing the,polypeptide encoded by the cDNA insert of the
DNA52187-1354 vector deposited on
May 6, 1998 as ATCC 209845.
35. P-R_0864
Applicants have identified a cDNA clone that encodes a novel polypeptide
having sequence similarity to
Wnt-4, wherein the polypeptide is designated in the present application as
"PR0864".
In one embodiment, the invention provides an isolated nucleic acid molecule
comprising DNA encoding a
PR0864 polypeptide. In one aspect, the isolated nucleic acid comprises DNA
encoding the PR0864 polypeptide
having amino acid residues l to 351 of Figure 83 (SEQ ID N0:226), or is
complementary to such encoding nucleic
acid sequence, aril remains stabiy bound to it under at least moderate, and
opaottatly, under high stringency
conditions. In another aspect, the isolated nucleic acid comprises DN,A
encoding the PR0864 polypeptide having
amid acid residues about 23 to 351 of Figure 83 (SEQ ID N0:226), or is
complementary to such encoding nucleic
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acid sequence, and remains stably bound to it under at least moderate, and
optionally, under high stringency
conditions. The isolated nucleic acid sequence may comprise the cDNA insert of
the DNA48328-1355 vector
deposited on May 6, 1998 as ATCC 209843 which includes the nucleotide sequence
encoding PR0864.
In another embodiment, the invention provides isolated PR0864 polypeptide. In
particular, the invention
provides isolated native sequence PR0864 polypeptide, which in one embodiment,
includes an amino acid sequence
comprising residues 1 to 351 of Figure 83 (SEQ ID N0:226). In another
embodiment, the invention provides an
isolated PR0864 polygeptide absent the signal sequence, which includes an
amino acid sequence comprising residues
from about 23 to 351 of Figure 83 (SEQ ID N0:226). Optionally, the PR08b4
polypeptide is obtained or is
obtainable by expressing the polypeptide encoded by the cDNA insert of the
DNA48328-1355 vector deposited on
May 6, 1998 as ATCC 209843
36. PR, 0792
Applicants have identified a cDNA clone that encodes a novel polypeptide
having homology to CD23,
wherein the golypeptide is designated in the present application as "PR0792".
In one embodiment, the invention provides an isolated nucleic acid molecule
comprising DNA encoding a
PRO'792 polypeptide. In one aspect, the isolated nucleic acid comprises DNA
encoding the PR0792 polypeptide
having amino acid residues 1 to 293 of Figure 85 (SEQ ID NO:231), or is
complementary to such encoding nucleic
acid sequence, and remains stably bound to it under at least moderate, and
optionally, under high stringency
conditions. In another aspect, the isolated nucleic acid comprises DNA
encoding the PR0792 polypeptide having
amino acid residues X to 293 of Figure 85 (SEQ ID N0:231) where X is any amino
acid from 50 to 59 of Figure 85
(SEQ ID N0:231), or is compiemen<ary to such encoding rnicleic acid sequence,
and remains stably bound to it under
at least moderate, and optionally, under high stringency conditions. The
isolated nucleic acid sequence may comprise
the cDNA insert of the DNA56352-1358 vector deposited on May 6, 1998 as ATCC
209846 which includes the
nucleotide sequence encoding PR0792.
1n another embodiment, the invention provides isolated PR0792 polypeptide. In
particular, the invention
provides isolated native sequence PR0792 polypeptide, which in one embodiment,
includes an amino acid sequence
comprising residues i to 293 of Figure 85 (SEQ ID N0:231). An additional
embodiment of the present invention
is directed to PR0792 polygeptide comprising amino acids X to 293 of Figure 85
(SEQ ID N0:231), where X is any
amino acid from 50 to 59 of. Figure 85 (SEQ ID N0:231). Optionally, the PR0792
poiypeptide is obtained or is
obtainable by expressing the poIypeptide encoded by the cDNA insert of the
DNA56352-1358 vector deposited on
May 6, 1998 as ATCC 209846.
37. ~'R0866
Appii~s have identified a cDNA clone that encodes a novel polypeptide having
homology to mindin and
spondin proteins, wherein the polypeptide is designated in the present
application as "PR0866".
In one embodiment, the invention provides an isolated nucleic acid molecule
comprising DNA encoding a
PR0866 polypeptide. In one aspect, the isolated nucleic acid comprises DNA
encoding the PR0866 polypeptide
having amitw acid residues 1 to 331 of Figure 87 (SEQ ID N0:236), or is
complementary to such encoding nucleic
acid sequence, and remains stably bound to it under at least moderate, and
optionally, under high suingency
conditions. In another aspect, the isolated trucleic acid comprises DNA
encoding the PR0866 polypeptide having
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amino acid residues about 27 to 229 of Figure 87 (SEQ ID NO:236), or is
complemezttary to such encoding nucleic
acid sequence, and remains stably bound to it under at least moderate, and
optionally, under high stringency
conditions. The isolated nucleic acid sequence may comprise the cDNA insert of
the DNA53971-1359 vector
deposited on April 7, 1998 as ATCC 209750 which includes the nucleotide
sequence encoding PR0866.
In another embodiment, tire invention provides isolated PR0866 polypeptide. In
particular, the invention
provides isolated native sequence PR0866 polypeptide, which in one embodiment,
includes an amino acid sequence
comprising residues 1 to 331 of Figure 87 {SEQ ID NO:236). Another embodiment
of the present invention is
directed to PR0866 polypeptides comprising amino acids about 27 to 331 of
Figure 87 (SEQ ID N0:236).
Optionally, the PR0866 polypeptide is obtained or is obtainable by expressing
the polypeptide encoded by the cDNA
insert of the DNA53971-1359 vector deposited on April ?, 1998 as ATCC 209750.
38. PR087~
Applicants have identified a cDNA clone that encodes a novel palypeptide
having homology to CyP-60,
wherein the polypeptide is designated in the present application as "PR0871".
In one embodiment, the invention provides an isolated nucleic acid molecule
comprising DNA encoding a
PR0871 polypeptide. In one aspect, the isolated nucleic acid comprises I)NA
encoding the PR0871 polypeptide
having amino acid residues 1 to 472 of Figure 89 (SEQ ID N0:245), or is
complementary to such encoding nucleic
acid sequence, and remains stably bound to it under at least moderate, and
optionally, under high stringency
conditions. In other aspects, the isolated nucleic acid comprises DNA encoding
the PR0871 polypeptide having
amino acid residues about 22 to 472 of Figure 89 (SEQ ID N0:245), or is
complementary to such encoding nucleic
acid sequence, and remains stably bound to it under at least moderate, and
optionally, under high stringency
conditions. The isolated nucleic acid sequence may comprise the cDNA insert of
the DNA50919-1361 vector
deposited on May 6, 1998 as ATCC 209848 which includes the nucleotide sequence
encoding PR0871.
In another embodiment, the invention provides isolated PR087I polypeptide. In
particular, the invention
provides isolated native sequence PR0871 polypeptide, which in one embodiment,
includes an amino acid sequence
comprising residues 1 to 472 of Figure 89 (SEQ i;D N0:245). An additional
embodiment of the present invention
is directed to PR0871 polypeptidea comprising amino acids about 22 to 472 of
Figure 89 (SEQ ID N0:245).
Optionally, the PR0871 polypeptide is obtained or is obtainable by expressing
the polypeptide encoded by the cDNA
insert of the DNA50919-1361 vector deposited on May 6, 1998 as ATCC 209848.
39. PR08~3
Applicants have identified a cDNA clone that encodes a novel polype~ptide
having homology to
carboxylesterase, wherein the polypeptide is designated in the present
application as "PRO873".
In one embodiment, the invention provides an isolated nucleic acid molecule
comprising DNA encoding a
PRO873 polypeptide. In one aspect, the isolated nucleic acid comprises DNA
encoding the PR0873 polypeptide
having amino acid residues 1 to 545 of Figure 91 (SEQ ID N0:254), or is
complementary to such encoding nucleic
acid sequence, and remains stably bound to it under at least moderate, and
optionally, under high suingency
conditions. In other aspects, the isolated nucleic acid comprises DNA.
encoding the PR0873 polypeptide haviag
amino acid residues about 30 to about 545 of Figure 91 (SEQ ID N0:254), or is
complementary to such encoding
ra~cleic acid sequence, and remains stably bound to it under at least
mode:race, and optionally, under high stringency
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conditions. The isolated nucleic acid sequence may comprise the cDNA insert of
the DNA44179-1362 vector
deposited on May 6, 1998 as ATCC 209851 which includes the nucleotide sequence
encoding PR0873.
In another embodiment, the invention provides isolated PRO873 polypeptide. In
particular, the invention
provides isolated native sequence PR0873 polypeptide, which in one embodiment,
includes an amino acid sequence
comprising residues 1 to 545 of Figure 91 (SEQ ID N0:254). Additional
embodiments of the present invention are
directed to PR0873 polypeptides comprising amino acids about 30 to about 545
of Figure 91 (SEQ ID NO:254).
Optionally, the PR0873 polypeptide is obtained or is obtainable by expressing
the poiypeptide encoded by the cDNA
insert of the DNA44179-1362 vector deposited on May 6, 1998 as ATCC 209851.
40. . PR
Applicants have identified a cDNA clone that encodes a novel polypeptide
having homology to CD33 and
OB binding protein-2, wherein the polypeptide is designated in the present
application as "PR0940".
h~ one embodiment, the invention provides an isolated nucleic acid molecule
comprising DNA encoding a
PR0940 polypeptide. In one aspect, the isolated nucleic acid comprises DNA
encoding the PR0940 polypeptide
having amino acid residues 1 to 544 of Figure 93 (SEQ ID N0:259), or is
complementary to such encoding nucleic
acid sequence, and remains stably bound to it under at least moderate, and
optionally, under high stringency
conditions. In other aspects, the isolated nucleic acid comprises DNA encoding
the PR0940 polypeptide having
amino acid residues about 16 to 544 of Figure 93 (SEQ ID N0:259) or 1 or about
16 to X of Figure 93 (SEQ ID
N0:259}, where X is any amino acid from 394 to 403 of Figure 93 (SEQ ID
N0:259), or is complementary to such
encoding nucleic acid sequence, and remains stably bound to it under at least
moderate, and optionally, under high
stringency conditions. The isolated nucleic acid sequence may comprise the
cDNA insert of the DNA54002-1367
vector deposited on April 7, 1998 as ATCC 209754 which includes the nucleotide
sequence encoding PR0940.
In another embodiment, the invention provides isolated PR0940 polypeptide. In
particular, the invention
provides isolated native sequence PR0940 polypeptide, which in one embodiment,
includes an amino acid sequence
comprising residues 1 to 544 of Figure 93 (SEQ ID N0:259}. Other embodimerns
of the present invention are
directed to PR0940 polypeptides comprising amino acids about 16 to 544 of
Figure 93 (SEQ 1D N0:259) or I or
about 16 to X of Figure 93 (SEQ ID N0:259), where X is any amino acid from 394
to 403 of Figure 93 (SEQ ID
N0:259). Optionally, the PR0940 polypeptide is obtained or is obtainable by
expressing the polypeptide encoded
by the cDNA insert of the DNA54002-1367 vector deposited on April 7, 1998 as
ATCC 209754.
41. PRO9$1
Applicants have identified a cDNA clone that encodes .a novel polypeptide
having homology to a cadherin
protein, wherein the polypeptide is designated in the present application as
"PR0941".
In one embodiment, the invention provides an isolated nucleic acid molecule
comprising DNA encoding a
PR0941 polypeptide. In one aspect, the isolated nucleic acid comprises DNA
encoding the PR094i polypeptide
having amino acid residues 1 to 772 of Figure 95 (SEQ ID N0:2b4), or is
complementary to such encoding nucleic
acid sequence, and remains stably bound to it under at least moderate, and
optionally, under high stringency
conditions. In other aspects, the isolated nucleic acid comprises DNA encoding
the PR0941 polypeptide having
amino acid residues about 22 to 772 of Figure 95 (SEQ ID N0:264) or l or about
22 to X of Figure 95 (SEQ ID
N0:264), where X is arty amino acid from 592 to 601 of Figure 95 (SEQ ID
N0:264), or is complementary to such
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encoding nucleic acid sequence, and remains stably bound to it under at least
moderate, and optionally, under high
stringency conditions. The isolated nucleic acid sequence may comprise the
cDNA insert of the DNA53906-1368
vector deposited on April 7, 1998 as ATCC 209747 which includes the nucleotide
sequence encoding PR0941.
In another embodiment, the invention provides isolated PRO941 polypeptide. In
particular, the invention
provides isolated native sequence PR0941 polypeptide, which in one embodiment,
includes an amino acid sequence
comprising residues 1 to 772 of Figure 95 (SEQ ID N0:264). Additional
embodiments of the present invention are
directed to PR0941 polypeptides which comprise amino acid about 21 to 77:d of
Figure 95 (SEQ ID N0:264) or 1
or about 22 to X of Figure 95 (SEQ ID N0:264), where X is any amino acid from
592 to 601 of Figure 95 (SEQ ID
N0264). Optionally, the PR0941 polypepdde is obtained or is obtainable by
expressing the polypeptide encoded
by the cDNA insert of the DNAS3906-1368 vector deposited on April 7, 1998 as
ATCC 209747
In another embodiment, the invention provides an expressed sequence tag (EST)
designated herein as
DNA6415 comprising the nucleotide sequence of Figure 96 (SEQ ID N0:265).
42. PR0944
Applicants have identified a cDNA clone that encodes a novel polypeptide
having homology to Clostridium
IS perfringens enterotoxin receptor (CPE-R), wherein the polypegtide is
designated in the present application as
"PR0944" .
In one embodiment, the invention provides an isolated nucleic acid molecule
comprising DNA encoding a
PR0944 polypeptide. In one aspect, the isolated nucleic acid comprises DNA
encoding the PR0944 polypeptide
having amino acid residues 1 to 211 of Figure 98 (SEQ ID N0:270), or is
complementary to such encoding nucleic
acid sequence, and remains stably bound to it under at least moderate, and
optionally, under high stringency
conditions. In other aspects, the isolated nucleic acid comprises DNA encoding
the PR0944 polypeptide having
amino acid residues about 22 to 229 of Figure 98 (SEQ ID N0:270) or amino acid
I or about 22 to X of Figure 98
(SEQ ID N0:270) where X is any amino acid from 77 to 80 of Figure 98 (SEQ ID
N0:270), or is complementary
to such encoding nucleic acid sequence, and remains stably bound to it under
at least moderate, and optionally, under
high stringency conditions. The isolated nucleic acid sequence may comprise
the eDNA insert of the DNA52185
1370 vector deposited on May 14, 1998 as ATCC 209861 which includes the
nucleotide sequence encoding PR0944.
In another embodiment; the invention provides isolated PR0944 polypeptide. In
particular, the invention
pzovides isolaoed native sequence PR0944 polypegtide, which in one embodiment,
includes an amino acid sequence
comprising residues I to 211 of Figure 98 (SEQ ID N0:270). Additional
embodiments of the present invention are
directed to PR0944 poiypepddes comprising amino acids about 22 to 2I 1 of
Figure 98 (SEQ ID N0:270) or amino
acid 1 or about 22 to X of Figure 98 (SEQ ID N0:270), where X is any amino
acid from 77 to 86 of Figure 98 (SEQ
ID N0:270). Optionally, the PR0944 polypeptide is obtained or is obtainable by
expressing the polypeptide encoded
by the eDNA insert of the DNA52I85-1370 vector deposited on May 14,, 1998 as
ATCC 209861.
In another embodiment, the invention provides an expressed sequence tag (EST)
designated herein as
DNA14007 comprising the nucleotide sequence of Figure 99 (SEQ ID NO:27I).
In another embodirc~nt, the invention provides an expressed sequence tag (FST)
designated herein as
DNA12733 comprising the nucleotide sequence of Figure 100 (SEQ ID N0:272).
In another embodiment, the invention provides an expressed sequence tag (EST)
designated herein as
DNA12746 comprising the nucleotide sequence of Figure 101 (SEQ ID N0:273).
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In another embodiment, the invention provides an expressed sequence tag (EST)
designated herein as
DNA12834 comprising the nucleotide sequence of Figure 102 (SEQ ID N0:274).
In another embodiment, the invention provides an expressed sequence tag (EST)
designated herein as
DNA12846 comprising the nucleotide sequence of Fiugure 103 (SEQ ID N0:275).
In another embodiment, the invention provides an expressed sequence tag (EST)
designated herein as
DNA13104 comprising the nucleotide sequence of Figure 104 (SEQ ID N0:276).
In another embodiment, the invention provides an expressed sequence tag (ES'T)
designated herein as
DNA13259 comprising the nucleotide sequence of Figure 105 (SEQ ID N0:277).
In another embodiment, the invention provides an expressed sequence tag (EST)
designated herein as
DNAI3959 comprising the nucieotide sequence of Figure 106 (SEQ ID N0:278).
In another embodiment, the invention provides an expressed sequence tag {EST)
designated herein as
DNA13961 comprising the nucleotide sequence of Figure 107 {SEQ ID N0:279).
43. ~'R0983
Applicants have identified a cDNA clone that encodes a novel polypeptide
having homology to a vesicle
associated protein, VAP-33, wherein the polypeptide is designated in the
present application as "PR0983".
In one embodiment, the invention provides an isolated nucleic acid molecule
comprising DNA encoding a
PR0983 polypeptide. In one aspect, the isolated nucleic acid comprises DNA
encoding the PR0983 polypeptide
having amino acid residues 1 to 243 of Figure 109 (SEQ ID N0:284), or is
complementary to such encoding nucleic
acid sequence, and remains stably bound to it under at least moderate, and
optionally, under high stringency
conditions. In other aspects, the isolated nucleic acid comprises DNA encoding
the PR0983 polypeptide having
amino acid residue 1 to X of Figure 109 (SEQ ID N0:284) where X is any amino
acid from 219 to 228 of Figure
109 (SEQ ID N0:284), or is complementary to such encoding nucleic acid
sequence, and remains stably bound to
it under at least moderate, and optionally, under high stringency conditions.
The isolated nucleic acid sequence may
comprise the eDNA insert of the DNA53977-1371 vector deposited on May 14, 1998
as ATCC 209862 which
includes the nucleotide sequence encoding PR0983.
In another embodiment, the invention provides isolated PR0983 polypeptide. In
particular, the invention
provides isolated native sequence PR0983 polypeptide, which in one embodiment,
includes an amino acid sequence
comprising residues 1 to 243 of Figure 109 (SEQ ID N0:284). Additional
embodiments of the present invention are
directed to PR0983 poiypeptides comprising amino acid 1 to X of Figure I09
(SEQ ID N0:284), where X is any
amino acid from 219 to 228 of Figure 109 (SEQ ID N0:284). Optionally, the
PR0983 poiypeptide is obtained or
is obtainable by expressing the polypeptide encoded by the cDNA insert of the
DNA53977-137I vector deposited on
May 14, 1998 as ATCC 209862.
In another embodiment, the invention provides an expressed sequence tag (EST)
designated herein as
DNA17130 comprising the nucleotide sequence of Figure 110 (SEQ ID N0:285).
In another embodiment, the invention provides an expressed sequence tag (EST)
designated herein as
DNA23466 comprising the nucleotide sequence of Figure 111 (SEQ ID N0:286).
In another embodiment, the invention provides an expressed sequence tag (EST)
designated herein as
DNA26818 comprising the nucleotide sequence of Figure 112 (SEQ ID N0:287).
In another embodiment, the invention provides an expressed sequence tag (EST)
designated herein as
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DNA37618 comprising the nucleotide sequence of Figure I13 (SEQ ID N0:288).
In another embodiment, the invention provides an expressed sequence tag (EST)
designated herein as
DNA41732 comprising the. nucleotide sequence of Figure 114 (SEQ ID N0:289).
In another embodiment, the invention provides an expressed sequence tag (EST)
designated herein as
DNA4S980 comprising the nucleotide sequence of Figure l IS (SEQ ID N0:290).
In another embodiment, the invention provides an expressed sequence tag {EST)
designated herein as
DNA46372 comprising the nucleotide sequence of Figure 116 (SEQ ID N0:291).
44. R007
Applicants have identified a cDNA clone that encodes a novel polypeptide
having homology to proteases,
wherein the polypeptide is designated in the present application as "PROIOS7".
In one embodiment, the invention provides an isolated nucleic acid. molecule
comprising DNA encoding a
PROlOS7 polypeptide. In one aspect, the isolated nucleic acid comprises DNA
encoding the PR01057 polypeptide
having amino acid residues I to 413 of Figure 118 (SEQ 1D N0:296), or is
complementary to such encoding nucleic
acid sequence, and remains stably bound to it under at least moderate, and
optionally, under high stringency
1S conditions. in other aspects, the isolated nucleic acid comprises DNA
encoding the PROlOS7 poiypeptide having
amino acid residues about 17 to 413 of Figure 1 I8 (SEQ ID N0:296), or is
complementary to such encoding nucleic
acid sequence, and remains stably bound to it under at least moderate, and
optionally, under high stringency
conditions. The isolated nucleic acid sequence may comprise the eDNA insert of
the DNAS72S3-1382 vector
deposited on May I4, 1998 as ATCC 209867 which includes the nucleotide
sequence encoding PROlOS7.
In another embodiment, the invention provides isolated PROlOS7 polypeptide. In
particular, the invention
provides isolated native sequence PROlOS7 polypeptide, which in one
embodiment, includes an amino acid sequence
comprising residues I to 413 of Figure 118 (SEQ ID NO:296). Additional
embodiments of the present invention are
directed to PROlOS7 polypeptides comprising amino acids about 17 to 413 of
Figure 118 (SEQ iD N0:296).
Optionally, the PROIOS7 polypeptide is obtained or is obtainable by expressing
the poiypeptide encoded by the cDNA
2S insert of the DNAS72S3-1382 vector deposited on May 14, 1998 as ATCC
209867.
45. PROI87I
Applicants have identified a cDNA clone that encodes a novel polypeptide
having homology to
thrombospondin, wherein the polypeptide is designated in the present
application as "PROI071".
In one embodiment, the invention provides an isolated nucleic acid molecule
comprising DNA encoding a
PR01071 polypeptide. In one aspect, the isolated nucleic acid comprises DNA
encoding the PR01071 polypeptide
having amino acid residues 1 to S2S of Fgure I20 {SEQ ID N0:30i), or is
complementary to such encoding nucleic
acid sequence, and remains stably bound to it under at least moderate, and
optionally, under high stringency
conditions. In other aspects, the isolated tmcleic acid comprises DNA encoding
the PR01071 polypeptide having
amino acid residues about 26 to 525 of Figure 120 (SEQ ID N0:301), or is
complementary to such encoding irueIeic
acid sequence, and remains stably bound to it under at least moderate, and
optionally, under high stringency
conditions. The isolated nucleic acid sequence may comprise the eDNA insert of
the DNAS8847-1383 vector
deposited on May 20, 1998 as ATCC 209879 which includes the nucleotide
sequence encoding PR01071.
In another embodiment: the invention provides isolated PROIO'71 polypeptide.
In particular, the invention
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provides isolated native sequence PR01071 polypeptide, which in one
embodiment, includes an amino acid sequence
comprising residues 1 to 525 of Figure 120 (SEQ ID N0:301). Additional
embodiments of the present invention are
directed to PR0107I polypeptides comprising amino acids about 26 to 525 of
Figure 120 (SEQ ID N0:301).
Optionally, the PR01071 polypeptide is obtained or is obtainable by expressing
the polypeptide encoded by the cDNA
insert of the DNA58847-1383 vector deposited on May 20, 1998 as ATCC 209879.
46. PR01072
Applicants have identified a cDNA clone that encodes a novel polypeptide
having homology to reductase
proteins, wherein the polypeptide is designated in the present application as
"PR01072".
In one embodiment, the invention provides an isolated nucleic acid molecule
comprising DNA encoding a
PR01072 polypeptide. In one aspect, the isolated nucleic acid comprises DNA
encoding the PR01072 polypeptide
having amino acid residues 1 to 336 of Figure 122 (SEQ ID N0:303), or is
complementary to such encoding nucleic
acid sequence, and remains stably bound to it under at least moderate, and
optionally, under high stringency
conditions. In other aspects, the isolated nucleic acid comprises DNA encoding
the PR01072 polypept:ide having
amino acid residues about 22 to 336 of Figure 122 (SEQ ID N0:303), or is
complementary to such encoding nucleic
IS acid sequence, and remains stably bound to it under at least moderate, and
optionally, under high stringency
conditions. The isolated nucleic acid sequence may comprise the cDNA insert of
the DNA5874?-1384 vector
deposited on May 14, 1998 as ATCC 209868 which includes the nucleotide
sequence encoding PR01072.
In another embodiment, the invention provides isolated PR01072 polypeptide: In
particular, the invention
provides isolated native sequence PR01072 polypeptide, which in one
embodiment, includes an amino acid sequence
comprising residues 1 to 336 of Figure 122 (SEQ 1D N0:303). Additional
embodiments of the present invention are
directed to PR01072 polypeptides comprising amino acids about 22 to 336 of
Figure 122 (SEQ ID N0:303).
Optionally, the PR01072 polypeptide is obtained or is obtainable by expressing
the polypeptide encoded by the cDNA
insert of the DNA58747-1384 vector deposited on May 14, 1998 as ATCC 209868.
In another embodiment, the invention provides an expressed sequence tag (ES'I~
designated herein as
DNA40210 comprising the nucleotide sequence of Figure 123 (SEQ ID N0:304).
47. PR01075
Applicants have identified a cDNA clone that encodes a novel polypeptide
having homology to protein
disulfide isomerase, wherein the polypeptide is designated in the present
application as "PR01075".
In one embodiment, tl~ invention provides an isolated nucleic acid molecule
comprising DNA encoding a
PR01075 polypeptide. In one aspect, the isolated nucleic acid comprises DNA
encoding the PRO1075 polypeptide
having amino acid residues 1 fo 406 of Figure 125 (SEQ ID N0:309), or is
complementary to such encoding rrucleic
acid sequence, and remains stably bound to it under at least moderate, and
optionally, under high stringency
conditions. In other aspects, the isolated nucleic acid comprises DNA encoding
the PR01075 polypeptide having
amino acrd residues about 30 to 406 of Figure 125 (SEQ ID N0:309), or is
complementary to such encoding nucleic
acid sequence, and remains stably bound to it under at least moderate, and
optionally, under high stringency
conditions. The isolated nucleic acid sequence may comprise the cDNA insert of
the DNA57689-1385 vector
deposited on May 14, 1998 as ATCC 209869 which includes the nucleotide
sequence encoding PROI075
In anther embodiment, the invention provides isolated PR01075 polypeptide. In
particular, the invention
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provides isolated native sequence PR01075 poiypeptide, which in one
embodiment, includes an amino acid sequence
comprising residues 1 to 406 of Figure 125 (SEQ ID N0:309). Additional
embodiments of the present invention are
directed to PR01075 polypeptides comprising amino acids about 30 to 406 of
Figure 125 (SEQ ID N0:309).
Optionally, the PR01075 polypeptide is obtained or is obtainable by expressing
the polypeptide encoded by the cDNA
insert of the DNA57689-1385 vector deposited on May i4, 1998 as ATCC 2 09869.
In another embodiment, the invention provides an expressed sequence tag (EST)
designated herein as
DNA13059 comprising the nucleotide sequence of Figure 126 (SEQ ID N0:310).
In another embodiment, the invention provides an expressed sequence tag (EST)
designated herein as
DNA19463 comprising the nucleotide sequence of Figure 127 (SEQ ID N0:311).
48. ~'ROl8I
Appli~rns have identified a cDNA cio~ that encodes a novel polypeptide having
homology to the cornichon
protein, wherein the polypeptide is designated in the present application as
"PR0181".
In of embodittnt, the invention provides an isolated nucleic acid molecule
comprising DNA encoding a
PR0181 polypeptide. In one aspect, the isolated nucleic acid comprises DNA
encoding the PROI81 polypeptide
having amino acid residues 1 to 144 of Figure 129 (SEQ ID N0:322), or is
complementary to such encoding nucleic
acid sequence, and remains stably bound to it under at Ieast moderate, and
optionally, under high stringency
conditions: In other aspects, the isolated nucleic acid comprises DNA encoding
the PR0181 polypepdde having
amino acid residues about 21 to 144 of Figure 129 (SEQ ID N0:322} or amino
acid I or about 2I to X of Figure 129
{SEQ ID N0:322) where X is any amino acid from 52 to 61 of Figure 129 (SEQ ID
N0:322), or is complementary
to such encoding rnxleic acid sequence, and remains stably bound to it under
at least moderate, and optionally, under
high stringency conditions. The isolated nucleic acid sequence may comprise
the cDNA insert of the DNA23330-
1390 vector deposited on April 14, 1998 as ATCC 2U9775 which includes the
nucleotide sequence encoding PR0181.
1n another embodiment, the invention pro~iides isolated PR018I polypeptide. In
particular, the invention
provides isolated native sequence PR0181 polypeptide, which in one embodiment,
includes an amino acid sequence
comprising residues 1 w 144 of Figure 129 (SEQ ID N0:322). Additional
earlbodiments of the present invention are
directed to PR018I polypepudes comprising amino acids about 2I to 144 of
Figure 129 {SEQ ID N0:322) or amino
acid 1 or about 2I to X of Figure 129 (SEQ ID N0:322}, where X is any amino
acid from 52 to 61 of Figure 129
(SEQ m N0:322). Optionatlx, the PR0181 polypeptide is obtained or is
obtainable by expressing the polypeptide
encoded by the cDNA insert of the DNA23330-1390 vector deposited on April 14,
1998 as ATCC 209775.
In another embodiment, the invention provides an expressed sequence tag (EST)
designated herein as
DNA13242 comprising the mtcleotide sequence of Figure 130 (SEQ ID N0:323).
49. PR0195
Applicants have identified a cDNA clot that encodes a novel transmembrane
polypeptide, wherein the
polypeptide is designated in the 'present application as "PR0195".
In one embodiment, the invention provides an isolated nucleic acid molecule
comprising DNA encoding a
PROI95 polypeptide. In one aspect, the isolated nucleic acid comprises DNA
encoding the PR0195 potypeptide
having amino acid residues 1 to 323 of Figure 132 (SEQ ID N0:330), or is
complementary to such encoding nucleic
acid sequence, and remains stably bound to it under at least moderate, and
optionally, under high stringency
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conditions. In other aspects, the isolated nucleic acid comprises DNA encoding
the PR0195 polypepiide having
amino acid residues about 32 no 323 of Figure 132 (SEQ ID N0:330) or amino
acid 1 or about 32 to X of Figure 132
(SEQ B~ N0:330) where X is any amino acid from 236 to 245 of Figure 132 (SEQ
ID N0:330), or is complementary
to such encoding nucleic acid sequence, and remains stably bound to it under
at least moderate, and optionally, under
high stringency conditions. The isolated nucleic acid sequence may comprise
the cDNA insert of the DNA26847-
1395 vector deposited on April 14, 1998 as ATCC 209772 which includes the
nucleotide sequence encoding PR0195.
In another embodiment, the invention provides isolated PR0195 polypeptide. In
particular, the invention
provides isolated native sequence PR0195 polypeptide, which in one embodiment,
includes an amino acid sequence
comprising residues 1 to 323 of Figure 132 (SEQ 1D N0:330). Additional
embodiments of the present invention are
directed to PR0195 polypeptides comprising amino acids about 32 to 323 of
Figure i32 (SEQ ID N0:330) or amino
acid I or about 32 to X of Figure i32 (SEQ ID N0:330), where X is any amino
acid from 236 to 245 of Figure 132
(SEQ ID N0:330). Optionally, the PR0195 polypeptide is obtained or is
obtainable by expressing the palypeptide
encoded by the cDNA insert of the DNA26847-1395 vector deposited on April 14,
1998 as ATCC 209772.
1n another embodiment, the invention provides an expressed sequence tag (EST)
comprising the nucleotide
sequence of Figure 133 (SEQ ID N0:331), herein designated DNA15062.
In another embodiment, the invention provides an expressed sequence tag (EST)
comprising the nucleotide
sequence of Figure I34 (SEQ TD N0:332), herein designated DNA13199.
50. PR0865
Applicants have identified a cDNA clone that encodes a novel secreted
polypeptide, wherein the polypeptide
is designated in the present application as "PR0865".
In one embodiment, the invention provides an isolated nucleic acid molecule
comprising DNA encoding a
PR0865 polypeptide. In one aspect, the isolated nucleic acid comprises DNA
encoding the PR0865 polypepdde
having amino acid residues 1 to 468 of Figure 136 (SEQ ID N0:337), or is
complementary to such encoding nucleic
acid sequence, and remains stably bound to it under at least moderate, and
optionally, under high stringency
conditions. In other aspects, the isolated nucleic acid comprises DNA encoding
the PR0865 polypeptide having
amino acid residues about 24 tn 229 of Figure 136 (SEQ ID N0:337), or is
complementary to such encoding nucleic
acid sequence, and remains stably bound to it under at least moderate, and
optionally, under high stringency
conditions. The isolated nucleic acid sequence may comprise the cDNA insert of
the DNA539?4-1401 vector
deposited on April 14, 1998 as ATCC 209774 which includes the nucleotide
sequence encoding PR0865.
In another embodiment, the invention provides isolated PR0865 polypeptide. In
particular, the invention
provides isolated native sequence PR0865 polypeptide, which in one embodiment,
includes an amino acid sequence
comprising residues 1 to 468 of Figure 136 (SEQ ID N0:337). An additional
embodiment of the present invention
is directed to a PR0865 polypeptide comprising amino acids about 24 to 468 of
Figure 136 (SEQ ID N0:337).
Optionally, the PR0865 polypeptide is obtained or is obtainable by expressing
the polypeptide encoded by the cDNA
insert of the DNA53974-1401 vector deposited on April 14, 1998 as ATCC 209774.
In another embodiment, the invention provides an expressed sequence tag (EST)
comprising the nucleotide
sequence of Figure 137 (SEQ ID N0:338), herein designated as DNA37642.
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51. PRQ827
Applicants have identified a cDNA clone that encodes a novel polypeptide
having homology to integrin
proteins, wherein the polypeptide is designated in the present application as
"PRO827".
In one embodiment, the invention provides an isolated nucleic acid molecule
comprising DNA encoding a
PR0827 polypeptide. In one aspect, the isolated nucleic acid comprises DNA
encoding the PR0827 polypeptide
having amino acid residues 1 to 124 of Figure 139 (SEQ ID N0:346), or is
complementary to such encoding nucleic
acid sequence, and remains stabiy bound to it under at least moderate, and
optionally, under high stringency
conditions. In other aspects, the isolated nucleic acid comprises DNA encoding
the PR0827 polypeptide having
amino acid residues about 23 to 124 of Figure I39 (SEQ ID N0:346), or is
complementary to such encoding nucleic
acid sequence, and remains stably bound to it under at Least moderate, and
optionally, under high stringency
IO conditions. The isolated nucleic acid sequence may comprise the eDNA insert
of the DNA57039-1402 vector
deposited on April 14, 1998 as ATCC 209777 which includes the nucleotide
sequence encoding PR0827.
In another embodiment, the invention provides isolated PR0827 polypeptide. In
particular, the invention
provides isolated native sequence pR0827 polypeptide, which in one embodiment,
includes an amino acid sequence
comprising residues 1 to 124 of Figure 139 (SEQ ID N0:346). An additional
embodiment of the present invention
is directed to a PR0827 polypeptide comprising amino acids about 23 to 124 of
Figure 139 (SEQ ID N0:346).
Optionally, the PR0827 polypeptide is obtained or is obtainable by expressing
the polypeptide encoded by the cDNA
insert of the DNA57039-1402 vector deposited on April 14, 1998 as ATCC 2097?7.
SZ. PR01114
2d Applicants have identified a cDNA clone that encodes a novel polypeptide
having homology to cytoltine
receptor family! proteins, wherein the poiypeptide is designated in the
present application as "PROI114".
In one embodiment, the invention provides an isolated nucleic acid molecule
comprising DNA encoding a
PR01114 poiypeptide. In one aspect, the isolated nucleic acid comprises DNA
encoding the PR01114 polypeptide
having amino acid trsidues 1 to 311 of Figure 142 (SEQ ID N0:352), or is
complementary to such encoding nucleic
acid sequence, and remains stably bound to it under at least moderate, and
optionally, under high stt~ingency
conditions. In other aspects, the isolated nucleic acid comprises DNA encoding
the PR01114 polypeptide having
amino acid residues about 30 to 311 of Figure i42 (SEQ ID N0:352} or amino
acid 1 or about 30 to X of Figure 142
(SEQ ID N0:352), where, X is any amino acid from 225 to 234 of Figure 142 (SEQ
ID N0:35Z), or is
completnentaty to stilt encoding nucleic acid sequence, and remains stably
bound to it under at least moderate, and
optionally, under high stringency conditions. The isolated nucleic acid
sequence may comprise the cDNA insert of
the DNA57033-1403 vector deposited on May 27, 1998 as ATCC 209905 which
includes the nucleotide sequence
encoding PR01114.
In another embodiment, the invention provides isolated PRO! 1:!4 polypeptide.
In particular, the invention
Provides isolated native sequence PROs 114 polypeptide, which in one
embodiment, includes an amino acid sequence
comprising residues 1 to 311 of Figure 142 (SEQ ID N0:352). Additional
embodiments of the present imrention are
directed to PR01114 polypepttdes comprising amino acids about 30 to 311 of
Figure 142 (SEQ ID N0:352) or amino
acid 1 or about 30 to X of Figure 142 (SEQ ID N0:352), where X is any amino
acid from 225 to 234 of Figure 142
(SEQ ID N0:352). Ogtsonally, the pR01114 polypeptide is obtained or is
obtainable by expressing the polypeptide
encoded by the cDNA insert of the DNA57033-1403 vector deposited on May 27,
1998 as ATCC 209905.
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In another embodiment, the invention provides an expressed sequence tag (EST)
designated herein as
DNA48466 comprising the nucleotide sequence of Figure 143 (SEQ ID N0:353).
A cDNA clone (DNA57033-1403) has been identified that encodes a novel
interferon receptor polypeptide,
designated in the present application as "PROl I14 interferon receptor".
In one embodiment, the invention provides an isolated nucleic acid molecule
comprising DNA encoding a
PR01114 interferon receptor polypeptide.
In one aspect, the isolated nucleic acid comprises DNA having at least about
80% sequence identity,
preferably at least about 85% sequence identity, more preferably at least
about 90'~v sequence identity, most
preferably at least about 95 k sequence identity to (a) a DNA molecule
encoding a PR01114 interferon receptor
polypeptide having the sequence of amino acid residues from about I or about
30 to about 311, inclusive of Figure
142 (SEQ ID N0:352), or (b) the complement of the DNA molecule of (a).
In another aspect, the invention concerns an isolated nucleic acid molecule
encoding a PR01114 interferon
receptor polypeptide comprising DNA hybridizing to the complement of the
nucleic acid between about nucleotides
250 or about 337 and about 1182, inclusive, of Figure 141 (SEQ lD N0:351).
Preferably, hybridization occurs under
stringent hybridization and wash conditions.
In a further aspect, the invention concerns an isolated nucleic acid molecule
comprising DNA having at least
about 80%n sequence identity, preferably at least about 85% sequence identity,
mote preferably at least about 90%
sequence identity, most preferably at least about 95% sequence identity to (a)
a DNA molecule encoding the same
mature polypeptide encoded by the human protein cDNA in ATCC Deposit No.
209905 (DNA57033-1403) or (b)
Lhe complement of the nucleic acid molecule of (a). In a preferred embodiment,
the nucleic acid comprises a DNA
encoding the same mature polypeptide encoded by the human protein cDNA in ATCC
Deposit No. 209905
(DNAS?033-1403).
In still a further aspect, the invention concerns an isolated nucleic acid
molecule comprising (a) DNA
a polypeptide having at least about 8096 sequence identity, preferably at
least about 85% sequence identity,
more preferably at Ieast about 90% sequeace identity, most preferably at least
about 95% sequence identity to the
sequence of amino acid residues 1 or about 30 to about 311, inclusive of
Figure 142 (SEQ ID N0:352), or (b) the
complement of the DNA of (a).
In a further aspect, the invention concerns an isolated nucleic acid molecule
having at least 10 nucleotides
and produced by hybridizing,a test DNA molecule under stringern conditions
with (a) a DNA molecule encoding a
PR01114 interferon receptor polypeptide having the sequence of amino acid
residues from 1 or about 30 to about
311, iuch~sive of Figure 142 (SEQ 1D N0:352), or (b) the complement of the DNA
molecule of (a), and, if the DNA
molecule has at least about an 80 ~ sequence identity, prefereably at least
about an 85 % sequence identity, more
preferably at least about a 90 ~ sequence identity, most preferably at least
about a 95 ~ sequence identity to (a) or
(b), isolating the test DNA molecule.
by a specific aspect, the invention provides an isolated nucleic acid molecule
comprising DNA encoding a
PR01114 interferon receptor polypeptide, with or without the N-terminal signal
sequence and/or the initiating
metbionine, a3nd its soluble, i.e., transtnetnbrane domain deleted or
inactivated variants, or is complementary to such
encoding nucleic acid molecule. The signal peptide bas been tentatively
identified as extending from about amino
acid position 1 to about amino acid position 29 in the sequence of Figure 142
(SEQ ID N0:352). The transmembrane
domain has been tentatively identified as extending from about amino acid
position 230 to about amino acid position
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255 in the PROs t 14 interferon receptor amino acid sequence (Figure 142, SEQ
ID N0:352).
In another aspect, the invention concerns an isolated nucleic acid molecule
comprising (a) DNA encoding
a polypeptide scoring at least about 80% positives, preferably at least about
85% positives, more preferably at least
about 90~ positives, most preferably at least about 95% positives when
compared with the amino acid sequence of
residues 1 or about 30 to about 311, inclusive of Figure. I42 (SEQ ID N0:352),
or (b) the complement of the DNA
S of (a).
Another embodiment is directed to fragments of a PR01114 interferon receptor
polypeptide coding sequence
that may find use as hybridization probes. Such nucleic acid fragments are
fiom about 20 to about 80 nucleotides
in length, preferably from about 20 to about 60 nucleotides in length, more
preferably from about 20 to about 50
nucleotides in length and most preferably from about 20 to about 40
nucleotides in length and may be derived from
the nucleotide sequence shown in Figure 141 ($EQ ID N0:351).
In another embodiment, the invention provides a vector comprising DNA encoding
PR01114 interferon
receptor or its variants. The vector may comprise any of the isolated nucleic
acid molecules hereinabove identified.
A host ceU comprising such a vector is also provided. By way of example, the
host cells may be CHO cells,
E. colt, or yeast. A process for producing PROs 114 interferon receptor
polypeptides is further provided and
comprises culturing host cells under conditions suitable for expression of
PR01114 interferon receptor and recovering
PR01 I 14 interferon receptor from the cell culture.
In another embodiment, the invention provides isolated PRO/ ! I4 interferon
receptor polypeptide encoded
by any of the isolated nucleic acid sequences hereinabove identified.
In a specific aspect, the invention provides isolated native sequence PR01114
interferon receptor
polypeptide, which in certain embodiments, includes an amino acid sequence
comprising residues 1 or about 30 to
about 31 ! of Figure 142 (SEQ ID N0:352).
In another aspect, the invention concerns an isolated PROI114 interferon
receptor polypeptide, comprising
an amino acid sequence /raving at least about 80 k sequence identity,
preferably at least about 85 % sequence identity,
more preferably at least about 909b sequence identity, most preferably at
least about 95 % sequence identity to the
sequence of amino acid residues 1 or about 30 to about 311, inclusive of
Figure I42 (SEQ ID N0:352).
In a futtlrer aspect, the imerrtion concerns an isolated PR01114 interferon
receptor polypepude, comprising
an amino acid scquerrce scoring at least about 80% positives, preferably at
least about 85 % positives, more preferably
at least about 90% positives, most preferably at Least about 95% positives
when compared with the amino acid
sequence of residues 1 or about 30 to about 311, inclusive of Figure 142 (SEQ
ID N0:352).
In yet another aspect, the invention concerns an isolated PR01114 interferon
receptor polypeptide,
comprising the sequence of amir>o acid residues t or about 30 to about 311,
inclusive of Figure 142 (SEQ ID
N0:352), or a fragrneru thereof sufficient to provide a binding site for an
anti-PR01 I 14 interferon receptor antibody.
Preferably, the PR01114 interferon receptor fragment retains a qualitative
biological activity of a motive PROl l l4
interferon receptor poIypeptide.
In a still further aspect, the invention provides a polypeptide produced by
(t) hybridizing a test DNA
molecule under stringent conditions with {a) a DNA molecule encoding a PR01114
interferon receptor polypeptide
having the seqrreace of amrim acid residues from about I or about 30 to about
31I, inclusive of Figure I42 (SEQ ID
N0:352), or (b) the complemeru of the DNA mrolecule of (a), and if the test
DNA molecule has at least about an 80%
sequence identity, preferably at least above an 859b sequence identity, more
preferably at least about a 90~ sequer~e
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identity, most preferably at least about a 95 'I6 sequence identity to (a) or
(b), (ii) culturing a host cell comprising the
test DNA molecule under conditions suitable for expression of the polypeptide,
and {iii) recovering the polypeptide,
from the cell culture.
In another embodiment, the invention provides chimeric molecules comprising a
PRO1114 interferon
receptor polypeptide fused to a heterologous polypeptide or amino acid
sequence. An example of such a chimeric
molecule comprises a PR01114 interferon receptor polypeptide fused to an
epitope tag sequence or a Fc region of
an iznmunoglobulin.
In another embodiment, the invention provides an antibody which specifically
binds to a PROl 1 I4 interferon.
receptor poiypeptide. Optionally, the antibody is a monoclonal antibody.
In yet another ettlbodirttent, the invention concerns agonists and antagonists
of a native PROl 114 interferon
: receptor polypeptide. In a particular embodiment, the agonist or antagonist
is an anti-PR01114 interferon receptor
antibody.
In a further embodiment, the invention concerns a method of identifying
agonists or antagonists of a native
PR41114 interferon receptor polypeptide by contacting the native PR01114
interferon receptor polypeptide with a
candidate molecule and monitoring a biological activity mediated by said
polypeptide.
in a still further embodiment, the invention concerns a composition comprising
a PRO11I4 interferon
receptor polypeptide, or an agonist or antagonist as hereinabove defined, in
combination with a phata~aceutically
acceptable carrier.
53. PR0237
Applicants have identified a cDNA clone that encodes a novel polypeptide
having homology to carbonic
anhydrase, wherein the polypeptide is designated in the present application as
"PR0237".
1n one embodiment, the invention provides an isolated nucleic acid molecule
comprising DNA encoding a
PR0237 polypeptide. In one aspect, the isolated nucleic acid comprises DNA
encoding the PR0237 polypeptide
having amino acid residues 1 to 328 of Figure 145 {SEQ ID N0:358), or is
complementary to such encoding nucleic
acid sequence, and remains stably bound to it under at least moderate, and
optionally, under high stringency
conditions. In other aspects, the isolated nucleic acid comprises DNA encoding
the PR0237 polypeptide having
amino acid residues about 24 to 328 of Figure 145 (SEQ ID N0:358) or amino
acid 1 or about 24 to X of Figure 145
{SEQ ID N0:358), where X is any amine acid from 172 to 181 of Figure 145 (SEQ
ID N0:358), or is
complementary to such encoding nucleic acid sequence, and remains stably bound
to it under at least moderate, and
optionally, under high stringency conditions. The isolated nucleic acid
sequence may comprise the cDNA insert of
the DNA34353-1428 vector deposited on May 12, 1998 as ATCC 209855 which
includes the nucleotide sequence
encoding PR0237.
In another embodiment, the invention provides isolated PR0237 polypeptide. In
particular, the invention
provides isolated native sequence PR0237 polypeptide, which in one embodiment,
includes an amino acid sequence
comprising residues 1 to 328 of Figure 145 (SEQ 1D N0:358). Additional
embodiments of the present invention are
directed to PR0237 poiypeptides comprising amino acids about 24 to 328 of
Figure 145 (SEQ ID N0:358) or amino
acid 1 or about 24 to X of Figure 145 (SEQ ID N0:358), where X is arty amino
acid from 172 to 181 of Figure 145
(SEQ ID N0:358). Optionally, the PR0237 polypeptide is obtained or is
obtainable by expressing the polypeptide
encoded by the cDNA insert of the DNA34353-1428 vector deposited on May 12,
1998 as ATCC 209855.
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54. PR 5 1
Applicants have identified a cDNA clone that encodes a novel polypeptide
having homology to a trypsin
inhibitor protein, wherein the polypeptide is designated irt the present
application as "PR0541 ".
In one embodiment, the invention provides an isolated nucleic acid molecule
comprising DNA encoding a
PR0541 polypeptide. In one aspect; the isolated nucleic acid comprises DNA
encoding the PR0541 polypeptide
having amino acid residues 1 to 500 of Figure 147 (SEQ ID N0:363), or is
complementary to such encoding nucleic
acid sequence, and remains stably bound to it under at least, moderate, and
optionally, under high stringency
conditions. In other aspects, the isolated nucleic acid comprises DNA encoding
the PR054i polypeptide having
amino acid residues about 21 to 500 of Figure 147 (SEQ 1D N0:363), or is
complementary to such encoding nucleic
acid sequence, and remains stably bound to it under at least moderate, and
optionally, under high stringency
conditions. The isolated nucleic acid sequence may comprise the cDNA insert of
the DNA45417-1432 vector
deposited on May 27, 1998 as ATCC 209910 which includes the nucleotide
sequence encoding PR0541.
In another embodiment, the invention provides isolated PR0541 poiypeptide. In
particular, the invention
provides isolated native sequence PR0541 polypeptide, which in one embodiment;
includes an amino acid sequence
corrtprising residues 1 to 500 of Figure 147 (SEQ 1D N0:363). Additional
embodiments of the present invention are
directed to PR054i polypeptides comprising amino acids about 21 to 500 of
Figure 147 (SEQ ID N0:363).
Optionally, the PR0541 polypeptide is obtait~d or is obtainable by expressing
the polypeptide encoded by the cDNA
insert of the DNA4541?-1432 vector deposited on May 27, 1998 as ATCC 209910.
55. ~R0273
Applicants have identified a cDNA clone that encodes a novel polypeptide,
wherein the polypeptide is
designated in the present application as "PR0273".
In one embodiment, the invention provides an isolated nucleic acid molecule
comprising DNA encoding a
PR0273 poiypeptide. In one aspect, the isolated nucleic acid comprises 'DNA
encoding the PR0273 polypeptide
having amino acid residues 1 through 111 of Figure 149 ($EQ ID N0:370), or is
complementary to such encoding
tnxxleic acid seq~ce, atui remains stably bound to it under at least moderate,
and optionally, under high stringency
conditions.
In another embodiment, the invention provides isolated PR0273 polypeptide. In
particular, the invention
provides isolated native sequence PR0273 polypeptide, which in one embodiment,
includes an amino acid sequence
comprising residues 1 through 111 of.Figure 149 (SEQ 1D N0:370).
56. PR0701
Applicants have identified a cDNA clone that encodes a novel golypeptide
having homology to neuroligins
I, 2, and 3, wherein the polypeptide is designated in the present application
as "PR0701".
In one err~oditnent, the invention provides an isolated nucleic acid molecuie
comprising DNA encoding a
PR070I polypeptide. In orte aspect, the isolated nucleic acid comprises DNA
encoding the PR0701 polypeptide
having amino acid residues 1 through 81b of Figure 1S1 (SEQ ID N0:375), or is
complementary to such encoding
tatcleic acid sequence, and remiains stably bound to it under at least
moderate, and optionally, under high stringency
conditions. The isolated ttucleic acid sequence may comprise the cDNA insert
of the vector deposited with the ATCC
on March 31, 1998 which includes the nucleotide sequence encoding PR0701.
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In another embodiment, the invention provides isolated PR0701 polypeptide. In
particular, the invention
provides isolated native sequence PR070i polypeptide, which in one embodiment,
includes an amino acid sequence
comprising residues 1 through 816 of Figure 151 (SEQ ID N0:375). An additional
embodiment of the present
invention is directed to an isolated extracellular domain of a PR0701
polypeptide. Optionally, the PR0701
polypeptide is obtained or is obtainable by expressing the polypeptide encoded
by the cDNA insert of the vector
S deposited with the ATCC on March 31, 1998.
57. PR0704
Applicants have identified a cDNA clone that encodes a novel polypeptide
having sequence identity with
VIP36, wherein the polypeptide is designated in the present application as
"PR0704".
10: In one embodiment, the invention provides an isolated nucleic acid
molecule comprising DNA encoding a
PR0704 polypeptide. In one aspect, the isolated nucleic acid comprises DNA
encoding the PR0704 polypeptide
having amino acid residues 1 through 348 of Figure 153 (SEQ ID N0:380), or is
complementary to such encoding
nucleic acid sequence, and remains stabIy bound to it under at least moderate,
and optionally, under high stringency
conditions. The isolated nucleic acid sequence may comprise the cDNA insert of
the vector deposited on March 31,
15 1998 with the ATCC as DNA509I I-1288, which includes the nucleotide
sequence encoding PR0704.
In another embodiment, the invention provides isolated PR0704 polypeptide. In
particular, the invention
provides isolated native sequence PR0704 polypeptide, which in one embodiment,
includes an amino acid sequence
comprising residues 1 through 348 of Figure 153 (SEQ ID N0:380). An additional
embodiment of the present
invention is directed to an isolated extracellular domain of a PR0704
polypeptide. Optionally, the PR0704
20 polypeptide is obtained or is obtainable by expressing the polypeptide
encoded by the cDNA insert of the vector
deposited on March 31, 1998 with the ATCC as DNA50911-1288.
58. PRO?06
Applicants have identified a cDNA clone that encodes a novel polypeptide
having homology to prostatic acid
25 phosphatase precursor and lysosomal acid phosphatase precursor, wherein the
polypepdde is designated in the present
application as "PR0706" .
In one embodiment, the invention provides an isolated nucleic acid molecule
comprising DNA encoding a
PR0706 polypeptide. In one aspect, the isolated nucleic acid comprises DNA
encoding the PR0706 polypeptide
having amino acid residues 1 through 480 of Figure 155 (SEQ ID N0:385), or is
complementary to such encoding
30 nucleic acid sequence, and remains stably bound to it under at least
moderate, and optionally, under high stringency
editions. The isolated nucleic acid sequence may comprise the cDNA insert of
the vector deposited on April 21,
1998 with the ATCC as DNA48329-1290 which includes the nucleotide sequence
encoding PRO706.
In another embodiment, the invention provides isolated PR0706 polypeptide. In
particular, the invention
provides isolated native sequence PR0706 polypeptide, which in one embodiment,
it~ludes an amino acid sequence
35 comprising residues 1 through 480 of Figure 155 (SEQ 1D N0:385), or
comprising residues 19 through 480 of Figure
155 (SEQ ID N0:385). Optionally, the PR0706 polypeptide is obtained or is
obtainable by expressing the
polypeptide encoded by the cDNA insert of the vector deposited on April 21,
1998 with the ATCC as DNA48329-
1290.
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S9. R07 7
Applicants have identified a cDNA clone that encodes a novel polypeptide
having homology to cadherins,
particularly cadherin FIB3,_wherein the polypeptide is designated in the
present application as "PR0707".
In one embodiment, the invention provides an isolated nucleic acid molecule
comprising DNA encoding a
PR0707 polypeptide. In one aspect, the isolated nucleic acid comprises DNA
encoding.the PR0707 polypeptide
having amino acid residues 1 to 916 of Figure 157 (SEQ ID N0:390), or is
complementary to such encoding nucleic
acid sequence, and remains stabIy bound to it under at least moderate, and
optionally, under high stringency
conditions. The isolated nucleic acid sequence may comprise the cDNA insert of
the vector deposited on May 27,
1998 with the ATCC as DNA48306-1291 which includes the nucleotide sequence
encoding PR0707.
In another embodiment, the invention provides isolated PR07U7 polypeptide. In
particular, the invention
provides isolated native sequence PR0707 polypeptide, which in one embodiment,
includes an amino acid sequence
comprising residues 1 to 916 of Figure 1S7 {SEQ ID N0:390). An additional
embodiment of the present invention
is directed to an isolated extracelIular domain of a PR0707 polypeptide:
Optionally, the PR0707 polypeptide is
obtained or is obtainable by expressing the polypeptide encoded by the cDNA
insert of the vector deposited on May
27,.1998 with the ATCC as DNA48306-1291.
60. R(
Applirams have identified a cDNA clone that encodes a novel polypeptide having
homology to neuropsin,
wherein the poIypeptide is designated in the present application as "PR0322".
In one embodiment, the invention provides an isolated nucleic acid molecule
comprising DNA encoding a
PR0322 polypeptide. In one aspect, the isolated nucleic acid comprises DNA
encoding the PRO322 polypeptide
having amino acid residues l or 24 through 260 of Figure 159 (SEQ ID N0:395),
or is complementary to such
encoding nucleic acid sequence, and remains stably bound to it under at least
moderate, and optionally, under high
stringency conditions. The isolated nucleic acid sequence may comprise the
cDNA insert of the vector deposited on
March I I, 1998 as ATCC no. 209669 which includes the nucleotide sequence
encoding PR0322.
In another embodiment, the invention provides isolated PR0322 polypeptide. In
particular, the invention
provides isolated native sequence PR0322 polypeptide, which in one embodiment;
includes an amino acid sequence
comprising residues l or 24 through 260 of Figure 159 {SEQ ID N0:395). An
additional embodi~nt of the present
invention is directed to an isolated extraceIlular domain of a PR0322
polypeptide. Optionally, the PR0322
po3ypeptide is obtai~d or is obtainable by expressing the polypeptide encoded
by the cDNA insert of the vector
deposited on March I1, 1998 as ATCC no. 209669.
61. P Z
Applicants have identified a cDNA ciotre that encodes a novel polypeptide
having sequence identity with
ALS, wherein the poiypeptide is designated in the present application as
"PR0526".
In one embodiment, the invention provides an isolated nucleic acid molecule
comprising DNA encoding a
PR0526 polypeptide. In one aspect, the isolated nucleic acid comprises DNA
encoding the PR0526 polypeptide
having amino acid residues 1 to 473 of Figure 161 {SEQ ID N0:400), or is
complementary to such encoding nucleic
acid sequence, and remains stably bound to it under at least moderate, and
optionally, under high stringency
conditions. The isolated nucleic acid sequence may comprise the cDNA insert of
the vector deposited on March 26,
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WO 99146281 PCTIUS99105028
1998 with the ATCC as DNA44184-1319 which includes the nucleotide sequence
encoding PR0526.
In another embodiment, the invention provides isolated PR0526 polypeptide: In
particular. the invention
provides isolated native sequence PR0526 polypeptide, which in one embodiment,
includes an amino acid sequence
comprising residues 1 to 473 of Figure 161 (SEQ ID N0:400). Optionally, the
PR0526 polypeptide is obtained or
is obtainable by expressing the polypeptide encoded by the cDNA insert of the
vector deposited on March 26, 1998
with the ATCC as DNA44184-1319 which includes the nucleotide sequence encoding
PR0526.
62. PR 5 I
Applicants have identified a cDNA clone that encodes a novel poiypeptide
having sequence identity with
protocadherins, wherein the polypeptide is designated in the present
application as "PR0531".
In one embodiment, the invention provides an isolated nucleic acid molecule
comprising DNA encoding a
PR0531 polypeptide. In one aspect, the isolated nucleic acid comprises DNA
encoding the PR0531 polypeptide
having amino acid residues 1 to 789 of Figure 163 (SEQ 1D N0:405), or is
complementary to such encoding nucleic
acid sequence, and remains stably bound to it under at least moderate, and
optionally, under high stringency
conditions. The isolated nucleic acid sequence may comprise the cDNA insert of
the vector deposited on March 26,
1998 as DNA48314-1320 which includes the nucleotide sequence encoding PR0531.
In another embodiment, the invention provides isolated PR0531 polypeptide. In
particular, the invention
provides isolated native sequence PR053! polypeptide, which in one embodiment,
includes an amino acid sequence
comprising residues 1 to 789 of Figure 163 (SEQ ID N0:405). An additional
embodiment of the present invention
is directed to an isolated extracellular domain of a PR0531 polypeptide.
Optionaliy, the PR0531 polypeptide is
obtained or is obtainable by expressing the polypeptide encoded by the cDNA
insert of the vector deposited on March
26, 1998 as DNA48314-1320.
63. PR(~534
Applicams have identified a cDNA clone that encodes a novel polypeptide having
sequence identity with
disulfide isomerase (sometimes referred to herein as protein disulfide
isomerase), wherein the polypeptide is
designated in the present application as "PR0534".
In one embodiment, the invention provides an isolated nucleic acid molecule
comprising DNA encoding a
PR0534 polypeptide. In one aspect, the isolated nucleic acid comprises DNA
encoding the PR0534 polypeptide
having amino acid residues 1 to 360 of Figure 165 (SEQ ID N0:410), or is
complementary to such encoding nucleic
acid sequence, and remains stably bound to it under at least moderate, and
optionally, under high stringency
conditions. The isolated nucleic acid sequence may comprise the cDNA insert of
the vector deposited on March 26,
1998 as DNA48333-1321 which includes the nucleotide sequence encoding PR0534.
In another embodiment, the invention provides isolated PR0534 polypeptide. In
particular, the invention
provides isolated native sequence PR0534 polypeptide, which in one embodiment,
includes an amino acid sequence
comprising residues 1 to 360 of Figure 165 (SEQ ID N0:410). An additional
embodiment of the present invention
is directed to an isolated exuacellular domain of a PR0534 polypeptide.
Optionally, the PR0534 polypeptide is
obtained or is obtainable by expressing the polypeptide encoded by the cDNA
insert of the vector deposited on March
26, 1998 as DNA48333-1321.
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64. PROb97
Applicants have identified a cDNA clone that encodes a novel poIypeptide
having sequence identity with
shRPs, wherein the polypeptide is designated in the present application as
"FR0697".
In one embodiment; the invention provides an isolated nucleic acid molecule
comprising DNA encoding a
PR0697 polypeptide. In one aspect, the isolated nucleic acid comprises DNA
encoding the PR0697 polypeptide
having amino acid residues 1 through 295 of Figure 167 (SEQ 1D N0:415), or is
complementary to such encoding
nucleic acid sequence, and remains stably bound to it under at least moderate,
and optionally, under high stringency
conditions. The isolated nucleic acid sequetx;e may comprise the cDNA insert
of the vector deposited with the ATCC
on March 26, 1998 as DNA50920-1325 which includes the nucleotide sequence
encoding PR0697.
In another embodiment, the invention provides isolated PR0697 polypeptide. In
particular, the invention
provides isolated native sequence PR0697 polypeptide, which in one embodiment,
includes an amino acid sequence
comprising residues 1 through 295 of Figure 167 (SEQ ID N0:415). Optionally,
the PR0697 polypeptide is obtained
or is obtainable by expressing the polypeptide encoded by the cDNA insert of
the vector deposited with the ATCC
on March 26, 1998 as DNA50920-1325.
65. . P 7 7
Applicants have identified a cDNA clone that encodes a novel 12 transmembrane
polypeptide, wherein the
polypeptide is designated is the present application as "PR0717".
In one embodiment, the invention provides an isolated nucleic acid molecule
comprising DNA encoding a
PR0717 polypeptide. In one aspect, the isolated nucleic acid comprises DNA
encoding the PR0717 polypeptide
having amino acid residues 1 through 560 of Figure 169 (SEQ ID N0:420), or is
complementary to such encoding
nucleic acid sequence, and remains stably bound to it under at least moderate,
and optionally, under high stringency
conditions. The isolated nucleic acid sequence may comprise the cDNA insert of
the vector deposited on April 28',
1998 with the ATCC as DNA50988-1326 which includes the nucleotide sequence
encoding PR0717.
In another embodiment, the invention provides isolated PR0717 polypeptide. In
particular, the invention
provides isolated native sequence PR0717 polypeptide, which in one embodiment,
inchtdes an amino acid sequence
comprising residues 1 through 560 of Figure 169 (SEQ ID N0:420). An additional
embodiment of the present
invention is directed to an isolated extracelIular domain of a FR0717
polypeptide. Optionally, the PR0717
polypeptide is obtained or is obtainable by expressing the polypepdde encoded
by the cDNA insert of the vector
deposited on April 28, 1998 with the ATCC as DNA50988-1326.
b6. PROZ31
Applicants have identified a cDNA clone that encodes a novel polypeptide
having sequence identity with
protocadherin 4, wherein the polypeptide is designated in the present
application as "PR0731".
In one embodiment, the invention provides an isolated nucleic acid molecule
comprising DNA encoding a
PR0731 polypeptide. In one aspect, the isolated nucleic acid comprises DNA
encoding the PRO731 polypeptide
having at>utto acid residues 1 through 1 l84 of Figure 171 (SEQ ID N0:425), or
is complementary to such encoding
nucleic acid sequence, and remains stably bound to it under at least moderate,
and optionally, under high stringency
co~tians. The isolated nucleic acid sequence may comprise the cDNA insert of
the vector deposited on March 31,
1998 with the ATCC as DNA48331-1329 which includes the nucleotide sequence
encoding PR0731.
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In another embodiment, the invention provides isolated PR0731 polypeptide. In
particular, the invention
provides isolated native sequence PR0731 polypeptide, which in one embodiment,
includes an amino acid sequence
comprising residues 1 through 1184 of Figure 171 (SEQ ID N0:425). An
additional embodiment of the present
invention is directed to an isolated extracellular domain of a PR0731
poIypeptide. Optionally, the PR0731
polypeptide is obtained or is obtainable by expressing the polypeptide encoded
by the cDNA insert of the vector
deposited on March 31, 1998 with the ATCC as DNA48331-1329.
67. PR0218
Applicants have identified a cDNA clone that encodes a novel multi-
transmembrane protein having sequence
identity with membrane regulator proteins, wherein the polypeptide is
designated in the present application as
"PR0218" .
In one embodiment, the invention provides an isolated nucleic acid molecule
comprising DNA encoding a
PR0218 polypeptide. In one aspect, the isolated nucleic acid comprises DNA
encoding the PR0218 polypeptide
having amino acid residues 1 through 455 of Figure 173 (SEQ ID N0:430), or is
complementary to such encoding
nucleic acid sequence, and remains stably bound to it under at least moderate,
and optionally, under high stringency
conditions. The isolated nucleic acid sequence may comprise the cDNA insert of
the vector deposited on April 28,
1998 with the ATCC as DNA30867-1335 which includes the nucleotide sequence
encoding PR0218.
In another embodiment, the invention provides isolated PR0218 polypeptide. In
particular, the invention
provides isolated native sequence PR0218 polypeptide, which in one embodiment,
includes an amino acid sequence
comprising residues 1 through 455 of Figure 173 (SEQ ID N0:430). Optionally,
the PR0218 polypeptide is obtained
or is obtainable by expressing the polypeptide encoded by the cDNA insert of
the vector deposited on April 28, 1998
with the ATCC as DNA30867-1335.
1n another embodiment, the invention provides an expressed sequence tag (EST)
sequence comprising the
nucleotide sequence of Figure 174 (SEQ ID N0:431), designated herein as
DNA14472.
In another embodiment, the invention provides an expressed sequence tag (EST)
sequence comprising the
nucleotide sequence of Figure 175 (SEQ ID N0:432), designated herein as DNA
15846.
68. PRO_'If_r8_
Applicants have identified a cDNA clone that encodes a novel polypeptide
having stquence identity with
integrins, wherein the polypegtide is designated in the present application as
"PR0768" .
In one embodiment, the invention provides an isolated nucleic acid molecule
comprising DNA encoding a
PR0768 polypeptide. In one aspect, the isolated nucleic acid comprises DNA
encoding the PRO?68 polypeptide
having amino acid residues 1 through 1141 of Figure 177 (SEQ ID N0:437), or is
complementary to such encoding
rn~cleic acid sequence, and remains stably bound to it under at least
moderate, and optionally, under high stringency
conditions. The isolated nucleic acid sequence may comprise the cDNA insert of
the vector deposited on April 6,
1998 as DNA55737-1345 which includes the nucleotide sequence encoding PR0768.
1n another embodiment, the invention provides isolated PR0768 polypeptide. In
particular, the invention
provides isolated native sequence PR0768 polypeptide, which in one
emboditnent, includes an amino acid sequence
comprising residues 1 through 1141 of Figure 177 (SEQ ID IV0:437). An
additional embodiment of the present
invention is directed to an isolated extracellular domain of a PR0768
polypeptide. Optionally, the PR0768
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potypeptide is obtained or is obtainable by expressing the polypeptide encoded
by the cDNA insert of the vector
deposited on April 6, 1998 as DNA55?37-1345.
69. PR0771
Applicants have identified a eDNA clone that encodes a novel polypeptide
having sequence identity with
testican, wherein the polypeptide is designated in the present application as
"PR0771 ".
In one embodiment, the invention provides an isolated nucleic acid molecule
comprising DNA encoding a
PR0771 polypeptide. In one aspect, the isolated nucleic acid comprises DNA
encoding the PR0771 polypeptide
having atnino acid residues 1 through 436 of Figure 179 (SEQ ID N0:442), or is
complementary to such encoding
nucleic acid sequence, and remains stably bound to it under at least moderate,
and optionally, under high stringency
conditions. The isolated nucleic acid sequence may comprise the cDNA insert of
the vector deposited on April 7,
1998 with the ATCC as DNA49829-1346 which includes the nucleotide sequence
encoding PR0771.
In another embodiment, the invention provides isolated PR0771 polypeptide. In
particular, the invention
provides isolated native sequence PR0771 polypeptide, which in one embodiment,
includes an amino acid sequence
comprising residues 1 through 436 of Figure 179 (SEQ ID N0:442). Optionally,
the PR0771 polypeptide is obtained
or is obtainable by expressing the polypeptide encoded by the cDNA insert of
the vector deposited on April 7, 1998
with the ATCC as DNA49829-1346.
?'0. PR0T33
Applicants have identified a cDNA clone that encodes a novel polypeptide
having sequence identity with
the TIlST2 receptor binding protein, wherein the polypeptide is designated in
the present application as "PR0733".
In one embodiment, the invention provides an isolated nucleic acid molecule
comprising DNA encoding a
PR0733 polypeptide. In one aspect, the isolated nucleic acid comprises DNA
encoding the PR0733 polypeptide
having amino acid residues 1 through 229 of Figure 181 (SEQ ID N0:447), or is
complementary to such encoding
ta~leic mid sequence, and remains stably bound to it under at least moderate,
and optionally, under high stringency
conditions. The isolated nucleic acid sequence may comprise the cDNA insert of
the vector deposited on April 7,
1998 with the ATCC as DNA52196-1348 which includes the nucleotide sequence
encoding PR0733.
In another embodiment, the invention provides isolated PR0733 polypeptide. In
particular, the invention
provides isolated native sequence PR0733 polypeptide, which is one embodiment,
includes an amino acid sequence
. comprising residues 1 through 229 of Figure 181 (SEQ ID N0:447). An
additional embodiment of the present
invention is directed to an isolated extracellular domain of a PR0733
polypeptide. Optionally, the PR0733
polypeptide is obtained or is obtainable by expressing the polypeptide encoded
by the cDNA insert of the vector
deposited on April 7, 1998 as DNA52196-1348.
71. PR0162
Applicants have identified a cDNA clone that encodes a novel polypeptide
having sequence identity with
pattcreatitis-associated protein, wherein the polypeptide is designated in the
present application as "PR0162".
In one embodiment, the invention provides an isolated nucleic acid rrx~lecuie
comprising DNA encoding a
PR0162 polypeptide. In one aspect,.the isolated nucleic acid comprises DNA
encoding the PR0162 polypeptide
having amino acid residues 1 through 175 of Figure 183 (SEQ ID N0:452), or is
complementary to such encoding
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nucleic acid sequence, and remains stably bound to it under at least moderate,
and optionally, under high stringency
conditions. The isolated nucleic acid sequence may comprise the cDNA insert of
the vector deposited on May 6,
1998 with the ATCC as DNA56965-1356 which includes the nucleotide sequence
encoding PROi62.
In another embodiment, the invention provides isolated PR0162 polypeptide. In
particular, the invention
provides isolated native sequence PR0162 polypeptide, which in one embodiment,
includes an amino acid sequence
comprising residues I through 175 of Figure 183 (SEQ ID N0:452). Optionally,
the PR0162 polypeptide is obtained
or is obtainable by expressing the poiypeptide encoded by the cDNA insert of
the vector deposited on May 6, 1998
with the ATCC as DNA56965-1356.
72. PR0788
IO Applicants have identified a cDNA clone that encodes a novel polypeptide
having sequence identity with
anti-neoplastic urinary protein, wherein the polypeptide is designated in the
present application as "PR0788".
In one embodiment, the invention provides an isolated nucleic acid molecule
comprising DNA encoding a
PRO?88 poIypeptide. 1n one aspect, the isolated nucleic acid comprises DNA
encoding the PR0788 polypeptide
having amino acid residues 1 through 125 of Figure 185 (SEQ ID NO:454), or is
complementary to such encoding
nucleic acid sequence, and remains stably bound to it under at least moderate,
and optionally, under high stringency
conditions. The isolated nucleic acid sequence may comprise the cDNA insert of
the vector deposited on May 6,
1998 with the ATCC as DNA56405-1357 which includes the nucleotide sequence
encoding PR0788.
in another embodiment, the invention provides isolated PR0788 polypeptide. In
particular, the invention
provides isolated native sequence PR0788 polypeptide, which in one embodiment,
includes an amino acid sequence
comprising residues 1 through 125 of Figure 185 (SEQ ID N0:454). An additional
embodiment of the present
invention is directed to an isolated extracellular domain of a PR0788
polypeptide. Optionally, the PR0788
polypeptide is obtained or is obtainable by expressing the polypeptide encoded
by the cDNA insert of the vector
deposited on May 6, 1998 with the ATCC as DNA56405-1357.
73. R0 8
Applicants have identified a cDNA clone that encodes a novel polypeptide
having sequence identity with
dickkopf 1 (dkk-1), wherein the polypeptide is designated in the present
application as "PR41008".
In one embodiment,. the invention provides an isolated nucleic acid molecule
comprising DNA encoding a
PR01008 polypeptide. In one aspect, the isolated nucleic acid comprises DNA
encoding the PR01008 polypeptide
having amino acid residues 1 through 266 of Figure 187 (SEQ ID N0:456), or is
complementary to such encoding
nucleic acid sequence, and remains stably bound to it under at least moderate,
and optionally, under high stringency
conditions. The isolated nucleic acid sequence may comprise the cDNA insert of
the vector deposited on May 20,
1998 with the ATCC as DNA57530-1375 which includes the nucleotide sequence
encoding PR01008.
In another embodiment, the invention provides isolated PR01008 polypeptide. In
particular, the invention
provides isolated native sequence PR01008 polypeptide, which in one
embodiment, includes an amino acid sequence
comprising residues 1 through 266 of Figure 187 (SEQ ID N0:456). Optionally,
the PR01008 polypeptide is
obtained or is obtainable by expressing the polypeptide encoded by the cDNA
insert of the vector deposited on May
20, 1998 with the ATCC as DNA57530-1375.
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In another embodiment, the invention provides an expressed sequence tag (EST)
designated herein as
DNAI6508 comprising the nucleotide sequence of Figure I88 (SEQ ID N0:457).
~a. ~~Q39.1~
Applicants have identified a cDNA clone that encodes a novel polypeptide
having sequence identity with
disulfide isomerase and phospholipase C, wherein the poiypeptide is designated
in the present application as
"PR01012" .
In one embodiment, the invention provides an isolated nucleic acid molecule
comprising DNA encoding a
PR01012 polypeptide. In one aspect, the isolated nucleic acid comprises DNA
encoding the PR01012 polypeptide
having amino acid residues 1 through 747 of Figure 190 (SEQ ID N0:459), or is
complementary to such encoding
nucleic acid sequence; and remains stably bound to it under at least moderate,
and optionally, under high stringency
conditions. The isolated nucleic acid sequence may comprise the eDNA insert of
the vector deposited on May 14,
1998 with the ATCC as DNA56439-1376, which includes the nucleotide sequence
encoding PR01012.
In another embodiment, the invention provides isolated PR01012 polypeptide. In
particular, the invention
provides isolated native sequence PR01012 polypeptide, which in one
embodiment, includes an amino acid sequence
comprising residues 1 through 747 of Figure 190 (SEQ ID N0:459). Optionally,
the PR01012 polypeptide is
obtained or is obtainable by expressing the polypeptide encoded by the cDNA
insert of the vector deposited on May
14, 1998 with the ATCC as DNASb439-1376.
75. PR01014
Applicants have identified a cDNA clone that encodes a novel polypeptide
having sequence identity with
reductase, wherein the poIypeptide is designated in the present application as
"PR01014" .
In one embodiment, the invention provides an isolated nucleic acid molecule
comprising DNA encoding a
PR01014 polypeptide. In one aspect, the isolated nucleic acid comprises DNA
encoding the PR01014 polypeptide
having amino acid residues 1 through 300 of Figure l92 (SEQ ID N0:464), or is
complementary to such encoding
nucleic acid sequence, and remains stably bound to it under at least moderate,
ark optionally, under high stringency
conditions. The isolated nucleic acid sequence may comprise the cDNA insert of
the vector deposited on May 20,
1998 as DNA564t19-1377 with the ATCC which includes the nucleotide sequence
encoding PR01014.
In another embodiment, the invention provides isolated PR01014 polypeptide. In
particular, the invention
provides isolated native seque~e PR01014 polypeptide, which in one embodiment,
includes an atznno acid sequence
comprising residues 1 through 300 of Figure 192 (SEQ ID N0:464). Optionally,
the PR01014 polypeptide is
obtained or is obtainable by expressing the polypeptide encoded. by the cDNA
insert of the vector deposited on May
20, 1998 as DNA56409-1377 with the ATCC.
76. PROtOl~
Applicants have identified a cDNA clone that encodes a novel polypeptide
having sequence irierniry with
HNK-i sulfotransferase, wherein the polypeptide is designated in the present
application as "PR01017" .
In one embodiment; the invention provides an isolated nucleic acid molecule
comprising DNA et>coding a
PRO101? polypeptide. in otie aspect, the isolated nucleic acid comprises DNA
encoding the PR01017 polypeptide
having amino acid residues 1 through 414 of Figure 194 (SEQ ID N0:466), or is
complementary to such encoding
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nucleic acid sequence, and remains stably bound to it under at least moderate,
and optionally, under high stringency
conditions. The isolated nucleic acid sequence may comprise the cDNA insert of
the vector deposited on May 20,
1998 with the ATCC as DNA56112-1379 which includes the nucleotide sequence
encoding PR01017.
In another embodiment, the invention provides isolated PR01017 polypeptide. In
particular, the invention
provides isolated native sequence PR01017 polypeptide, which in one
embodiment, includes an amino acid sequence
comprising residues 1 through 414 of Figure 194 (SEQ ID N0:466). Optionally,
the PR01017 polypeptide is
obtained or is obtainable by expressing the polypeptide encoded by the cDNA
insert of the vector deposited on May
20, 1998 with the ATCC as DNA56112-1379.
77. PR0474
Applicants have identified a cDNA clone that encodes a novel polypeptide
having sequence identity with
dehydrogenase, wherein the polypeptide is designated in the present
application as "PR0474".
In one embodiment, the invention provides an isolated nucleic acid molecule
comprising DNA encoding a
PR0474 polypeptide. In one aspect, the isolated nucleic acid comprises DNA
encoding the PR0474 polypeptide
having amino acid residues 1 through 270 of Figure 196 (SEQ iD N0:468), or is
complementary to such encoding
nucleic acid sequence, and remains stably bound to it under at least moderate,
and optionally, under high stringency
cot~itions. The isolated nucleic acid sequence may comprise the cDNA insert of
the vector deposited on May I4,
1998 with the ATCC as DNA56045-1380 which includes the nucleotide sequence
encoding PR0474.
In another embodiment, the invention provides isolated PR0474 polypeptide. In
particular, the invention
provides isolated native sequence PR0474 polypeptide, which in one embodiment,
includes an amino acid sequence
c~n~prising residues 1 through 270 of Figure 196 (SEQ ID N0:468). Optionally,
the PR0474 polypeptide is obtained
or is obtainable by expressing the polypeptide encoded by the cDNA insert of
the vector deposited on May 14, 1998
with the ATCC as DNA56045-1380.
78. PR 1 1
Applicants have identified a cDNA clone that encodes a novel polypeptide
having sequence identity with
IL-17, wherein the polypeptide is designated in the present application as
"PR01031".
In one embodiment, the invention provides an isolated nucleic acid molecule
comprising DNA encoding a
PR01031 polypeptide. In one aspect, the isolated nucleic acid comprises DNA
encoding the PR01031 polypeptide
having amino acid residues 1 through 180 of Figure 198 (SEQ ID N0:470), or is
complementary to such encoding
nucleic acid sequence, and remains stably bound to it under at least moderate,
and optionally, under high stringency
conditions. The isolated nucleic acid sequence may comprise the cDNA insert of
the vector deposited on May 14,
1998 with the ATCC as DNA~9294-1381 which includes the nucleotide sequence
encoding PR0103I.
In another emboditrtear, the invention provides isolated PROI031 polypeptide.
In particular, the invention
provides isolated native sequence PR01031 polypeptide, which in one
embodiment, includes an amino acid sequence
comprising residues 1 through 180 of Figure 198 (SEQ ID N0:470). Optionally,
the PROI031 polypeptide is
obtained or is obtainable by expressing the polypeptide encoded by the cDNA
insert of the vector deposited on May
14, 1998 with the ATCC as DNA59294-1381.
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79. P O 3
Applicants have identified a cDNA clone that encodes a navel polypeptide
having sequence identity to
protein disulfide isomerase,. wherein the polypeptide is designated in tine
pre sent application as "PRO938".
In one embodiment, the invention provides an isolated nucleic acid molecule
comprising DNA encoding a
PR0938 polypeptide. In one aspect, the isolated nucleic acid comprises DNA
encoding the PR0938 polypeptide
having amino acid residues 1 to 349 of Figure 200 (SEQ ID N0:472), or is
complementary to such encoding nucleic
acid sequence, and remains stably bound to it under at least moderate, and
optionally, under high stringency
conditions. In other aspects, the isolated nucleic acid comprises DNA encoding
the PR0938 polypeptide having
amino acid residues about 23 to 349 of Figure 200 (SEQ ID N0:472) or amino
acid 1 or about 23 to X of Figure 200
(SEQ ID N0:472), where X is any amino acid from 186 to 195 of Figure 200 (SEQ
ID N0:472), or is
complementary to such encoding nucleic acid sequence, and remains stably bound
to it under at least moderate, and
optionally, under high stringency conditions. The isolated nucleic acid
sequt:nce may comprise the cDNA insert of
the DNA56433-1406 vector deposited on May 12, 1998, as ATCC Accession No.
209857 which includes the
nucleotide sequence encoding PR0938:
In another embodiment, the invention provides isolated PR0938 polypeptide. In
particular, the invention
I5 provides isolated native sequence PR0938 polypeptide, which in one
embodiment, includes an amino acid sequence
comprising residues 1 to 349 of Figure 200 (SEQ ID N0:472). Additional
embodiments of the present invention are
directed to PR0938 polypeptides comprising amino acids about 23 to 349 of
Figure 200 (SEQ ID N0:472) or amino
acid 1 or about 23 to X of Figure 200 (SEQ ID N0:472), where X is any amino
acid from 186 to 195 of Figure 200
(SEQ 1D N0:472). Optionally, the PR0938 polypeptide is obtained or is
obtainable by expressing the polypeptide
encoded by the cDNA inserrof the DNA56433-1406 vector deposited on May I2,
1998, as ATCC Accession No:
209857.
80. PR 1082
Applicants have identified a cDNA clone that encodes a novel polypeptide
having sequence identity with
a iectin-Like oxidized LDL receptor, wherein the polypeptide is designated irt
the present application as °PR01082".
In one embodiment, the invention provides an isolated nucleic acid molecule
comprising DNA encoding a
PR01082 polypeptide. In one aspect, the isolated nucleic acid comprises DNA
encoding the PR01082 poIypeptide
having amino acid residues 1, through 201 of Figure 202 (SEQ ID N0:477), or is
complementary to such encoding
nucleic acid sequence, and remains stably bound to is under at least moderate,
and optionally, under high stringency
conditions. The isolated nucleic acid sequence may comprise the cDNA insert of
the vector deposited on May 14,
1998 with the ATCC as DNA53912-1457 which includes the nucleotide sequence
encoding PR01082.
In another embodiment, the invention provides isolated PR01082 polypeptide. In
particular, the invention
provides isolated native sequetxe PR01082 polypeptide, which in one
embodiment, includes an amino acid sequet~e
comprising residues 1 through 201 of Figure 202 (SEQ ID N0:477). An additional
embodiment of the present
invention is directed to an isolated domain of a PR01082 polypeptide,
excluding the transtnembrane domain.
Optionally, the PR01082 polygeptide is obtained or is obtainable by expressing
the polypeptide encoded by the cDNA
insert of the vector deposited on May 14, 1998 with the ATCC as DNA53912-145?.
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81. PR_ 01083
Applicants have identified a cDNA clone that encodes a novel polypeptide
having sequence identity with
a 7TM receptor, latrophilin-related protein 1, and a macrophage restricted
cell surface glycoprotein, wherein the
poiypeptide is designated in the present application as "PR01083".
In one embodiment, the invention provides an isolated nucleic acid molecule
comprising DNA encoding a
PR01083 polypeptide. In one aspect, the isolated nucleic acid comprises DNA
encoding the PR01083 polypeptide
having amino acid residues 1 through 693 of Figure 204 (SEQ ID N0:483), or is
complementary to such encoding
trucleic acid sequence, and remains stably bound to it under at ieasi
moderate, and optionally, under high stringency
conditions. The isolated nucleic acid sequence may comprise the cDNA insert of
the vector deposited on May 12,
1998 with the ATCC as DNA50921-1458 which includes the nucleotide sequence
encoding PR01083.
In another embodiment, the invention provides isolated PR01083 polypeptide. In
particular, the invention
provides isolated native sequence PR01083 polypeptide, which in one
embodiment, includes an amino acid sequence
comprising residues 1 through 693 of Figure 204 (SEQ ID N0:483). An additional
embodiment of the present
invention is directed to an isolated exuacellular domain of a PR01083
polypeptide. Optionally, the PR01083
polypeptide is obtained or is obtainable by expressing the polypeptide encoded
by the cDNA insert of the vector
deposited on May 12, 1998 with the ATCC as DNA50921-1458.
In another embodiment, the invention provides an expressed sequence tag (EST)
designated herein as
DNA24256 which comprises the nucleotide sequence of Figure 205 (SEQ ID
N0:484).
82. PR020Q,
The objects of this invention, as defined generally supra, are achieved at
least in part by the provision of
a novel polypeptide, VEGF-E also herein designated PR0200, (SEQ ID N0:488) and
the nucleic acid encoding
therefor, SEQ ID N0:487, residues 259 through 1293.
In one embodiment, the invention provides an isolated nucleic acid molecule
comprising DNA encoding a
VEGF-E polypeptide. In one aspect, the isolated nucleic acid comprises DNA
encoding the VEGF-E polypeptide
having amino acid residues 1 through 345 of Figure 207 (SEQ iD N0:488), or is
complementary to such encoding
nucleic acid sequence, and remains stably bound to it under low stringency
conditions. In another embodiment,
variants are provided wherein the VEGF-E nucleic acid has single or multiple
deletions, substitutions, insertions,
truncations or combinations.thereof.
In another embodiment, the invention provides isolated VEGF-E polypepiide. In
particular, the invention
provides an isolated native sequence VEGF-E polypeptide, which in one
embodiment; includes an amino acid
sequence comprising residues 1 through 345 of Figure 20? (SEQ ID N0:488). In
another embodiment, variants are
provided wherein the VEGF-E polypeptide has single or multiple deletions,
substitutions, insertions, tnutcations or
combinations thereof:
In yet further embodiments, the present invention is directed to compositions
useful for treating indications
where proliferation, survival and/or differentiation of cells is desired,
comprising a therapeutically effective amount
of a VEGF-E polypeptide hereof in admixture with a pharmaceutically acceptable
carrier.
The invention fi>rlhes include<c associated embodiments of VEGF-E such as
modified VEGF-E polypeptides
and modified variants which have the same biological applications as VEGF-E,
and pharmaceutical compositions
incorporating same. Inhibitors of VEGF-E are also provided.
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83. 1aR0285 ar~c~,PR0286
Applicants have identified two novel cDNA clones that encode novel human Toll
polypeptides, designated
in the presem application as PR0285 (encoded by DNA4002I-1154) and PR0286
(encoded by DNA42663-1154).
In one embodiment, the invention provides an isolated nucleic acid molecule
comprising a DNA encoding
a polypeptide having at Ieast about 80% sequence identity, preferably at least
about 85% sequence identity, more
preferably at least about 90% sequence identity, most preferably at least
about 95% sequence identity to (a) a DNA
molecule encoding a PR0285 polypeptide having amino acid residues 27 to 839 of
Figure 209 (SEQ ID N0:496);
or (b) to a DNA tnolecuIe encoding a PR0286 polypeptide having amino acid
residues 27 to 825 of Figure 211 (SEQ
ID N0:498) or (c) the complement of the DNA molecule of (a) or (b). The
complementary DNA molecule preferably
remains stably bound to such encoding nucleic acid sequence under at least
moderate, and optionally, under high
stringency conditions.
In a further embodiment, the isolated nucleic acid molecule comprises a
polynucleotide that has at least about
90%, preferably at least about 95% sequence identity with a polynucleotide
encoding a polypeptide comprising the
sequence of amino acids 1 to 839 of Figure 209 (SEQ ID N0:496j; or at least
about 90%, preferably at least about
95 % sequence identity with a polynucleotide encoding a polypeptide comprising
the sequence of amino acids 1 to
1041 of Figure 211 (SEQ ID N0:498).
In a specific embodiment, the invention provides an isolated nucleic acid
molecule comprising DNA
encoding native or variant PR0285 and PR0286 polypeptides, with or wifhout the
N-teraur~al. signal sequence, and
with or without the transmetrtbrane regions of the respective full-length
sequences. In one aspect, the isolated nucleic
acid comprises DNA encoding a mature, full-length native PR0285 or PR0286
polypeptide having amirw acid
residues 1 to 1049 of Figure 209 (SEQ ID N0:496) and 1 to 1041 of Figure 211
(SEQ ID NO: 498), or is
complementary to such encoding nucleic acid sequence, In another aspect, the
invention concerns an isolated nucleic
acid molecule that comprises DNA encoding a native PR0285 or PR028b
polypeptide without an N-terminal signal
sequence, or is complementary to such encoding nucleic acid sequence. In yet
another embodiment, the invention
concerns nucleic acid et~oding transmembrane-domain deleted or inactivated
forms of the full-length native PR0285
ZS or PR0286 proteins.
In another embodiment, the invention the isolated nucleic acid molecule
comprises the clone (DNA40021-
1154) deposited on October 17, 1997, under ATCC number 209389; or the clone
(DNA42663-1154) deposited on
October 17, 1997, under ATCC number 209386.
In yet another embodiment, the invention provides a vector comprising DNA
encoding PR0285 and PR0286
polypeptides, or their variants. Thus, the vector may comprise any of the
isolated nucleic acid molecules hereinabove
defined.
In another embodi>nettt, the invention provides isolated PR0285 and PR0286
polypeptides. In particular,
the invention provides isolated native sequetxe PR0285 and PR0286
polypeptides, which in one embodiment, include
the amino acid sequences comprising residues 1 to 1049 and 1 to 1041 of
Figures 209 and 21 I (SEQ ID NOS:496
and 498), respectively. The invention also provides for variants of the PR0285
and PR0286 polypeptides which are
encoded by any of the isolated nucleic acid molecules hereinabove defined.
Specific variants include, but are not
limited to, deletion (tnmcated) variants of the full-length native sequence
PR0285 and PR0286 polypeptides which
lack the respective N-terminal signal sequences andlor have their respective
transmembrane and/or cytoplasmic
domains .deleted or inactivated.
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The invention also specifically includes antibodies with dual specificities,
e.g., bispecifrc antibodies binding
more than one Toll polypeptide.
In yet another embodiment, the invention concerns agonists and antagonists of
the native PR0285 and
PR0286 polypeptides. In a particular embodiment, the agonist or antagonist is
an anti-PR028S or anti-PR0286
antibody. .
In a further embodiment, the invention concerns screening assays to identify
agonists or antagonists of the
native PR028S and PR0286 polypeptides.
In a still further embodiment, the invention concerns a composition comprising
a PR0285 or PR0286
polypeptide, or an agonist or antagonist as hereinabove defined, in
combination with a pharmaceutically acceptable
carrier.
The invention further concerns a composition comprising an antibody
specifically binding a PR0285 or
PR0286 polypeptide, in combination with a pharmaceutically acceptable carrier.
The invention also concerns a method of treating septic shock comprising
administering to a patient an
effective amount of an antagonist of a PR028S or PR0286 polypeptide. In a
specific embodiment, the antagonist
is a blocking antibody specifically binding a native PR0285 or PR0286
polypeptide.
IS
84. PR0213-13,~R01330 and PR01449
The present invention concerns compositions and methods for the diagnosis and
treatment of neoplastic cell
growth and proliferation in mammals, including humans. The present invention
is based on the identification of genes
that are amplified in the genome of tumor cells. Such gene amplification is
expected to be associated with the
overexpression of the gene product and contribute to tumorigenesis.
Accordingly, the proteins encoded by the
amplified genes are believed to be useful targets for the diagnosis andlor
treatment (including prevention) of certain
cancers, and may act as predictors of the prognosis of tumor treatment.
In one embodiment, the present invention provides an isolated nucleic acid
molecule comprising DNA encoding a
PR0213-1, PR01330 and/or PR01449 polypeptide. In one aspect, the isolated
nucleic acid comprises DNA
encoding the PR0213-1, PRO1330 andlor PR01449 polypeptide having amino acid
residues 1-to 29S of Figure 213
(SEQ ID NO:S06), 20 to 2?3 of Figure 21S (SEQ ID NO:SOB) and 20 to 273 of
Figure 217 (SEQ ID NO:S10),
respectively, or is complementary to such encoding nucleic acid sequence, and
remains stably bound to it under at
least rnaderate; and optionally, under high stringency conditions. The
isolated nucleic acid sequence may comprise
the eDNA insert of the vector designated as DNA30943-1163 (ATCC 209791)
deposited on April 21, 1998;
DNA64907-1163-1 (ATCC 203242) deposited on September 9, 1998 and/or DNA64908-
1163-1 (ATCC 203243)
deposited on September 9, 1998.
In another embodiment, the present invention comprises an isolated nucleic
acid molecule having at least
about 80% sequence identity, preferably at Ieast about 85% sequence identity,
more preferably at least about 90~
sequence idemity, most preferably at least about 9S~ sequence identity to (a)
a DNA molecule encoding a PR0213-1,
PR01330 andlor FR01449 polypeptide having amino acid residues 1 to 295 of
Figure 213 (SEQ ID NO:S06), 20
to 273 of Figure 215 (SEQ ID N0:508) and 20 to 273 of Figure 217 (SEQ ID
NO:S10), respectively; or (b) the
complement of the DNA molecule of (a).
In another embodiment, the invention provides ~an isolated PR0213-1, PR01330
andlor PR01449
polypeptide. In.particular, the invention provides isolated native sequence
PR0213-1, PR01330 and/or PR01449
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polypepride, which in one embodirrtent, includes an amino acid sequence
comprising residues 1 to 29S of Figure 213
{SEQ ID NO:S06), 20 to 273 of Figure 215 (SEQ ID N0:508) or 20 to 273 of
Figure 217 (SEQ ID NO:S10),
respectively. Optionally, the PR0213-1, PR01330.and/or PROI449 poiypeptide is
obtained or obtainable by
expressing the polypeptide encoded by the cDNA insert of the DNA30943-1 I63
(ATCC 209791), DNA64907-I 163-I
(ATCC 203242) or DNA64908-l I63-I {ATCC 203243).
In another aspect, the invention provides an isolated PR0213-I, PR01330,
andlor PR01449 poly~peptide,
comprising an amino acid sequence having at least about 80% sequence identity,
preferably at least about 8596
seguence identity, more preferably at least about 95 % sequence identity to
amino acid residues 1 to 29S of Figure
213 (SEQ ID NO:S06), 20 to 273 of Figure 21S (SEQ ID NO:S08) or 20 to 273 of
Figure 2I7 (SEQ ID NO:S10),
inclusive.
In yet another embodiment, the invention provides an isolated PR0213-1,
PR01330, andlor PROI449
polypeptide, comprising the amino acid residues I to 29S of Figure 213 (SEQ ID
NO:S06), 20 to 273 of Figure 2IS
(SEQ ID N0:508) or 20 to 273 of Figure 217 (SEQ ID NO:S10), or a fragment
thereof sufficient to provide a binding
site for an anti-PR0213-1, anti-PROI330 andlor anti-PROI449 antibody.
Preferably, the PR0213-1, PR01330,
and/or PR01449 fragment retains a qualitative biological activity of a native
PR0213-1, PROi330, andlor PR01449
polypeptide.
1n a further aspect, tip invention concerns an isolated PR0213-1, PR01330,
and/or PR01449 polypeptide,
comprising an amino acid sequence scoring at least about 80% positives,
preferably at least about 85 q6 positives,
more preferably at least about 90~ positives, most preferably at least about
95 96 positives when compared with the
amino acid sequence of residues 1 to 29S of Figure 213 (SEQ ID NO:SO~, 20 to
273 of Figure 215 (SEQ ID
NO:S08) and 20 to 273 of Figure 21? (SEQ ID NO:S10), respectively.
In still a further aspect, the invention provides a polypeptide produced by
(l) hybridizing a test DNA
~lecule under stringent conditions with: (a) a DNA molecule encoding a PR0213-
1, PR01330, andlor PR01449
polypeptide having the amino acid residues from 1 to 29S of Figure 213 (SEQ ID
NO:S06), 20 to 273 of Figure 21S
(SEQ ID NO:S08) and 20 to 273 of Figure 217 (SEQ ID NO:S10), respectively; or
the complement of the DNA
2S molecule of (a), and if said test DNA molecule has at least about an 80 %
seguence identity to (a) or (b); (ii) culturing
a host cell comprising said test DNA molecule under conditions suitable for
the expression of said polypeptide, and
(iii) recovering said polypeptide froth the cell culnue.
In one embodiment, the present invention concerns an isolated antibody which
binds a PR0213-1, PROI330
andlor PR01449 polypeptide. In one aspect, the antibody induces death of a
cell oyerexpressing a PR0213-1,
PR01330 andlor PROI449 polypeptide. In another aspect, the antibody i,s a
monoclonal antibody, which preferably
has nonhuman complememarity determining region (CDR) residues and human
framework region {,FR) residues,
The antibody may be labeled ate may be immobilized on a solid support. In a
further aspect, the antibody is an
antibody fragment; a single-chain antibody, or an anti-idiotypic antibody.
In another embodiment, the invention concerns a composition comprising an
antibody which binds a
3S PR0213-1, PR01330 atxllor PR01449 polypeptide in admixture with a
phatynaceutically acceptable carrier. In one
aspect, the composition comprises a therapeutically effective amount of the
antibody. In another aspect, the
composition comprises a further active ingredient; which may, for example, be
a further antibody or a cytotoxic or
clLemotherapeutic agent. Preferably, the composition is sterile.
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In a further embodiment, the invention concerns nucleic acid encoding an anti-
PR0213-1, anti-PR01330
and/or anti-PROla49 antibody, and vectors and recombinant host cells
comprising such nucleic acid.
The invention further concerns antagonists and agonists of a PR0213-1, PR01330
and/or PR01449
polypeptide that inhibit one or more of the functions or activities of the
PR0213-1, PR01330 andlor PR01449
polypeptide.
In a further embodiment, the invention concerns isolated nucleic acid
molecules that hybridize to the
complement of the nucleic acid molecules encoding the PR0213-1, PR01330 andlor
PR01449 polypeptides. The
nucleic acid preferably is DNA, and hybridization preferably occurs under
stringent conditions. Such nucleic acid
molecules can act as antisense molecules of the amplified genes identified
herein, which, in turn, can find use in the
modulation of the respective amplified genes, or as antisense primers in
amplification reactions. Furthermore, such
sequences can be used as part of ribozyme andlor triple helix sequence which,
in turn, may be used in regulation
of the amplified genes.
In another embodiment, the invention concerns a method for determining the
presence of a PR02I3-1,
PROI330 andlor PRO1449 polypeptide comprising exposing a cell suspected of
containing the PR0213-1, PR01330
and/or PR01449 polypeptide to an anti-PR02I3-1, PR01330 andlor PROI449
antibody and determining binding of
the antibody to the cell.
In yet another embodiment, the present invention concerns a method of
diagnosing tumor in a matnrttal,
comprising detecting the level of expression of a gene encoding a PR0213-I,
PR01330 and/or PR01449 polypepdde
(a) in a test sample of tissue cells obtained from the mammal, and (b) in a
control sample of known normal tissue cells
of the same cell type, wherein a higher expression level in the test sample
indicates the presence of tumor iu the
mammal from which the test tissue cells were obtained.
In another embodiment, the present invention concerns a method of diagnosing
tu~r in a mammal,
cantprising (a) contacting an anti-FR0213-1, anti-PR01330 andlor anti-PR01449
antibody with a test sample of tissue
cells obtained from the mammal, and (b) detecting the formation of a complex
between the anti PR0213-1, anti-
PR01330 and/or anti-PR01449 antibody and the PR0213-1, PR01330 and/or PR01449
polypeptide in the test
sample. The detection may be qualitative or quantitative, and may be performed
in comparison with monitoring the
complex formation in a control sample of known normal tissue cells of the same
cell type. A larger quantity of
complexes formed in the test sample indicates the presence of tumor in the
mammal from which the test tissue cells
were obtained. The antibody preferably carries .a detectable label. Complex
formation can be monitored, for
example, by light microscopy, flaw cytometry, fluorimetry, or other techniques
known in the art. The test sample
is usually obtained from an individual suspected to have neoplastic cell
growth or proliferation (e.g. cancerous cells).
In another embodiment, the present invention rnncerns a cancer diagnostic kit,
comprising an anti-PR0213-
1, anti-PR01330 andlor anti-PR01449 atuibody and a carrier (e.g. a buffer) in
suitable packaging. The kit preferably
contains instructions for using the antibody to detect the PR0213-1, PR01330
andlor PR01449 polypeptide.
In yet another embodiment, the invention concerns a method for inhibiting the
growth of tumor cells
cotnptising exposing a cell which overexpresses a PR0213-1, PR01330 and/or
PROI4419 polypeptide to an effective
amount of an agent inhibiting the expression andlor activity of the PR0213-1;
PR01330 andlor PROlA49
polypeptide. The agent preferably is an anti-PR0213-1, anti-PR01330 and/or and-
PR01449 anu'body, a small
organic and inorganic molecule, peptide, phosphopeptide, antisense or ribozyme
molecule, or a triple helix molecule.
In a specific aspect, the agent, e.g. anti-PR0213-1, anti-PR01330 and/or anti-
PR01449 antibody induces cell death.
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In a further aspect, the tumor cells are further exposed to radiation
treatment and/or a cytotoxic or chemotherapeutic
agent.
In a further embodiment, the invention concerns an article of manufacture,
comprising:
a) a container;
b) a label on the container; and
c) a composition comprising an active agent contained within the container;
wherein the composition is
effective for inhibiting the growth of tumor cells, the label on the container
indicates that the composition can be used
for treating conditions characterized by overexpression of a PR0213-1, PR01330
and/or PR01449 polypeptide, and
the active agent in the composition is an agent inhibiting the expression
andlor activity of the PR0213-1, PR01330
andlor PR01449 polypeptide. In a preferred aspect, the active agent is an anti-
PR0213-1, anti-PR01330 andlor anti
PR01449 antibody.
In yet a further embodiment, the invention provides a method for identifying a
compound capable of
inhibiting the expression andlor activity of a PR0213-1, PR01330 and/or
PR01449 polypeptide, comprising
contacting a candidate compound with a PR0213-1, PR01330 andlor PR01449
polypeptide under conditions and
for a tire sufficient to allow these two components to interact. . In a
specific aspect, either the candidate compound
or the PR0213-1, PR01330 andlor PR01449 polypeptide is immobilized on a solid
support. In another aspect, the
non immobilized component carries a detectable label.
85. PR0298
Applicants have identified a cDNA clone that encodes a novel polypeptide. The
DNA is designated in the
present application as "DNA39975-1210", encoding a novel mufti-transmembxane
protein, referred to as "PR0298".
1n one embodiment, the invention pnwides an isolated nucleic acid molecule
comprising DNA having at least
about 809& , preferably at least about 85 ~, more preferably at least about 90
%, most preferably at least about 95 %
sequence identity to (a) a DNA molecule encoding PR0298, comprising the
sequence of amino acids 1 to 364 of
Figure 219 (SEQ ID NO:515), or (b) the complett~nt of the DNA molecule of (a).
1n one aspect, the isolated nucleic
acid comprises DNA encoding a PR0298 polypeptide having amino acid residues 1
to 364 of Figure 219 (SEQ ID
NO:515}, or is complerrterimry to such encoding nucleic acid sequence, and
remains stably bound to it under at least
moderate, and optionally, under high stringency conditions.
In a further embodiment, the invention concerns an isolated nucleic acid
molecule comprising DNA having
at least as 809b sequence identity to (a) a DNA molecule encoding the same
tnataue polypeptide encoded by the
human prosein cDNA in ATCC Deposit No. 209783 (DNA39975-12I0); or (b) the
complement of the DNA molecule
of (a).
In a still further embodiment, the invention concerns mtcieic acid which
comprises a DNA molecule
encoding the same mature polypeptide encoded by the human protein cDNA in ATCC
Deposit No. 209783
(DNA39975-1210).
In another embodiment, the invention provides isolated PR029?3 polypeptide. In
particular, the invention
provides isolated native sequence PR0298 polypeptide, which in one embodiment,
includes as amino acid sequence
comprising residues 1 to 364 of Figure 219 (SEQ ID NO:515).
in another embodiment, the invention provides an expressed sequence tag (EST)
designated DNA26832
comprising the nucleotide sequence of Figure 220 (SEQ ID N0:516).
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86. PR 337
Applicants have identified a cDNA clone (DNA43316-1237) that encodes a novel
polypeptide, designated
in the present application as- "PR0337".
In one embodiment, the invention provides an isolated nucleic acid molecule
having at least about 80%
sequence identity to (a) a DNA molecule encoding a PR0337 polypeptide
comprising the sequence of amino acids
1 to 344 of Figure 222 (SEQ ID N0:523), or (b) the cotrtplement of the DNA
molecule of (a). The sequence Identity
preferably is about 85%, more preferably about 90%, most preferably about
95'0. In one aspect, the isolated nucleic
acid has at least about 803'0, preferably at least about 85 %, more preferably
at least about 90%, and most preferably
at least about 95 (including 96, 97> 98 and 99%) sequence identity with a
polypeptide having amino acid residues 1
to 344 of Figure 222 (SEQ ID N0:523). Preferably, the highest degree of
sequence identity occurs within the
itnmunoglobulin and major histocompatibility domains (amino acids 113 to 130
of Figure 222, SEQ ID N0:523).
In a further embodiment, the isolated nucleic acid molecule comprises DNA
encoding a neurouimin
polypeptide haying amino acid residues 1 to 344 of Figure 222 (SEQ ID N0:523),
or is complementary to such
encoding nucleic acid sequence, and remains stably bound to it under at least
moderate, and optionally, under high
stringency conditions. In another aspect, the invention provides a nucleic
acid of the full length protein of clone
DNA43316-1237, deposited with the ATCC under accession number ATCC 209487,
alternatively the coding
sequence of clone DNA43316-1237, deposited under accession number ATCC 209487.
In yet another embodiment, the invention provides isolated PR0337 polypeptide.
In particular, the invention
provides isolated native sequence PR0337 polypeptide, which in one embodiment,
includes an amino acid sequence
comprising residues 1 to 344 of Figure 222 (SEQ ID N0:523). Native PR0337
polypeptides with or without the
native signal sequence (amino acids 1 to about 28 in Figure 222 (SEQ ID
N0:523), and with or without the initiating
methionine are specifically included. Alternatively, the invention provides a
PR0337 poIypeptide encoded by the
nucleic acid deposited under accession number ATCC 20948?.
In yet another embodiment, the invention provides an expressed sequence tag
(EST) comprising the
nucleotide sequences identified in Figure 223 as DNA42301 (SEQ ID N0:524).
87. PR0403
Applicants have identified a cDNA clone (DNA55800-1263) that encodes a novel
polypeptide, designated
in the present application as "PR0443".
In one embodiment, the invention provides an isolated nucleic acid molecule
having at least about 809'0
sequence identity to (a) a DNA molecule encoding a PR0403 palypeptide
comprising the sequence of amino acids
1 to ?36 of Figure 225 (SEQ ID N0:526), or (b) the complement of the DNA
molecule of (a). The sequence identity
preferably is about 8S%, more preferably about 90%, most preferably about 95
%. In one aspect, the isolated nucleic
acid has at least about 80%, preferably at least about 85 % , more preferably
at least about 90% , and most preferably
at least about 95 % sequence identity with a poiypeptide having amino acid
residues 1 to 736 of Figure 225 (SEQ ID
NO:526). Preferably, the highest degree of sequence identity occurs within:
(1) the putative N-glycosylatation sites
{amino acid residues I32, 136,177, 237, 282, 349, 505, 598 and 606; {2) Cys
residues conserved with the Kell blood
group protein family (amino acid residues 65, 70, 88 and 96) and the putative
zinc binding motif (amino acid residues
570 579).
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In a further embodiment, the isolated nucleic acid molecule comprises 1f?NA
encoding a PR04a3 polypeptide
having amino acid residues 1 to 736 of Figure 225 {SEQ ID N0:526), or is
complementary to such encoding nucleic
acid sequence, and remains stably bound to it under at least moderate, and
optionally; under high stringency
conditions. In another aspect, the invention provides a nucleic acid of the
full length protein of clone DNA55800-
1263, deposited with the ATCC under accession number ATCC 209680,
alternatively the coding sequence of clone
DNA55800-1263, deposited under accession number ATCC 209680.
In yet another embodiment, the invention provides isolated PR0403 polypeptide.
In particular, the invention
provides isolated native sequence PR0403 polypeptide, which in one embodiment,
includes an amino acid sequence
comprising residues 1 to 736 of Figure 225 (SEQ ID N0:526). Native PRO403
polypeptides with or the initiating
methionine are specifically included. Alternatively, the invention provides a
PR0403 polypeptide encoded by the
nucleic acid deposited under accession number ATCC 209680.
In yet another embodiment, the invention provides an expressed sequence tag
(EST)~and other sequence
fragtnentc comprising the nucleotide sequences identified herein as DNA344I5
(Figures 226A-B; SEQ ID N0:527);
DNA49830 (Figure 227 ; SEQ ID N0:528) and DNA49831 (Figure 228; ~EQ ID
N0:529).
88. Additional Embodimen
In other embodiments of the present invention, the invention provides vectors
comprising DNA encoding
any of the above or below described polygeptides. A host cell comprising any
such vector is also provided. By way
of example, the hit cells may be CHO cells, E. coli, or yeast. A process for
producing any of the above or below
described polypeptides is further provide and comprises culturing host cells
under conditions suitable for expression
of the desired polypeptide and recovering the desired polypeptide from the
cell culture.
In other embodiments, the invention provides chimeric molecules comprising any
of the above or below
descn'bed polypepttdes fused to a heterologous polypeptide or amino acid
sequence. An example of such a chimeric
molecule comprises arty of the above or below described palypeptides fused to
an epitope tag sequence or a Fc region
of an immunoglobulin.
In artottrer embodiment, the invention provides an antibody which specifically
binds to arty of the above or
below described polypeptides. Optionally, the antibody is a monoclonal
antibody.
In yet other embodiments, the invention provides oligonucleotide probes useful
for isolating genomic and
cDNA nucleotide sequences, wherein those probes may be derived from any of the
above or below described
nucleotide sequences.
$RIEF DFSs:.~~Q~~,~>V~~
Figure 1 shows a nucleotide sequence (SEQ ID NO:I) of a native sequence PRO2I3
cDNA, wherein SEQ
ID NO:1 is a clone designated herein as "UNQ187" andlor "DNA30943-1163".
Figtue 2 shows the amino acid sequence (SEQ 1D N0:2) derived from the coding
sequence of SEQ ID NO;1
shown in Figure I .
Figure 3 shows a nucleotide sequence (SEQ ID N0:6) of a native sequence PRO274
cDNA, wherein SEQ
ID N0:6 is a clone designated herein as "UNQ241" and/or "DNA39987-1184".
Figure 4 shows the amino acid sequence (SEQ >D N0:7) derived from the coding
sequence of SEQ ID N0:6
shown in Figure 3.
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Figure S shows an EST nucleotide sequence designated herein as DNA17873 (SEQ
ID N0:8).
Figure 6 shows an EST nucleotide sequence designated herein as DNA36157 (SEQ
ID N0:9).
Figure ? shows an EST nucleotide sequence designated herein as DNA28929 (SEQ
ID NO:10}.
Figure 8 shows a nt~clwtide sequence (SEQ ID N0:18) of a native sequence
PR0300 cDNA, wherein SEQ
ID N0:18 is a clone designated herein as "UNQ263" andlor "DNA40625-1189".
Figure 9 shows the amino acid sequence (SEQ ID N0.19) derived from the coding
sequence of SEQ ID
N0:18 shown in Figure 8.
Figure 10 shows a nucleotide sequence (SEQ ID N0:27) of a native sequence
PR0284 cDNA, wherein SEQ
ID N0:27 is a clone designated herein as "UNQ247" and/or "DNA23318-1211".
Figure 11 shows the amino acid sequence (SEQ ID N0:28) derived from the coding
sequence of SEQ ID
N0:27 shown in Figure 10.
Figure 12 shows an EST nucleotide sequence designated herein as DNAI2982 (SEQ
1D N0:29).
Figure 13 shows an EST nucleotide sequence designated herein as DNAI5886 (SEQ
ID N0:30).
Figtzne 14 shows a nucleotide sequence ($EQ ID N0:35) of a native sequence
PR0296 cDNA, wherein SEQ
ID N0:35 is a clone designated herein as "UNQ260" andlor "DNA39979-1213".
Figure 15 shows the amino acid sequence (SEQ ID N0:36) derived from the coding
sequence of SEQ ID
N0:35 shown in Figure 14.
Figure 16 shows an EST nucleotide sequence designated herein as DNA23020 (SEQ
ID N0:3?).
Figure 17 shows an EST nucleotide sequence designated herein as DNA2197I (SEQ
ID N0:38).
Figure 18 shows an EST nucleotide sequence designated herein as DNA29037 {SEQ
ID N0:39).
Figure 19 shows a nucleotide sequence (SEQ ID N0:44) of a native sequence
PR0329 cDNA, wherein SEQ
ID N0:44 is a clone designated herein as "UNQ291 " and/or "DNA40594-1233".
Figure 20 shows the amino acid sequence (SEQ ID N0:45) derived from the coding
sequence of SEQ ID
N0:44 shown in Figure 19.
Figure 21 shows a nucleotide sequence (SEQ ID NO:51) of a native sequence
PR0362 cDNA, wherein SEQ
1D NO:51 is a clone designated herein as "UNQ317" and/or "DNA454I6-125I"
Figure 22 shows the amino acid sequence (SEQ ID N0:52} derived from the coding
sequence of SEQ ID
NO:51 shown in Figure 21.
Figure 23 shows a nuchtide sequence (SEQ ID N0:58) of a native sequence PR0363
cDNA, wherein SEQ
ID N0:58 is a clone designated herein as "UNQ318" and/or "DNA45419-1252" .
Figure 24 shows the amino acid.sequence (SEQ ID N0:59) derived from the coding
sequence of SEQ ID
NO:58 shown in Figure 23.
Figure 25 shows a nucleotide sequence (SEQ U7 NO:b3) of a native sequence
PR0868 cDNA, wherein SEQ
ID N0:63 is a clone designated herein as "UNQ437" and/or "DNA52594-1270".
Figure 26 shows the amino acid sequence (SEQ iD N0:64) derived from the coding
sequence of SEQ ID
N0:63 shown in Figure 25.
Figure 27 shows a nucleotide sequence (SEQ ID N0:68) of a native sequence
PR0382 cDNA, wherein SEQ
ID N0:68 is a clone designated herein as "UNQ323" andlor "DNA45234-1277".
Figure 28 shows the amino acid sequence (SEQ ID N0:69) derived from the coding
sequence of SEQ ID
N0;68 shown in Figure 27.
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Figure 29 shows a nucleotide sequence (SEQ ID N0:73) of a native sequence
PR054S cDNA, wherein SEQ
ID NO:?3 is a clone designated herein as "UNQ346" and/or "DNA49624-I279".
Figure 30 shows the, amino acid sequence {SEQ ID N0:74) derived :from the
coding sequence of SEQ ID
NO:73 shown in Figure 29.
Figure 31 shows an EST rntcleotide sequence designated herein as DNA13217 (SEQ
ID N0:75).
Figure 32 shows a nucleotide sequence (SEQ ID N0:84) of a native sequence
PR06I7 cDNA, wherein SEQ
ID N0:84 is a clone designated herein as "UNQ353" and/or "DNA48309-1280".
Figure 33 shows the amino acid sequence (SEQ ID N0:85) derived from the coding
sequence of SEQ ID
N0:84 shown in Figure 32.
Figure 34 shows a nucleotide sequence (SEQ ID N0:89) of a native sequence
PR0700 cDNA, wherein SEQ
ID N0:89 is a clone designated herein as "UNQ364" and/or "DNA46776-1284".
Figure 35 shows the amino acid sequence (SEQ ID N0:90) derived from the coding
sequence of SEQ ID
N0:89 shown in Figure 34.
Figure 36 shows a nucleotide sequence (SEQ 1D N0:96) of a native sequence
PR0702 cDNA, wherein SEQ
ID N0:96 is a clone designated herein as "UNQ366" and/or "DNA50980-1286".
Figure 37 shows the amino acid sequence (SEQ ID N0:97) derived from the coding
sequence of SEQ ID
N0:96 shown in Figure 36.
Figure 38 shows a nucleotide sequence (SEQ ID N0:101) of a nauive sequence
PR0703 cDNA, wherein
SEQ ID NO:IOI is a clone designated herein as "UNQ367" and/or "DNA50913-1287".
Figure 39 shows the amino acid sequence {SEQ ID N0:102) dezived from the
coding sequence of SEQ ID
N0:10I shown in Figure 38.
Figure 40 shows a nucleotide sequence (SEQ ID NO:108) of a native sequence
PR0705 cDNA, wherein
SEQ ID N0:108 is a clone designated herein as "UNQ369" and/or "DNA5U914-1289".
Figure 41 stews the amino acid sequence (SF.Q ID N0:109) derived from the
coding sequence of SEQ ID
NO:108 shown in Figure 40.
Figures 42A-B show a nucleotide sequence (SEQ ID N0:113) of a native sequence
PR0708 cDNA, wherein
SEQ ID N0:113 is a clone designated herein as "UNQ372" and/or "DNA48296-1292".
Figure 43 straws the amino acid sequence (SEQ ID N0:114) derived from the
coding sequence of SEQ ID
N0:113 shown in Figures 42A-B.
Figure 44 shows a nucleotide sequence (SEQ ID NO:118) of a native sequence
PR0320 cDNA, wherein
SEQ ID N0:118 is a clone designated herein as "UNQ281" and/or "DNA32284-1307".
Figure 45 shows the amino acid sequence (SEQ ID N0:119) derived from the
coding sequence of SEQ ID
N0:118 shown in Figure 44.
Figure 46 shows a nucleotide sequence (SEQ ID N0:123) of a native sequence
PR0324 cDNA, wherein
SEQ ID NO:123 is a clone designated herein as "UNQ285" and/or "DN,A36343-
1310".
Figure 47 shows the amino acid sequence (SF.Q ID NO:I24) derived from the
coding sequence of SEQ ID
N0:123 shown in Figure 46.
Figure 48 shows a nucleotide sequence (SEQ ID N0:131) of a native sequence
PR035I cDNA, wherein
SEQ ID N0:131 is a clone designated herein as "UNQ308" and/or "DNA40571-1315".
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Figure 49 shows the amino acid sequence (SEQ ID N0:132) derived from the
coding sequence of SEQ ID
N0:131 shown in Figure 48.
Figure 50 shows a nucleotide sequence (SEQ ID N0:136) of a native sequence
PR0352 cDNA, wherein
SEQ ID N0:136 is a clone designated herein as "UNQ309" and/or "DNA41386-1316"
Figure 51 shows the amino acid sequence (SEQ 1D NO:137) derived from the
coding sequence of SEQ ID
NO:I36 shown in Figure 50.
Figure 52 shows a nucleotide sequence (SEQ ID N0:144) of a native sequence
PR0381 cDNA, wherein
SEQ ID N0:144 is a clone designated herein as "UNQ322" andlor "DNA44194-1317".
Figure 53 shows the amino acid sequence (SEQ 1D N0:145) derived from the
coding sequence of SEQ ID
NO:I44 shown in Figure 52.
Figure 54 shows a nucleotide sequence {SEQ ID N0:149) of a native sequence
PR0386 cDNA, wherein
SEQ ID N0:149 is a clone designated herein as "UNQ326" andlor "DNA45415-1318".
Figure 55 shows the amino acid sequence (SEQ ID N0:150) derived from the
coding sequence of SEQ ID
N0:149 shown in Figure 54.
Figure 56 shows an EST nucleotide sequence designated herein as DNA23350 (SEQ
ID N0:151).
Figure 57 shows an EST nucleotide sequence designated herein as DNA23536 (SEQ
ID N0:152).
Figure 58 shows a nucleotide sequence (SEQ ID N0:156) of a native sequence
PR0540 cDNA, wherein
SEQ ID N0:156 is a clone designated herein as "UNQ341" andlor "DNA44189-1322".
Figure 59 shows the amino acid sequence (SEQ ID N0:157) derived from the
coding sequence of SEQ ID
N0:156 shown in Figure 58.
Figure 60 shows a nucleotide sequence {SEQ ID N0:161) of a native sequence
PR0615 cDNA, wherein
SEQ ID N0:16I is a clone designated herein as "UNQ352" and/or "DNA48304-1323".
Figure 61 shows the amino acid sequence (SEQ ID N0:162) derived from the
coding sequence of SEQ ID
NO:161 shown in Figure 60.
Figure 62 shows a nucleotide sequence {SEQ ID N0:168) of a native sequence
PR0618 cDNA, wherein
SEQ ID N0:168 is a clone designated herein as "UNQ354" andlor "DNA49152-1324".
Figure 63 shows the amino acid sequence (SEQ ID N0:169) dexived from the
coding sequence of SEQ ID
N0:168 shown in Figure 62.
Figure 64 shows an EST nucleotide sequence designated herein as DNA35597 (SEQ
ID NO:I70).
Figure 65 shows a nucleotide sequence (SEQ ID N0:17?) of a native sequence
PR0719 cDNA, wherein
SEQ ID N0:17? is a clone designated heiein as "UNQ387" and/or "DNA49646-1327".
Figure 66 shows the amino acid sequence (SEQ ID N0:178) derived from the
coding sequence of SEQ ID
N0:177 shown in Figure 65.
Figure 67 shows a nucleotide sequence (SEQ ID N0:182) of a native sequence
PR0724 cDNA, wherein
SEQ ID NO:182 is a clone designated herein as "UNQ389" and/or "DNA49631-1328".
Figure 58 shows the amino acid sequence (SEQ ID N0:183) derived from the
coding sequence of SEQ ID
N0:182 shown in Figure 67.
Figure 69 shows a nucleotide sequence (SEQ ID N0:189) of a native sequence
PR0772 cDNA, wherein
SEQ ID N0:189 is a clone designated herein as "UNQ410" and/or "DNA4964S-1347".
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Figure 70 shows the amino acid sequence (SEQ ID N0:190) derived from the
coding sequence of SEQ ID
N0:189 shown in Figure 69.
Figure ?1 shows an EST nucleotide sequence designated herein as DNA43509 (SEQ
ID N0:191).
Figure 72 shows a nucleotide sequence (SEQ ID N0:195) of a native sequence
PR0852 cDNA, wherein
SEQ ID N0:195 is a clone designated herein as "UNQ418" andlor "DNA45493-1349".
Figure 73 shows the amino acid sequence (SEQ ID N0:196) derived from the
coding sequence of SEQ ID
N0:195 shown in Figure 72:
Figure 74 shows a nucleotide sequence (SEQ ID N0:205) of a native sequence
PR0853 cDNA, wherein
SEQ ID N0:205 is a clone designated herein as "UNQ419" andlor "DNA48227-1350".
Figure 75 shows the amino acid sequence (SEQ ID N0:206) derived from the
coding sequence of SEQ ID
N0:245 shown in Figure ?4.
Figures 76A-B show a nucleotide sequence (SEQ ID N0:210) of a native sequence
PR0860 cDNA, wherein
SEQ ID N0:210 is a clone designated herein as "UNQ42I" andlor "DNA41404-1352".
Figure 77 shows the amino acid sequence (SEQ ID N0:211) derived from the
coding sequence of SEQ ID
N0:210 shown in Figures 76A-B.
Figure 78 shows a nucleotide sequence (SEQ ID N0:215) of a native sequence
PR0846 cDNA, wherein
SEQ ID N0:215 is a clone designated herein as "UNQ422" andlor "DNA44196-1353".
Figure 79 shows the amino acid sequence (SEQ ID N0:216) derived from the
coding sequence of SEQ ID
N0:215 shown in Figure 78.
Figure 80 shows a nucleotide sequence (SEQ ID N0:220) of a native sequence
PR0862 cDNA, wherein
SEQ ID N0:220 is a clone designated herein as "UNQ424" andlor "DNA52187-1354".
Figure 81 shows the amino acid sequence (SEQ ID N0:221) derived from the
coding sequence of SEQ ID
N0:220 shown in Figure 80.
Figure 82 shows a nucleotide sequence (SEQ ID N0:225) of a native sequence
PR0864 cDNA, wherein
SEQ ID N0:225 is a clone designated herein as "UNQ426" andlor "DNA48328-1355".
Figure 83 shows the amino acid sequence (SEQ ID N0:22b) derived from the
coding sequence of SEQ ID
N0:225 shown in Figure ~,.
Figure 84 shows a nucleotide sequence (SEQ 1D N0:230) of a native sequence
PR0792 cDNA, wherein
SEQ ID N0:230 is a clone designated herein as "UNQ431" andlor °DNA56352-
1358".
Figure 85 shows the amino acid sequence (SEQ ID N0:231) derived from the
coding sequence of SEQ ID
N0:230 shown in Figure 84.
Figure 86 shows a nucleotide sequence (SEQ ID N0:235) of a native sequence
PR0866 cDNA, wherein
SEQ ID N0:235 is a clone designated herein as "UNQ435" andlor "DNA53971-1359".
Figure 87 shows the amino acid sequence (SEQ ID N0:236) derived from the
coding sequence of SEQ ID
N0:235 shown in Figure 86.
Figure 88 shows a nucleotide sequence (SEQ ID N0:244) of a native sequence
PR0871 cDNA, wherein
SEQ ID N0:244 is a clone designated herein as "UNQ438" andlor "DT1A50919-1361
".
Figure 89 shows the amino acid sequence (SEQ ID N0:245) derived from the
coding sequence of SEQ ID
N0:244 shown in Figure 88.
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Figure 90 shows a nucleotide sequence (SEQ ID N0:253) of a native sequence
PR0873 cDNA, wherein
SEQ ID N0:253 is a clone designated herein as "UNQ440" andlor "DNA44179-1362".
Figure 91 shows the. amino acid sequence (SEQ ID N0:254} derived from the
coding sequence of SEQ ID
N0:253 shown in Figure 90.
Figure 92 shows a nucleotide sequence (SEQ ID N0:258} of a native sequence
PR0940 cDNA, wherein
SEQ ID N0:258 is a done designated herein as °UNQ477" and/or "DNA54002-
1367".
Figure 93 shows the amino acid sequence (SEQ ID N0:259) derived from the
coding sequence of SEQ ID
N0:258 shown in Figure 92.
Figure 94 shows a nucleotide sequence (SEQ ID N0:263) of a native sequence
PR0941 cDNA, wherein
SEQ ID N0:263 is a clone designated herein as "UNQ478" and/or "DNA53906-1368".
Figure 95 shows the amino acid sequence (SEQ ID N0:264) derived from the
coding sequence of SEQ ID
NO:263 shown in Figure 94.
Figure 96 shows an EST nucleotide sequence designated herein as DNA6415 (SEQ
ID N0:265).
Figure 97 shows a nucleotide sequence (SEQ ID N0:269) of a native sequence
FR0944 cDNA, wherein
SEQ ID N0:269 is a clone designated herein as "UNQ481" andlor "DNA52185-1370".
Figure 98 shows the amino acid sequence (SEQ ID N0:270} derived from the
coding sequence of SEQ ID
N0:269 shown in Figure 97.
Figure 99 shows an EST nucleotide sequence designated herein as DNA140t17 (SEQ
ID N0:271).
Figure 100 shows an EST nucleotide sequence designated herein as DNA12773 (SEQ
ID N0:272).
Figure 101 shows an EST nucleotide sequence designated herein as DNAi2746 (SEQ
ID N0:273).
Figure 102 shows an EST nucleotide sequence designated herein as DNA12834 (SEQ
ID N0:274).
Figure 103 shows an EST nucleotide sequence designated herein as DNA12846 (SEQ
ID N0:275).
Figure 104 shows an EST nucleotide sequence designated herein as DNA13104 {SEQ
ID N0:276).
Figure 105 shows an EST nucleotide sequence designated herein as DNA13259 (SEQ
ID N0:277).
Figure 106 shows an EST nucleotide sequence designated herein as DNA13959 {SEQ
ID N0:278).
Figure 107 shows an EST nucleotide sequence designated herein as DNA23961 (SEQ
ID N0:279).
Figure 108 shows a nucleotide sequence (SEQ ID N0:283) of a native sequence
PR0983 cDNA, wherein
SEQ ID N0:283 is a clone designated herein as pUNQ484" andlor "DNA5397?-1371".
Figure I09 shows the amino acid sequetxe (SEQ 1D N0:284) derived from the
coding sequence of SEQ ID
N0:283 shown in Figure 108.
Figure I IO shows an EST nucleotide sequence designated herein as DNA17130
(SEQ ID N0:285}.
Figure 11 i shows an EST nucleotide sequence designated herein as DNA23466
(SEQ ID N0:286).
Figure 1 i2 shows an EST nucleotide sequence designated herein as DNA26818
(SEQ 1D N0:287).
Figure 113 shows an EST nucleotide sequence designated herein as DNA37618 (SEQ
ID N0:288).
Figure 114 shows an EST nucleotide sequence designated herein as DNA41732 (SEQ
ID N0:289).
Figure 115 shows an EST nucleotide sequence designated herein as DNA45980 (SEQ
ID N0:290).
Figure 116 shows an EST nucleotide sequence designated herein as DNA463?2 (SEQ
ID N0:291).
Figure 117 shows a nucleotide sequence (SEQ ID N0:295) of a native sequence
PR01057 cDNA, wherein
SEQ ID N0:295 is a clone designated herein as "UNQ522" andlor "DNA57253-1382".
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Figure 118 shows the amino acid sequence (SEQ ID N0:296) derived from the
coding sequence of SEQ ID
N0:295.shown in Figure 117.
Figure l l9 shows a nucleotide sequence {SEQ ID N0:300) of a native sequence
PR01071 cDNA, wherein
SEQ ID N0:300 is a clone designated herein as "UNQ528" andlor "DNA58847-1383".
Figure 120 shows the amino acid sequence (SEQ ID N0:301) derived from the
coding sequence of SEQ ID
N0:300 shown in Figure 119.
Figure 121 shows a nucleotide sequence {SEQ ID N0:302) of a native sequence
PR01072 cDNA, wherein
SEQ ID N0:302 is a clone designated herein as "UNQ529" and/or "DNA58747-1384".
Figure 122 shows the amino acid sequence (SEQ ID N0:303) derived from the
coding sequence of SEQ ID
N0:302 shown in Figure 121.
Figure 123 shows an EST nucleotide sequence designated herein as DNA40210 (SEQ
ID N0:304).
Figure 124 shows a nucleotide sequence (SEQ ID N0:308) of a native sequence
PR01075 cDNA, wherein
SEQ ID N0:308 is a clone designated herein as "UNQ532" andlor "DNA57b89-1385".
Figure 125 shows the amino acid sequence (SEQ ID N0:309) derived from the
coding sequence of SEQ ID
N0:308 shown in Figure I24.
Figure 126 shows an EST nucleotide sequence designated herein as DNAI3059 (SEQ
ID N0:3I0).
Figure 127 shows an EST nucleotide sequence designated herein as DNA194b3 (SEQ
ID N0:311).
Figure I28 shows a nucleotide sequence (SEQ ID N0:321) of a native sequence
PR0181 cDNA, wherein
SEQ iD N0:321 is a clone designated herein as "UNQ155" andlor "DNA23330-1390".
Figure I29 shows the amino acid sequence (SEQ ID N0:322) derived from the
coding sequence of SEQ ID
N0:321 shown in Figure 128.
Figure 130 shows an EST nucleotide sequence designated herein as DNA13242 (SEQ
ID N0:323).
Figure 131 shows a nucleotide sequence (SEQ ID N0:329) of a native sequence
PR0195 cDNA, wherein
SEQ ID N0:329 is a clone designated herein as "UNQ169" and/or "DNA'26847-
I395".
Figure 132 shows the amino acid sequence (SEQ ID N0:330) derived from the
coding sequence of SEQ ID
N0:329 shown in Figure 131.
Figure I33 shows an EST nucleotide sequence designated herein as DNA15062 (SEQ
ID N0:331).
Figure 134 shows an EST nucleotide sequence designated herein as DNA13199 {SEQ
ID N0:332).
Figu~te 135 shows a nucleotide sequence (SEQ ID N0:336) of a native sequence
PR0865 cDNA, wherein
SEQ ID N0:336 is a clone designated herein as "UNQ434" andlor "DNA53974-1401".
Figure 136 shows the amino acid sequence (SEQ ID N0:33?) derived from the
coding sequence of SEQ ID
N0:336 shown in Figure 135.
Figure 137 shows an EST nucleotide sequence designated herein as DNA37642 (SEQ
ID N0:338).
Figure 138 shows a nucleotide sequence (SEQ 1D N0:345) of a native sequence
PR0827 cDNA, wherein
SEQ ID N0:345 is a clone designated herein as "UNQ468" andlor "DNA57039-1402".
Figure I39 shows the amitm acid sequence (SEQ 1D N0:346) derived from the
coding sequence of SEQ ID
N0:345 shown in Figure 138. .
Figure 140 shows an EST nucleotide sequence designated herein as DNA47751 (SEQ
ID N0:347).
Figure I41 shows a nucleotide sequence (SEQ ID N0:351) of a native sequence
PR01114 cDNA, wherein
SEQ ID N0:351 is a clone designated herein as "UNQ557" andlor "DNA57033-1403".
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Figure 142 shows the amino acid sequence (SEQ ID N0:352) derived from the
coding sequence of SEQ ID
N0:351 shown in Figure 141.
Figure 143 shows an EST nucleotide sequence designated herein as DNA48466 (SEQ
1D N0:353).
Figure 144 shows a nucleotide sequence {SEQ ID N0:357) of a native sequence
PR0237 cDNA, wherein
SEQ ID N0:357 is a clone designated herein as "UNt2211" and/or "DNA34353-
1428".
Figure 145 shows the amino acid sequence {SEQ 1D N0:358) derived from the
coding sequence of SEQ ID
N0:357 shown in Figure 144.
Figure 146 shows a nucleotide sequence (SEQ ID N0:362) of a native sequence
PR0541 cDNA, wherein
SEQ ID N0:362 is a clone designated herein as "UNQ342" and/or "DNA45417-1432".
Figure 147 shows the amino acid sequence (SEQ ID N0:363) derived from the
coding sequence of SEQ ID
N0:362 shown in Figure 146.
Figure 148 shows a nucleotide sequence (SEQ ID N0:369) of a native sequence
PR0273 cDNA> wherein
SEQ ID N0:369 is a clone designated herein as "UNQ240" andlor "DNA39523-1192".
Figure 149 shows the amino acid sequence (SEQ ID N0:370) derived from the
coding sequence of SEQ ID
N0:369 shown in Figure 148.
Figure 150 shows a nucleotide sequence (SEQ ID N0:374) of a native sequence
PR0701 cDNA, wherein
SEQ ID N0:374 is a clone designated herein as "UNQ365" andlor "DNA44205-1285",
Figure 151 shows the amino acid sequence (SEQ ID N0:375) derived from the
coding sequence of SEQ ID
N0:374 shown in Figure 150.
Figure 152 shows a nucleotide sequence (SEQ.ID N0:379) of a native sequence
PR0704 cDNA, wherein
SEQ ID N0:379 is a clone designated herein as "UNQ368" atxl/or "DNA50911-
1288".
Figure 153 shows the amino acid sequence (SEQ ID N0:380) derived from the
coding sequence of SEQ ID
N0:379 shown in Figure 152.
Figure 154 shows a nucleotide sequence (SEQ ID N0:384) of a native sequence
PRO706 cDNA, wherein
SEQ ID N0:384 is a clone designated herein as "UNQ370" andlor "DNA48329-1290".
Figure 155 shows the amino acid sequence (SEQ ID N0:385) derived from the
coding sequence of SEQ ID
N0:384 shown in Figure 154.
Figure 156 shows a nucleotide sequence (SEQ ID N0:389) of a native sequence
PR0707 cDNA, wherein
SEQ ID N0:389 is a clone designated herein as "UNQ371" and/or "DNA48306-
1291".'
Figure 157 shows the atnino acid sequence (SEQ ID N0:390) derived from the
coding sequence of SEQ ID
N0:389 shown in Figure 156.
Figure 158 shows a nucleotide seduence (SEQ ID N0:394) of a native sequence
PR0322 cDNA, wherein
SEQ ID N0:394 is a clone designated herein as "UNQ283" and/or °DNA48336-
1309" .
Figure 159 shows the amino acid sequence (5EQ ID N0:395) derived from the
coding sequence of SEQ ID
N0:394 shown in Figure 158.
Figure 160 shows a nucleotide sequence (SEQ ID N0:399) of a native sequence
PR0526 cDNA, wherein
SEQ ID N0:399 is a clone designated herein as "UNQ330" and/or "DNA44184-1319".
Figure 161 shows dte amino acid sequence (SEQ ID N0:4p0) derived from the
coding sequence of SEQ ID
N0:399 shown in Figure 160.
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Figure 162 shows a nucleotide sequence (SEQ ID N0:404) of a nati~oe sequence
PR0531 cDNA, wherein
SEQ ID N0:404 is a clone designated herein as "UNQ332" and/or "DNA48314-1320"_
Figure i63 shows the amino acid sequence (SEQ ID N0:405) derived from the
coding sequence of SEQ ID
N0:404 shown in Figure 162.
Figure 164 shows a nucleotide sequence (SEQ ID N0:409) of a native sequence
PR0534 cDNA, wherein
S SEQ ID N0:409 is a clone designated herein as "UNQ33S" andlor "DNA48333-
1321".
Figure i65 shows the amino acid sequence (SEQ 1D N0:410) derived from the
coding sequence of SEQ ID
N0:409 shown in Figure 164.
Figure 166 shows a nucleotide sequence (SEQ ID N0:4I4) of a native sequence
PR0697 cDNA, wherein
SEQ 1D N0:414 is a clone designated herein as "UNQ36i" and/or "DNA50920-1325".
Figure 167 shows the amino acid sequence (SEQ ID N0:415) derived from the
coding sequence of SEQ ID
N0:414 shown in Figure 166: ,,
Figure I68 shows a nucleotide sequence (SEQ ID N0:419) of a native sequence
PR07I7 cDNA, wherein
SEQ ID N0:419 is a clone designated herein as "UNQ385" andJor "DNA50988-1326".
Figure i69 shows the amino acid sequence (SEQ ID N0:420) derived from the
coding sequence of SEQ iD
i5 N0:419 shown in Figure 168:
Figures 170A-B show a nucleotide sequence (SEQ ID N0:424) of a native sequence
PR0731 cDNA,
wherein SEQ ID N0:424 is a cio~ designated herein as "UNQ39S" and/or "DNA48331-
1329".
Figure 171 shows the amino acid sequence (SEQ ID N0:425) derived from the
coding sequence of SEQ ID
N0:424 shown in Figures I70A-B.
Figure 172 shows a nucleotide sequence (SEQ ID N0:429} of a naive sequence
PR0218 cDNA, wherein
SEQ ID N0:429 is a clone designated herein as "UNQ192" andlor 'DNA30867-1335".
Figure i73 shows the amino acid sequence (SEQ ID N0:430) derived from the
coding sequence of SEQ ID
N0:429 shown in Figure 172.
Figure 174 shows an EST nucleotide sequence designated herein as DNA14472 (SEQ
ID N0:431).
Figure 175 shows an EST nucleotide sequence designated herein as DNA15846 (SEQ
iD N0:432).
Figures 176A-B show a nucleotide sequence (SEQ ID N0:436) of a native sequence
PR0768 cDNA,
wherein SEQ ID NQ:436 is a clone designated herein as "UNQ446" and/or
'DNA55737-1345".
Figure i77 shows the amino acid sequence (SEQ iD'N0:437) derived from the
coding sequence of SEQ ID
N0:436 shown in Figures 176A-B.
Figure 178 shows a nucleotide, sequence (SEQ ID N0:441) of a native sequence
PR0771 cDNA, wherein
SEQ ID N0:441 is a clone designated herein as "UNQ409" andlor "DNA49829-1346".
Figure 179 shows the amino acid sequence (SEQ ID N0:442) derived froth the
coding sequence of SEQ ID
N0:441 shown in Figure 178.
Figures 180A-B show a nucleotide sequence (SEQ ID N0:446) of a native sequence
PR0733 cDNA,
wherein SEQ ID N0:446 is a clone designated herein as 'UNQ42I" andlor
"DNA52196-1348".
Figure 181 shows the amino acid sequetxe (SEQ ID N0:447) derived from the
coding sequence of SEQ ID
N0:446 shown in Figures I80A-B.
Figure 182 shows a nucleotide sequence (SEQ ID N0:451) of a native seauetice
PR0162 eDNA, wherein
SEQ iD N0:451 is a clone designated herein as "UNQ429" and/or "DNA56965-1356".
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Figure 183 shows the amino acid sequence (SEQ ID N0:452) derived from the
coding sequence of SEQ 1D
N0:451 shown in Figure 182.
Figure 184 shows a.nucleotide sequence (SEQ ID N0:453) of a native sequence
PR0788 cDNA, wherein
SEQ ID N0:453 is a clone designated herein as "UNQ430" andlor "DNA56405-1357".
Figure 185 shows the amino acid sequence (SEQ ID N0:454) derived from the
coding sequence of SEQ ID
N0:453 shown in Figure 184.
Figure 186 shows a nucleotide sequence (SEQ ID N0:455) of a native sequence
PR01008 cDNA, wherein
SEQ ID N0:455 is a clone designated herein as "UNQ492" andlor "DNA57530-1375".
Figure 187 shows the amino acid sequence (SEQ ID N0:456) derived from the
coding sequence of SEQ ID
N0:455 shown in Figure 186.
Figure 188 shows an EST nucleotide sequence designated herein as DNA16508 (SEQ
ID N0:457).
Figures 189A-B show a nucleotide sequence (SEQ ID NO:458) of a native sequence
PR01012 cDNA,
wherein SEQ ID NO:458 is a clone designated herein as "UNQ495" andlor
"DNA56439-137b".
Figure 190 shows the amino acid sequencx (SEQ ID N0:459) derived from the
coding sequence of SEQ ID
N0:458 shown in Figures 189A-B.
Figure 191 shows a nucleotide sequence {SEQ ID N0:463) of a native sequence
PR01014 cDNA, wherein
SEQ ID N0:463 is a clone designated herein as "UNQ497" andlor "DNA56409-1377".
Figure 192 shows the amino acid sequence (SEQ ID N0:464) derived from the
coding sequence of SEQ ID
N0:4b3 shown in Figure 191.
Figure 193 shows a nucleotide sequence {SEQ ID N0:465) of a native sequence
PRO101? cDNA, wherein
SEQ ID N0:465 is a clone designated herein as "UNQ500" andlor "DNA56112-1379"
.
Figure 194 shows the amino acid sequence (SEQ ID N0:466) derived from the
coding sequence of SEQ 113
N0:465 shown in Figure 193.
Figure 195 shows a nucleotide sequence (SEQ ID N0:467) of a native sequence
PR0474 cDNA, wherein
SEQ ID NO:467 is a clone designated herein as "UNQ502" andlor "DNA56045-1380".
Ftgure 196 shows the amino acid sequence (SEQ ID N0:4b8) derived from the
coding sequence of SEQ ID
N0:467 shown in Figure 195.
Figure 197 shows a nucleotide sequence (SEQ ID N0:469) of a native sequence
PRO1031 cDNA, wherein
SEQ ID N0:469 is a clone designated herein as "UNQ516" and/or "DNA59294-1381".
Figure 198 shows the amino acid sequence (SEQ ID N0:4?0) derived from the
coding sequence of SEQ ID
N0:469 shown in Figure 197.
Figure 199 shows a nucleotide sequence (SEQ ID N0:471) of a native sequence
PR0938 cDNA, wherein
SEQ ID N0:471 is a clone designated herein as "UNQ475" and/or "DNA56433-1406".
Figure 200 shows the amino acid sequence (SEQ ID N0:472) derived from the
coding sequence of SEQ ID
N0:471 shown in Figure 199.
Figure 201 shows a nucleotide sequence (SEQ ID N0:476) of a native sequence
PR01082 cDNA, wherein
SEQ ID N0:476 is a clone designated herein as "UNQ539" andlor "DNA53912-1457".
Figure 202 shows the amino acid sequence (SEQ ID N0:477) derived from the
coding sequence of SEQ ID
N0:476 shown in Figure 201.
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Figure 203 shows a nucleotide sequence (SEQ ID N0:482) of a native: sequence
PR01083 cDNA, wherein
SEQ ID N0:482 is a clone designated herein as "UNQ540" andlor "DNA50921-1458".
Figure 204 shows the amino acid sequence (SEQ ID N0:483) derived from the
coding sequence of SEQ ID
N0:482 shown in Figure 203.
Figure 205 shows an EST nucleotide sequence designated herein as DNA24256 (SEQ
ID N0:484).
Figure 206 shows a nucleotide sequence (SEQ ID N0:487} of a native sequence
PR0200 cDNA, wherein
SEQ ID N0:487 is a clone designated herein as "UNQ1?4" and/or "DNA29101-1122".
Figure 207 shows the amino acid sequence (SEQ ID N0:488) derived from the
coding sequence of SEQ ID
N0487 shown in Figure 206.
Figure 208 shows a nucleotide sequence {SEQ 1D N0:495) of a native sequence
PR0285 cDNA, wherein
SEQ ID N0:495 is a clone designated herein as "DNA44021-1154".
Figure 209 shows the amino acid sequence (SEQ ID N0:496) derived from the
coding sequence of SEQ ID
NO:495 shown in Figure 208.
Figures 210A-B show a nucleotide sequence (SEQ ID N0:497) of a native sequence
PR0286 cDNA;
wherein SEQ ID N0:49? is a clone designated herein as "DNA42663-1154".
Figure 211 shows the amitw acid sequence (SEQ ID N0:498) derived from the
coding, sequence of SEQ ID
N0:497 shown in Figures 210A-8.
Figure 212 shows a nucleotide sequence {SEQ ID N0:505) of a native sequence
PR0213-1 cDNA, wherein
SEQ ID N0:505 is a clone: designated herein as °DNA30943-I-l I63-
1".
Figure 213 shows tire amino acid sequence (SEQ ID N0:506} derived from the
coding sequence of SEQ ID
NO:505 shown in Figure 212.
Figure 2I4 shows a nucleotide sequence (SEQ ID N0:507) of a native sequence
PR01330 cDNA, wherein
SEQ ID N0:507 is a clone designated herein as "DNA64907-I I63-1".
Figure 215 shows the amine acid sequence (SEQ ID N0:508) derived from the
coding sequence of SEQ ID
N0:507 shown in Figure 214.
Figure 216 shows a rexleotide sequence (SEQ ID N0:509) of a native sequence
PROI449 cDNA, wherein
SEQ ID N0:509 is a clone designated herein as "DNA64908-1163-1".
Figure 217 shows the amino acid sequence (SEQ ID N0:510) derived from the
coding sequence of SEQ ID
N0:509 shown in Figure 216.
Figure 218 shows a nucleotide sequence (SEQ ID N0:514) of a native sequence
PR0298 cDNA, wherein
SEQ ID N0:514 is a clone designated herein as "UNQ261" and/or "DNA39975-1210".
Figure 219 shows the amino acid sequence (SEQ ID N0:515) derived from the
coding sequence of SEQ ID
N0:5I4 shown in Figure 218.
Figure 220 shows an EST nucleotide sequence designated herein as DNA26832 (SEQ
ID N0:516).
Figure 221 shows a nucleotide sequence (SEQ ID N0:522) of a native sequence
PR0337 cDNA, wherein
SEQ ID N0:522 is a clone designated herein as "DNA43316-1237".
Figure 222 shows the amino acid sequence (SEQ ID N0:523) derived from the
coding sequence of SEQ ID
N0:522 shown in Figure 221.
Figure 223 shows an EST nucleotide sequence designated hereon as DNA42301 (SEQ
ID N0:524).
I09
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Figure 224 shows a nucleotide sequence (SEQ ID N0:525) of a native sequence
PR0403 cDNA, wherein
SEQ ID N0:525 is a clone designated herein as "DNA55800-1263".
Figure 225 shows the amino acid sequence (SEQ iD N0:526) derived from the
coding sequence of SEQ ID
N0:525 shown in Figure 224.
Figures 226A-B show an EST nucleotide sequence designated herein as DNA344I5
(SEQ ID N0:527).
Figure 22? shows an EST nucleotide sequence designated herein as DNA49830 (SEQ
ID N0:528).
Figure 228 shows an EST nucleotide sequence designated herein as DNA49831 (SEQ
iD N0:529).
DETAILED DESCRIPTION OF THE PREFERRED EMBODIMENTS
I. Deflnitiotts
The terms "PRO polypeptide" and "PRO" as used herein and when immediately
followed by a numerical
designation refer to various polypeptides, wherein the complete designation
(i.e., PRO/number) refers to specific
polypeptide sequences as described herein. The terms "PROlnumber polypeptide"
and "PROlnumber" as used herein
encompass native sequence poiypeptides and polypeptide variants (which are
further defined herein). The PRO
poIypeptides described herein may be isolated from a variety of sources, such
as from human tissue types or from
another source, or prepared by recombinant or synthetic methods.
A "native sequence PRO polypeptide" comprises a polypeptide having the same
amino acid sequence as the
corresponding PRO polypeptide derived from nature. Such native sequence PRO
polypeptides can be isolated from
nature or can be produced by recombinant or synthetic means. The term "native
sequence PRO polypeptide"
specifically encompasses naturally-occurring truncated or secreted forms of
the specific PRO polypeptide (e.g., an
extracellular domain sequence), naturally-occurring variant forms (e.g.,
alternatively spliced forms) and naturally-
occurring allelic variants of the polypeptide. In various embodiments of the
invention, the native sequence PR0213
poiypeptide is a mature or full length native sequence PR0213 polypeptide
comprising amino acids 1 to 295 of Figure
2 (SEQ ID N0:2), the native sequence PR0274 polypeptide is a mature or full-
length native sequence PR0274
polypeptide comprising amino acids 1 to 492 of Figure 4 (SEQ 1D N0:7), the
native sequence PR0300 polypeptide
is a mature or full-length native sequence PR0300 polypeptide comprising amino
acids 1 to 457 of Figure 9 (SEQ
ID N0:19), the native sequence PR0284 polypeptide is a mature or full-length
native sequence PR0284 polypeptide
comprising amino acids 1 to 285 of Figure 11 (SEQ 1D N0:28}, the native
sequence PR0296 polypeptide is a mature
or full-length native sequence PR0296 polypeptide comprising amino acids 1 to
204 of Figure IS (SEQ ID N0:36),
the native sequence PR0329 polypeptide is a mature or full-length native
sequence PR0329 polypeptide comprising
amino acids 1 to 359 of Figure 2D (SEQ ID N0:45), the native sequence PR0362
polypeptide is a mature or full-
length native sequence PR0362 polypeptide comprising amino acids 1 to 321 of
Figure 22 (SEQ ID N0:52), the
native sequence PR0363 polypeptide is a mature or full-length native sequence
PR0363 polypeptide comprising
amino acids 1 to 373 of Figure 24 (SEQ ID N0:59), the native sequence PR0868
polypeptide is a mature or full-
length native sequence PR0868 polypeptide comprising amino acids 1 to 655 of
Figure 26 (SEQ ID N0:64), the
native sequence PR0382 polypeptide is a mature or full-length native sequence
PR0382 polypeptide comprising
amino acids 1 to 453 of Figure 28 (SEQ ID N0:69), the native sequence PR0545
polypeptide is a mature or full-
iength native sequence PR0545 polypeptide comprising amino acids 1 to 735 of
Figure 30 (SEQ ID N0:74), the
native sequence PR0617 polypeptide is a mature or foil-length native sequence
PR0617 polypeptide comprising
amino acids 1 to 6? of Figure 33 (SEQ ID NO:85), the native sequence PR0700
polypeptide is a mature or full-length
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native sequence PR0700 polypeptide comprising amino acids I to 432 of Figure
35 (SEQ ID N0:90), the native
sequence PR0702 polypeptide is a mature or full-length native sequence PR0702
polypeptide comprising amino acids
1 to 277 of Figure 37 (SEQ ID N0:97), the native sequence PR0703 polypeptide
is a mature or full-length native
sequence PR0703 poiypeptide comprising amino acids I to 730 of Figure 39 (SEQ
ID N0:102), the native sequence
PR0705 polypeptide is a mature or full-length native sequence PR0705
polypeptide comprising amino acids 1 to 555
of Figure 41 (SEQ ID NO:i09), the native sequence PR0708 polypeptide is a
mature or full-length native sequence
PR07U8 polypeptide comprising amino acids 1 to 515 of Figure 43 (SEQ ID
N0:114), the native sequence PR0320
polypeptide is a mature or full-lengdt native sequence PR0320 polypeptide
comprising amino acids 1 to 338 of Figure
45 (SEQ ID N0:119), the native sequence PRU324 polypeptide is a mature or full-
length native sequence PR0324
polypeptide comprising amino acids 1 to 289 of Figure 47 (SEQ 1D N0:124), the
native sequence PR0351
polypeptide is a mature or full-length native sequence PR0351 polypeptide
comprising amino acids I to 571 of Figure
49 (SEQ m N0:132), the native sequence PR0352 polypeptide is a mature or full-
length native sequence PR0352
polypeptide comprising amino acids l to 316 of Figure 51 (SEQ ID N0:137), the
native sequence PR0381
polypeptide is a mature or full-length native sequet~e PR0381 polypeptide
comprising amino acids l to 21 I of Figure
S3 (SEQ ?3) N0:145), the native sequence PR0386 polypeptide is a mature or
full-length native sequence PR0386
1S polypeptide comprising amino acids 1 to 215 of Figure 55 (SEQ ID NO:150),
the native sequence PR0540
polypeptide is a mature or full-length native sequence PR0540 polypeptide
comprising amino acids 1 to 412 of Figure
59 (SEQ ID N0:157), the native sequence PR0615 polypeptide is a mature or full-
length native sequence PR06i5
polypeptide comprising amino acids l to 224 of Figure 61 (SEQ ID N0:162), the
native sequence PR0618
polypeptide is a mature or full-length native sequence PR06I8 polypeptide
comprising amino acids I to 802 of Figure
63 (SEQ ID N0:169), the native sequence PR0719 polypeptide is a mature or full-
length native sequence PR0719
polypeptide comprising atnnno acids 1 to 354 of Figure 66 (SEQ ID NOa78), the
native sequence PR0724
polypeptide is a mature or full-length native sequence PR0724 polypeptide
comprising amino acids 1 to 713 of Figure
68 (5EQ ID NO:I83), the native sequence PR0772 poiypeptide is a mature or full-
length native sequence PR0772
polypeptide comprising amino acids 1 to 152 of Figure 70 (SEQ ID N0:190), the
native sequence PR0852
polypeptide is a mature or foil-length native sequence PR0852 polypeptide
comprising amino acids 1 to 518 of Figure
73 (SEQ ID N0:196), the native sequence PR0853 polypeptide is a mature or full-
length native sequence PR0853
polypeptide comprising amino acids 1 to 3?7 of Figure 75 (SEQ Il) N0:206), the
native sequence PR0860
polypeptide is a mature or full-length native sequence PR0860 polypeptide
comprising amino acids 1 to 985 of Figure
77 (SEQ ID N0:211), the native sequence PR0846 polypeptide is a mature or full-
length native sequence PR0846
polypeptide comprising amino acids l ~ to 332 of Figure 79 (SEQ ID N0:216),
the native sequence PR0862
polypepude is a mature or full-length native sequet~ PR0862 polypeptide
comprising amino acids 1 to 146 of Figure
81 (SEQ ID N0:221), the native sequence PR0864 polypeptide is a mature or full-
length native sequence PR0864
polypeptide comprising amino acids 1 to 331 of Figure 83 (SEQ ID N0:226), the
native sequence PR0792
polypeptide is a mature or full-length native sequence PR0792 polypeptide
comprising amino acids 1 to 293 of Figure
85 (SEQ 1D N0:231), the native sequence PRO866 polypeptide is a mature or full-
length native sequence PR0866
polypepude comprising amino acids 1 to 331 of Figure 87 (SEQ ID N0:236), the
native sequence PR0871
polypeptide is a mature or full-length native sequence PR087I polypeptide
comprising amino acids 1 to 472 of Figure
89 (SEQ ID N0:245), the native sequence PR0873 polypeptide is a mature or full-
length native sequence PR0873
polypeptide comprising amino acids l to 545 of Figure 91 (SEQ ID N0:254), the
native sequence PR0940
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polypeptide is a mature or full-length native sequence PR0940 polypeptide
comprising amino acids 1 to 544 of Figure
93 (SEQ ID N0:259), the native sequence PR0941 polypeptide is a mature or full-
length native sequence PR0941
polypeptide comprising amino acids 1 to 772 of Figure 95 (SEQ ID N0:264), the
native sequence PR0944
polypeptide is a mature or full-length native sequence PR0944 polypeptide
comprising amino acids 1 to 211of Figure
98 (SEQ ID N0:270), the native sequence PR0983 polypeptide is a mature or full-
length native sequence PR0983
polypeptide comprising amino acids 1 to 243 of Figure 109 (SEQ ID N0:284), the
native sequence PROI057
polypeptide is a mature or full-length native sequence PR01057 polypeptide
comprising amino acids 1 to 413 of
Figure 118 (SEQ ID N0:296), the native sequence PROI071 polypeptide is a
mature or full-length native sequence
PR01071 polypeptide comprising amino acids 1 to 525 of Figure 120 (SEQ ID
NO:301), the native sequence
PR01072 polypeptide is a mature or full-length native sequence PR01072
polypeptide comprising amino acids 1 to
336 of Figure 122 (SEQ ID N0:303), the native sequence PR01075 polypeptide is
a mature or full-length native
sequence PR01075 polypeptide comprising amino acids 1 to 406 of Figure 125
(SEQ ID N0:309), the native
sequence PRO181 polypeptide is a mature or full-length native sequence PR0181
polypeptide comprising amino acids
1 to 144 of Figure 129 (SEQ ID N0:322), the native sequence PR0195 polypeptide
is a mature or full-length native
sequence PR0195 polypeptide comprising amino acids 1 to 323 of Figure I32 (SEQ
ID N0:330), the native sequence
PR08b5 polypeptide is a mature or full-length native sequence PR0865
polypeptide comprising amino acids 1 to 468
of Figure 136 (SEQ 1D N0:337), the native sequence PR082? poIypeptide is a
mature or full-length native sequence
PR0827 polypeptide comprising amino acids 1 to 124 of Figure 39 (SEQ ID
N0:34b), the native sequence PR01114
polypeptide is a mature or full-length native sequence FRO1114 polypeptide
comprising amino acids l to 311 of
Figure 142 (SEQ ID N0:352), the native sequence PR0237 polypeptide is a matuze
or full-length native sequence
PR0237 polypeptide comprising amino acids 1 to 328 of Figure 145 (SEQ ID
N0:358).> the native sequence PR0541
polypeptide is a mature or full-length native sequence PR0541 polypepride'
comprising amino acids 1 to 500 of Figure
14? (SEQ ID N0:363), the native sequence PR0273 polypeptide is a mature oz
full-length native sequence PR0273
polypeptide comprising amino acids 1 through l 11 of Figure 149 (SEQ ID
NO:370), the native sequence PRO?01
polypeptide is a mature or full-length native sequence PR0701 polypeptide
comprising amino acids 1 to 816 of Figure
151 (SEQ 1D N0:375), the native sequence PRO704 polypeptide is a full-length
or mature native sequence PRO704
polypeptide comprising amino acids 1 or 40 through 348 of Figure 153 (SEQ ID
N0:380), the native sequence
PR0706 polypeptide is a mature or full-length native sequence PR0706
polypeptide comprising amino acids 1 to 480
of Figure 155 (SEQ ID N0385), the native sequence PR0707 polypeptide is a full-
length or mature native sequence
PR0707 polypeptide comprising amino acids 1 or 31 through 916 of Figure 157
(SEQ ID N0:390), the native
sequence PR0322 polypeptide is a mature or full-Length native sequence PR0322
polypeptide comprising amino acids
24 or 1 to 260 of Figure 159 (SEQ ID N0:395), the native sequence PR0526
polypeptide is a full-length or mature
native sequence PR052b polypeptide comprising amino acids 1 or 27 through 4'73
of Figure 161 (SEQ ID N0:400),
the native sequence PR0531 polypeptide is a mature PR0531 polypeptide
comprising amino acids l to 789 of Figure
1b3 (SEQ ID N0:405), the native sequence PR0534 polypeptide is a mature or
full-length native sequence PR0534
polypeptide comprising amino acids 1 to 360 of Figure i65 (SEQ ID N0:410), the
native sequence PROb97
polypeptide is a full-length or mature native sequence PR0697 polypeptide
comprising amino acids 1 or 21 through
295 of Figure 1b? (SEQ ID N0:415), the native sequence FR0717 polypeptide is a
mature or full-length native
sequence PR07I7 polypeptide comprising amino acids l through 560 of Figure 169
(SEQ ID N0:420), the native
sequence PRO731 polypeptide is a full-length or mature native sequence PR0731
polypeptide comprising amino acids
1I2
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1 or I4 through 1184 of Figure 171 (SEQ ID N0:425), the native sequence PR0218
polypeptide is a full-length or
mature native sequence PR0218 polypeptide comprising amino acids 1 or 24
through 455 of Figure 173 (SEQ ID
N0:430), the native sequence PR0768 polypeptide is a full-length or mature
native sequence PR0768 polypeptide
comprising amino acids 1 or 34 through 1141 of Figure 177 (SEQ ID N0:437), the
native sequence PR0771
polypeptide is a full-length or mature native sequence PRO?7i polypeptide
comprising amino acids 1 or 17 through
436 of Figure 179 (SEQ iD N0:442), the native sequence PR0733 polypeptide is a
mature or full-length native
seguence PRO733 polypeptide comprising amino acids 24 or 1 through 229 of
Figure 181 (SEQ ID N0:447), the
native sequence PR0162 polypeptide is a full-length or mature native sequence
PR0162 polypeptide comprising
amino acids 1 or 27 through 175 of Figure 183 (SEQ ID N0:452), the native
sequence PR0788 polypeptide is a full-
length or mature native sequence PR0788 polypeptide comprising amino acids 1
or 18 through I25 of Figure I85
(SEQ ID N0:454), the native sequence PR01008 polypegtide is a full-length or
mature native sequence PR01008
polypeptide comprising amino acids 1 or 24 through 266 of Figure 187 (SEQ ID
N0:456), the native sequence
PR01012 polypeptide is a mature or full-length native sequence PR01012
polypeptide comprising amino acids I
through 747 of Figure 190 (SEQ ID N0:459), the native sequence PR01014
polypeptide is a full-length or mature
native sequence PR01014 polypeptide rn~nprising amino acids 1 or 20 through
300 of Figure 192 (SEQ ID N0:464),
the native sequence PR01017 polypeptide is a full-length or mature native
sequence PR01017 polypeptide comprising
amino acids 1 or 32 through 414 of Figure 194 (SEQ ID N0:466), the native
sequence PR0474 poiypeptide is a
mature or full-length native sequence PRQ474 polypeptide c:otr>prising amino
acids 1 through 270 of Figure 196 (SEQ
ID N0:468), the native sequence PR01031 polypeptide is a full-length or mature
native sequence PR01031
polypeptide comprising amino acids 1 or 21 through 180 of Figure 198 (SEQ ID
N0:470), the native sequence
PRO938 poIypeptide is a mature or full-length native sequence PR0938
polypeptide comprising amino acids 1 to 349
of Figure 200 (SEQ ID N0:472); the native sequence PR01082 polypeptide is a
full-length or mature native sequence
PR01082 poIypeptide comprising amino acids 1 through 201 of Figure 202 (SEQ ID
N0:47?), the native sequence
PR01083 polypeptide is a full-length or mature native sequence PR01083
polypeptide comprising amino acids 1 or
26 through 693 of Figure 204 (SEQ ID N0:483), the native sequence VEGF-E
poIypeptide is a mature or full-length
native sequence VEGF-E polypeptide comprising amino acids 1 through 345 as
depicted in Figure 207 (SEQ ID
N0:488), the native sequence PR0285 is a mature or full-length native sequence
PR0285 polypeptide comprising
amino acids 1 to 1049 of Figure 209 (SEQ m N0:496), the native sequence PR0286
is a mature or full-length native
sequence PR0286 polypeptide comprising amino acids I to 1041 of 1 figure 211
(SEQ ID N0:298), the native
se,quen~ PR0298 is a mature or full length native sequence PR0298 comprising
amino acids 1 to 364 of Figure 219
(SEQ ID N0:515), the native sequeixe PR0337 is a mature or full-length native
sequence human neurotrimin
comprising amino acids 1 to 344 of Figure 222 (SEQ ID N0:523), with or without
the N-terminal signal sequence
(residues 1 to about 28), and with or without the initiating methionine at
position 1 and the native sequence PR0403
is a mature or full-length native sequence comprising amino acids 1 to 'l36 of
Figure 225 (SEQ ID NO:S26), with
or without the initiating methionine at position 1.
The PRO polypeptide "extracellular domain" or "ECD" refers to a form of the
PRO polypeptide which is
essentially free of the transmembrane and cytoplasmic domains. Ordinarily, a
PRO polypeptids ECD will have less
than 1 % of such transmembrane andlor cytoplasmic domains and preferably; will
have less than 0.596 of such
domains. 1t will be tmderstood that any transmembrane domains identified for
the PRO polypeptides of the present
invention are identified pursuant to criteria routinely employed in the art
for identifying that type of hydrophobic
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WO 99146281 PCTlUS99105028
domain. The exact boundaries of a transnaembrane domain may vary but most
likely by no more than about 5 amino
acids at either end of the domain as initially identified. Optionally,
therefore, an extracellular domain of a PRO
polypeptide may contain from about 5 or fewer amino acids on either or the
transmembrane domain as initially
identified.
"PRO polypeptide variant" means an active PRO polypeptide as defined above or
below having at least about
80% amino acid sequence identity with the full-length native sequence PRO
polypegtide sequence as disclosed herein.
Such PRO polypeptide variants include, for instance, PRO polypeptides wherein
one or more amino acid residues
are added, or deleted, at the N- or C-terminus of the full-length native amino
acid sequence. Ordinarily, a PRO
polypeptide variant will have at least about 80% amino acid sequence identity,
more preferably at least about 85%
amino acid sequence identity, and even more preferably at least about 90%
amino acid sequence identity, even more
preferably at least about 91 % amino acid sequence identity, even more
preferably at least about 92 % amino acid
sequence identity, even more preferably at least about 93 % amino acid
sequence identity, even more preferably at
least about 94% amino acid sequence identity, even more preferably at least
about 95 % amino acid sequence identity;
yet more preferably at least about 96 % amino acid sequence identity, yet more
preferably at least about 97 % amino
acid sequence identity, yet more preferably at least about 98% amino acid
sequence identity and most preferably at
least about 99% amino acid sequence identity with rite amino acid sequence of
the full-length native amino acid
sequence as disclosed herein.
"Percent (%) amino acid sequence identity" with respect to the PRO polypeptide
sequences identified herein
is defined as the percentage of amino acid residues in a candidate sequence
that are identical with the amino acid
residues in the specifrc PRO polypeptide sequence, after aligning the
sequences and introducing gaps, if necessary,
to achieve the maximum percent sequence identity, and not considering any
conservative substitutions as part of the
sequence identity. Alignment for purposes of determining percent amino acid
sequence identity can be achieved in
various ways that are within the skill in the art, for instance, using
publicly available computer software such as
BLAST, BLAST-2, ALIGN or Megalign (DNASTAR) software. The preferred software
alignment program is
BLAST. Those skilled in the art can determine: appropriate parameters for
measuring alignment, including any
algorithms needed to achieve maximal alignment over the full length of the
sequences being compared. The %
identity values used herein have been generated using the WU-BL.AST 2 computer
program (Altschul et al., Methods
jsb .~nz5mnolo~ ~:4b0-480 (1996); http:f/blast.wustl/edult>last/README.htm1).
Most of the WU-BLAST-2 search
parameters were set to the default values. The adjustable parameters were set
with the following values: overlap span
= 1, overlap fraction = 0.125, word threshold (T~ = 11, and scoring matrix =
BLOSUM62. The HSP S and HSP
S2 parameters, which are dynamic values used by BLAST-2, are established by
the program itself depending upon
the composition of the sequence of interest and composition of the database
against which the sequence is being
searched. However, the values may be adjusted to increase sensitivity. A %
sequence identity value is determined
by the fraction of matching identical residues divided by the total number of
residues in the aligned region.
"Percent (%) rntcleic acid sequence identity" with respect to PRO-encoding
nucleic acid sequences identified
herein is defined as the percentage of nucleotides in a candidate sequence
that are identical with the nucleotides in
the PRO nucleic acid sequence of interest, after aligning the sequences and
introducing gags, if necessary, to achieve
the maximum percent sequence identity. Alignment for purposes of determining
percent nucleic acid sequence
identity can be achieved in various ways that are within the skill in the art,
for instance, using publicly available
caa>puter software such as BLAST, BLAST-2, ALIGN or Megalign (DNASTAR)
software. Those skilled in the art
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WO 99146281 PCT IUS99105028
can determine appropriate parameters for measuring alignment, including any
algorithms needed to achieve maximal
aiigttment over the foil length of the sequences being compared. The identity
values used herein were generated by
the BLASTN module of WU-BLAST-2 set to the default parameters, with overlap
span and overlap fraction set to
1 and 0.125, respectively.
The term "positives", in the context of sequence comparison performed as
described above, includes
residues in the sequences compared that are nat identical but have similar
properties (e.g. as a result of conservative
substitutions). The °6 value of positives is determined by the fraction
of residues scoring a positive value in the
BLOSUM 62 matrix divided by the total number of residues in the aligned
region, as defined above.
The term "epitope tagged" where used herein refers to a chimeric polypeptide
comprising a PRO
polypeptide, or domain sequence thereof, fused to a "tag polypeptide". The tag
polypeptitie has enough residues to
provide an epitope against which an antibody may be made, or which can be
identified by some other agent, yet is
short enough such that it does ~t interfere with the activity of the PRO
pol;ypeptide of interest. The tag polypeptide
preferably is also fairly unique so that the antibody does not substantially
cross-react with other epitopes. Suitable
tag polypeptides generally have at least six amino acid residues and usually
between about 8 to about 50 amino acid
residues (preferably, between about 10 to about 20 residues).
"Isolated," when used to describe the various polypeptides disclosed herein,
means poIypeptide that has been
identified and separated andlor recovered from a component of its natural
environment. Contaminant components
of its natural environment are materials that would typically interfere with
diagnostic or therapeutic uses for the
polypc~de, and may include enzymes, hormones; and other proteinaceous or non-
proteinaceous solutes. In preferred
embodiments, the polypeptide will be purified (1) to a degree sufficient to
obtain at least 15 residues of N-terminal
oar internal amino acid sequence by use of a spinning cup sequenator, or (2)
to homogeneity by SDS-PAGE under non-
reducing or reducing conditions using Coomassie blue or, preferably, silver
stain. Isolated polypeptide includes
polypegtide in situ within recombinant cells, since at least one component of
the PRO polypeptide natural environment
will not be present. Ordinarily, however, isolated polypeptide will be
prepared by at least one purification step.
An "isolated" PRO poiypeptide-encoding nucleic acid is a nucleic acid molecule
that is identified and
separated from at least one contaminant nucleic acid .molecule with which it
is ordinarily associated in the natural
source of the PRO polypeptide nucleic acid. An isolated PRO polypeptide
nucleic acid molecule is other than in the
form or setting in which it is found in nature. Isolated PRO polypeptide
nucleic acid molecules therefore are
distinguished from the specific PRO polypeptide nucleic acid molecule as it
exists in natural cells. However, an
isolated PRO polypeptide nucleic acid molecule includes PRO polypeptide
rnicleic acid molecules contained in cells
, that ordinarily express the PRO golypegtide where, for example, the nucleic
acid molecule is in a chromosomal
location different from that of natural cells.
The term "control sequences" refers to DNA sequences necessary for the
expression of an operably linked
coding sequence in a particular host organism. The cornrol sequences that are
suitable for prokaryotes, for example,
include a promoter, optionally an operator sequence, and a ribosome binding
site. Eukaryotic cells are known to
utilize promoters, poiyademrlation signals, and enhancers.
Nucleic acid is "operably linked" when it is placed into a fitnetional
relationship with another nucleic acid
sequence. For example, DNA for a presequence or secretory leader is operahly
linked to DNA for a polypeptide
if it is expressed as a preprotein that participates in the secretion of the
polypeptide; a promoter or enhancer is
operabiy linked to a coding sequence if it affects the transcription of the
sequet~e; or a ribosome binding site is
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WO 99!46281 PCT/US99/05028
operably linked to a coding sequence if it is positioned so as to facilitate
translation. Generally, "operabiy linked"
means that the DNA sequences being linked are contiguous, and, in the case of
a secretory leader, contiguous and
in reading phase. However, enhancers do not have to be contiguous. Linking is
accomplished by Iigation at
convenient restriction sites. If such sites do not exist, the synthetic
oligonucleotide adaptors or Linkers are used in
accordance with conventional practice.
The term "antibody~ is used in the broadest sense and specifically covers
single anti-PRO polypeptide
monoclonal antibodies (including agonist, antagonist, and neutralizing
antibodies) and anti-PRO polypeptide antibody
compositions with polyepitopic specificity. The term "monoclonal antibody" as
used herein refers to an antibody
obtained from a population of substantially homogeneous antibodies, i.e., the
individual antibodies comprising the
population are identical except for possible naturally-occurring mutations
that may be present in minor amounts.
"Active" or "activity" for the purposes herein refers to fotm(s) of PRO
polypeptide which retain the biologic
andlor immunologic activities of the specific native or naturally-occurring
PRO polypeptide.
"Treatment" or "txeating" refers to both therapeutic treatment and
prophylactic or preventative measures.
Those in need of treatment include those already with the disorder as well as
those prone to have the disorder of those
in which the disorder is to be prevented.
"Mammal~ for purposes of treaunent refers to any animal classified as a
mammal, including humans,
domestic and farm animals, and zoo, sports, or pet animals, such as sheep,
dogs, horses, cats, cows, and the like.
Preferably, the mammal herein is a human.
"Carriers" as used herein include pharmaceutically acceptable carriers,
excipients, or stabilizers which are
nontoxic to the cell or mammal being exposed thereto at the dosages and
concentrations employed. Often the
physiologically acceptable carrier is an aqueous pH buffered solution.
Examples of physiologically acceptable
carriers include buffers such as phosphate, citrate, and other organic acids;
antioxidants including ascorbic acid; low
molecular weight (less than about 10 residues) polypeptide; proteins, such as
serum albumin, gelatin, or
ittamunoglobulins; hydrophilic polymers such as poLyvinyIpyrrolidone; amino
acids such as glycine, glutatnine,
asparagine, arginine or lysine; monosaccharides, disaccharides, and other
carbohydrates including glucose. mannose,
or dextrins; chelating agents such as EDTA; sugar alcohols such as mannitol or
sorbitol; salt-forming eounterions
such as sodium; and/or nonionic surfactants such as TWEENT"", polyethyle~
glycol (PEG), and PLURONICSt"".
The term "agonist" is used to refer to peptide and non-peptide analogs of the
native PRO polypeptides
(where native PRO polypeptide refers to pro-PRO polypeptide, pre-PRO
polypeptide, prepro-PRO polypeptide, or
mature PRO polypeptide) of the present invention and to antibodies
specifically binding such native PRO
polypeptides, provided that they retain at least one biological activity of a
native PRO polypeptide. Preferably, the
agonists of the present invention retain the qualitative binding recognition
properties and receptor activation properties
of the native PRO poiypeptide.
The term "antagonist" is used to refer to a molecule inhibiting a biological
activity of a native PRO
polypeptide of the present invention wherein native PRO polypeptide refers to
pro-PRO polypeptide, pre-PRO
polypeptide, prepro-PRO polypeptide, or mature PRO polypeptide. Preferably,
the antagonises herein inhibit the
binding of a native PRO polypeptide of the present im~ention to a binding
partner. A PRO polypeptide "antagonist"
is a ztmlecule which prevents, or interferes with, a PRO antagonist effector
function (e.g. a molecule which prevents
or interferes with binding and/or activation of a PRO poLypeptide receptor by
PRO polypeptide). Such molecules
can be screened for their ability to competitively inhibit PRO polypeptide
receptor activation by monitoring binding
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WO 99/46281 PCTIUS99/05028
of native PRO polypeptide in the presence and absence of the test antagonist
molecule, for example. An antagonist
of the invention also encompasses an antisense polynucleotide against the PRO
polypeptide gene, which antisense
polynucleotide blocks transcription or uanslation of the PRO polypeptide gene,
thereby inhibiting its expression and
biological activity.
"Stringency" of hybridization reactions is readily determinable by one of
ordinary skill in the art, and
generally is an empirical calculation dependent upon probe length, washing
tc;mperature, and salt concentration. In
general, longer probes require higher temperatures far proper annealing, while
shorter probes need lower
temperatures. Hybridization generally depends on the ability of denatured I)NA
to reanneal when complementary
strands are present in an environment below their melting temperature. The
higher the degree of desired homology
between the probe and hybridizabie sequence, the higher the relative
temperature which can be used. As a result,
it follows that higher relative temperatures would tend to make the reaction
conditions more stringent, while lower
temperatures less so. For additional details and explanation of stringency of
hybridization reactions, see Ausubel et
al., Current Protocols in Molecular Bioloev; Wiley Interscience Publishers,
(1995).
"Stringent conditions" means (1) employing tow ionic strength and high
temperature for washing, for
example, 0.015 sodium chlorideJ0.0015 M sodium citrateJO.l % sodium dodecyl
sulfate at 50°C, or (2) employing
during hybridization a denaturing agent, such as formamide, for example, 5096
(vollvol) formamide with 0.1 % bovine
serum aIburttin/0.l % Ficoll/0.1 % polyvinylpyrrolidane/50 nIVi sodium
phosphate buffer at pH 6.5 with 750 mM
sodium chloride, ?5 mM sodium citrate at 42°C. .Another example is use
of 50% formamide; 5 x SSC (0.75 M
NaCI, 0.075 M sodium citrate), 50 mM sodium phosphate (pH 618), 0.1 % sodium
pyrophosphate, 5 x Denhardt's
solution, sonicated salmon sperm DNA (50 ~cglml), 4.1 % SDS, and 10%a dextran
sulfate at 42°C, with washes at
42°C in 0.2 x SSC and 0.1 % SDS. Yet another example is hybridization
using a buffer of 10% dextran sulfate, 2
x SSC (sodium chloridelsodium citrate) and SO% formamide at 55°C,
followed by a high-stringency wash consisting
of 0.1 x SSC containing F.DTA at 55°C.
"Moderately srtingent cotx3itions" are described in Sambrook et al., supra,
and include the use of a washing
solution and hybridization conditions (e.g., temperature, ionic strength, and
%SDS) less stringent than described
above. An example of moderately saingent conditions is a condition such as
overnight incubation at 37°C in a
sohttion co~rising: 2096 formarnick, S x SSC (150 mM NaCI, 15 mM trisodium
citrate), 50 mM sodium phosphate
(pH 7.6), 5 x Denhardt's solution, 10% dextran sulfate, and 20 mglmL denatured
sheared salmon sperm DNA,
followed by washing the filters in 1 x SSC at about 37-50°C. The
skilled artisan will recognize how to adjust the
temperature, ionic strength, etc., as necessary to accommodate factors such as
probe length and the Like.
"Southern analysis" or "Southern blotting" is a method by which the presence
of DNA sequences in a
restriction endotntclease digest of DNA or a DNA-containing composition is
confirmed by hybridization to a known,
labeled oligonucleodde or DNA fragment. Southern analysis typically involves
electrophoretic separation of DNA
digests on agarose gels, denaturation of the DNA after electrophoretic
separation, and tramfer of the DNA to
nitr~ca xllulose, nyktrt, or another suitable membrane support for analysis
with a radiolabeled, biotinylated, or enzyme-
labeled probe as described in sections 9.37-9.52 of Sambrook et al. , M~1
color Cloning ~ A Labora~ ~nual (New
York: Cold Spring Harbor Laboratory Press, 1989).
"Northern analysis" or "Northern blotting" is a method used to identify RNA
sequences that hybridize to
a known probe such as an oliganucl~dde, DNA fragttettt, cDNA or fragment
thereof, or RNA fragment. The probe
is labeled with a radioisotope such as'~P, or by biotinylation, or with an
enzyme. The RNA to be analyzed is usually
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WO 99/46281 PCTlUS99105028
electrophoretically separated on an agarose or polyacrylamide gel, transferred
to nitrocellulose, nylon, or other
suitable membrane, and hybridized with the probe, using standard techniques
well known in the art such as those
described in sections ?.39-?.52 of Sambrook et al., supra.
As used herein, the term "immunoadhesin" designates antibody-like molecules
which combine the binding
specificity of a heterologous protein (an "adhesin") with the effector
functions of immunoglobulin constant domains.
Structurally, the itnmunoadhesins comprise a fusion of an amino acid sequence
with the desired binding speciftciry
which is other than the antigen recognition and binding site of an antibody
(i.e., is "heterologous"), and an
irnrrntnoglobuIin constant domain sequence. The adhesin part of an
immunoadhesin molecule typically is a contiguous
amino acid sequence comprising at least the binding site of a receptor or a
ligand. The immunoglobulin constant
domain sequence in the immunoadhesin tray be obtained from am immunoglobulin,
such as IgG-1, IgG-2, IgG-3,
or IgG-4 subtypes, IgA (including IgA-1 and IgA-2), IgE, IgD or IgM.
"Chronic" administration refers to administration of the agents) in a
continuous mode as opposed to an acute
mode, so as to maintain the initial therapeutic effect (activity) for an
extended period of time. "Intermittent"
administration is treatment that is not consecutively done without
interruption, but rather is cyclic in nature.
Administration "in combination with" one or more further therapeutic agents
includes simultaneous
(concurrent) and consecutive administration in any order.
As used herein, "vascular endothelial cell growth factor-E," or "VEGF-E,"
refers to a marntnalian growth
factor as ascribed herein, including the human amino acid sequence of Figure
207, a sequence which has homology
to VEGF and bone morphogenetic protein 1 and which includes complete
conservation of all VEGF cysteine residues,
which have been shown to be required for biological activity of VEGF. VEGF-E
expression includes expression in
human fetal bone, thymus, and the gastrointestinal tract. The biological
activity of native VEGF-E is shared by any
analogue or variant thereof that is capable of promoting selective growth
and/or survival of umbilical vein endothelial
cells, induces proliferation of pluripotent fibroblast cells, induces
immediate early gene c-fos in human endothelial
cell lines and causes myocyte hypertrophy in cardiac cells, or which possesses
an immune epitope that is
immunologically cross-reactive with an antibody raised against at least one
epitope of the corresponding native VEGF-
E. The human VEGF-E herein is active on rat and mouse cells indicating
conservation across species. Moreover,
the VEGF-E herein is expressed at the growth plate region and has been shown
to embrace fetal myocytes.
As used herein, "vascular endothelial cell growth factor;" or "VEGF," refears
to a mammalian growth factor
as defined in U.S. Patent 5,332,671. The biological activity of native VEGF is
shared by any analogue or variant
thereof that is capable of promoting selective growth of vascular endothelial
cells but not of bovine corneal endothelial
cells, lens epithelial cells, adrenal cortex cells, BHiK-21 fibroblasts, or
keratinocytes, or that possesses an immune
epitope that is immunologically cross-reactive with an antibody raised against
at least one epitope of the corresponding
native VEGF.
The terms "VEGF-E poiypeptide" and "VEGF-E" when used herein encompass native
sequence VEGF-E
poiypeptide and VEGF-E palypeptide variants (which are further defined
herein). The VEGF-E polypeptides may
be isolated from a variety of sources, such as from human tissue types or from
another source, or prepared by
recombinant or synthetic methods.
Inhibitors of VEGF-E include those which reduce or inhibit the activity or
expression of VEGF-E and
includes antisense molecules.
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WO 99146281 PCTIUS99145028
The abbreviation "ICDR" refers to the kinase domain region of the VEGF
molecule. VEGF-E has no
homology with VEGF in this domain.
The abbreviation _"FLT-1 " refers to the FMS-like tyrosine kinase binding
domain which is known to bind
to the corresponding FLT-I receptor. VEGF-E has no homology with VEGF in this
domain.
"Toll receptor2", "TLRZ" and "huTLR2" are used interchangeably, and refer to a
human Toll receptor
designated as "HuT'LRZ" by Rack et al:, Proc. htatl. Acad. Sci. USA ~5, 588-
593 (1998).
The term "expression vector" is used to define a vector, in which a raucleic
acid encoding a PRO polypeptide
herein is operably linked to control sequences capable of affecting its
expression is a suitable host cells. Vectors
ordinarily carry a replication site (although this is not necessary where
chromosomal integration will occur).
Expression vectors also include marker sequences which are capable of
providing phenotypic selection in transformed
cells. For example, E. colt is typically transformed using pBR322, a plasmid
derived from an E. colt species
(BoIivar, et al., Gene ~: 95 (I977]). pBR322 contains genes for ampicillin and
tetracycline resistance and thus
provides easy means for identifying transformed cells, whether for purposes of
cloning or expression. Expression
vectors also optimally will contain sequences which are useful for the control
of transcription and translation; e.g.,
promoters and Shine-Dalgatno sequences (for prokaryotes) or promoters and
enhancers (for mamnwlian cells). The
prottwters may be, but need not be, inducible; even powerful constitutive
promoters such as the CMV promoter for
mammalian hosts have been found to produce the LHR without host cell toxicity.
While it is conceivable that
expression vectors need not contain any expression control, replicative
sequences or selection genes, their absence
may hamper the identification of hybrid transformants and the achievement of
high level hybrid immunoglobulin
expression.
The term "tipopolysaccharide" or "LPS" is used herein as a synonym of
"endotoxin." Lipopolysaccharides
(LPS) are characteristic cornpor~nts of the outer membrane of Grate-negative
bacteria, e.g., Escherichia colt. They
consist of a polysaccharide part and a fat called Lipid A. The polysaccharide,
which varies from one bacterial species
to another, is made up of the O-specific chain (built from repeating units of
three to eight sugars) and the two-part
core. Lipid A virtually always includes two glucosamine sugars modified by
phosphate and a variable number of fatty
acids. For further information see, for example, ltietschel and Brade,
Scientific merican August 1992, 54-6I.
The term "septic slmek" is used herein in the broadest sense, including all
defunftions disclosed in Bone, ~t
Intern Med. l~,g, 332-333 (1991). Specifically, septic shock starts with a
systemic response to infection, a syndrome
called sepsis. When this syndrome results inhypotension and organ dysfunction,
it is called septic shock. Septic
shock may be initiated by gram-positive organisms and fungi, as welt as
endotoxin-containing Grate-negative
organisms. Accordingly, the present definition is not limited to "endotoxin
shock."
The phrases "gene amplification" and "gene duplication" are used
interchangeably and refer to a process
by which multiple copies of a gene or gene fragment arc formed in a particular
cell or cell line. The duplicated region
(a stretch of amplified DNA) is often referred to as "amplicon". Usually, the
amount of the messenger RNA
(mRNA) produced, i.e., the level of gene expression, also increases in the
proportion of the number of copies made
of the particular gene expressed.
"'I~mor", as used herein, refers to all neoplastic cell growth and
proliferation, whether malignant or benign,
and all pre-cancerous and cancerous cells and tissues: The terms "cancer" and
"cancerous" refer to or describe the
physiological condition in mammals that is typically characterized by
unregulated cell growth. Examples of cancer
include but are not limited to, carcinoma, lymphoma, blastoma, sarcoma, and
leukemia. More particular examples
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WO 99146281 , PCT/US99/05028
of such cancers include breast cancer, prostate cancer, colon cancer, squamous
cell cancer, small-cell Lung cancer,
non small cell lung cancer, gastrointestinal cancer, pancreatic cancer,
gliobIastoma, cervical cancer, ovarian cancer,
liver cancer, bladder cancer, hepatoma, colorectal cancer, endometrial
carcinoma, salivary gland carcinoma, kidney
cancer, vulval cancer, thyroid cancer, hepatic carcinoma and various types of
head and neck cancer.
The term "cytotoxic agent" as used herein refers to a substance that inhibits
or prevents the function of cells
andlor causes destruction of cells. The term is intended to include
radioactive isotopes (e.g. I131, I125, Y90 and
Re186), chemotherapeutic agents, and toxins such as enzymatically active
toxins of bacterial, fungal, plant or animal
origin, or fragments thereof.
A "chemotherapeutic agent" is a chemical compound useful in the treatment of
cancer. Examples of
chemotherapeutic agents include adriamycin, doxorubicin, epirubicin, 5-
fiuorouracil, cytosine arabinoside ("Ara-C "),
cyclophosphamide, thiotepa, busulfan, cytoxin, taxoids, e.g. paclitaxel
(Taxol; Bristol-Myers Squibb Oncology,
Princeton, NJ), and doxetaxei (Taxotere~~, Rhone-Pouienc Rorer, Antony,
France), toxotere, methotrexate, cisplatin,
melphalan, vinblastine, bleomycin, etoposide, ifosfamide, mitomycin C,
mitoxantrone, vincristine, vinorelbine,
carboplatin, teniposide, daunomycin, carminomycin, aminopterin, dactinomycin,
mitomycins, esperannicins (see U.S.
Pat. No. 4,675,187), melphalan and other related nitrogen mustards. Also
included in this defueition are hormonal
IS agents that act to regulate or inhibit hormone action on rumors such as
tamoxifen and onapristone.
A "growth inhibitory agent" when used herein refers to a compound or
composition which inhibits growth
of a cell, especially cancer cell overexpressing any of the genes identified
herein, either in viuo or in vivo. Thus,
the growth inhibitory agent is one which significantly reduces the percentage
of cells overexpressing such genes in
S please. Examples of growth inhibitory agents include agents that block cell
cycle progression (at a place other than
S phase), such as agents that induce G1 arrest and M-phase arrest. Classical M-
phase blockers include the vincas
(vincristine and vinbiastine), taxol, and topo II inhibitors such as
doxorubicin, epirubicin, daunorubicin, etoposide,
and bleomycin. Those agents that arrest Gl also spill over into S-phase
arrest, for example, DNA aikylating agents
such as tamoxifen, prednisone, dacarbazine, mechlorethamine, cisplatin,
methotrexate, S-fluorouracil, and ara-C.
Further information can be found in The Molecular Basis of Cancer, Mendeisohn
and Israel, eds., Chapter 1, entitled
"Cell cycle regulation, oncogens, and antineoplastic drugs "by Murakami et al.
(WB Saunders: Philadelphia, 1995),
especially p.13.
"Doxorubicin" is an athracycline antibiotic.
The term "cytakine" is a generic term for proteins released by one cell
population which act on another cell
as intercellular mediators. Examples of such cytokines are iymphokines,
monokines, and traditional polypeptide
hormones. Ixicluded among the cytokines are growth hormone such as human
growth hormone, N-methionyl human
growth hormone, and bovine growth hormone; parathyroid hormone; thyroxine;
insulin; proinsuliii; relaxin;
prorelaxin; and the like. As used herein, the term cytokine includes proteins
from natural sources or from
recombinant cell culture and biologically active equivalents of the native
sequence cytokines.
"Immunological cross-reactivity" as used herein means that the candidate
polypeptide is capable of
3S competitively inhibiting the qualitative biological activity of a PR0213-1,
PR01330, or PR01449 polypeptide having
this activity with polyclonal antisera raised against the known active PR0213-
1, PR01330, or PR01449 polypeptide.
Such antisera may be prepared in conventional fashion by injecting goats or
rabbits, for example, subcutaneousiy with
the known active analogue in complete Freund's adjuvant, followed by booster
intraperitoneal or. subcutaneous
injection in incomplete Freunds. The immunological cross-reactivity preferably
is "specific", which means that the
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WO 99/46281 PCT/US99105028
binding affinity of the immunological cross-reactive molecule (e.g., antibody)
identified, to the corresponding
PR0213-1, PR01330, or FROla49 polypegtide is significantly higher (preferably
at least about 2-times, more
preferably at least about 4-tithes, even more preferably at least about 6-
times, most preferably at least about 8-times
higher) than the binding afftniry of that molecule to any other known native
polypeptide.
"Native antibodies" and "native immunoglobulins" are usually heterotetrameric
glycoproteins of about
150,000 daltons, composed of two identical light (L) chains and two identical
heavy (H) chains. Each light chain
is linked to a heavy chain by one covalent disulfide bond, while the number of
disulfide linkages varies among the
heavy chains of differern itnmunoglobulin isotypes. Each heavy and light chain
also has regularly spaced intrachain
disulfide bridges. Each heavy chain has at one end a variable domain (VH)
followed by a number of constant
domains. Each Sight chain has a variable domain at one end (VL) and a constant
domain at its other end; the constant
IO domain of the light chain is aligned with the first constant domain of the
heavy chain, and the light chain variable
don~airris aligned with the variable domain of the heavy chain. Pat2icular
amino acid residues are believed to form
an interface between the light- and heavy-chain variable domains.
The term "variable" refers to the fact that certain portions of the variable
domains differ extensively in
sequence among antibodies and are used in the binding and specificity of .each
particular antibody for its particular
antigen. However, the variability is not evenly distributed throughout the
variable domains of antibodies. It is
concentrated in three segments called complementarily-determining regions
(CDRs) or hypervariable regions both
in the light-chain and the heavy-chain variable domains. The more highly
conserved portions of variable domains
are called the framework (FR). The variable domains of native heavy and light
chains each comprise four FR
regions, largely adopting a ~i-sheet configuration, connected by three CI)Rs,
which form loops connecting, and in
some cases forming part of, the (i-sheet structure. The CDRs in each chain are
held together in close proximity by
the FR regions and, with the CDRs from the other chain, cornribute to the
formation of the antigen-binding site of
ana'bodies (see Kabat et al., NIH Publ. No.91-3242, Vol. l, pages G47-(i69
(1991)). The constant domains are not
involved directly in binding an antibody to an antigen, but exhibit various
effector functions, such as participation
of the antibody in antibody-dependent celhzlar toxicity.
"Antibody fragments" comprise a portion of an intact antibody, preferably the
antigen binding or variable
region of the intact antibody. Examples of antibody fragments include Fab,
Fab', F(ab')2, and Fv fragments;
diabodies; linear antibodies (Zapata et al. , Protein Eng. 8(10): 105'7-1062
[1995J); single-chain antibody molecules;
and multispecifrc antibodies formed from antibody fragments.
Papain digestion of antibodies produces two identical antigen-binding
fragments, called "Fab" fragments,
each with a single antigen binding site, and a residual "Fc" fragment, a
designation reflecting the ability to crystallize
readily. Pepsin treatment yields an F(ab'~ fragttiettt that has two.antigen
combining sites and is still capable of cross-
linking antigen.
"Fv" is the minimum antibody fragment which contains a complete antigen-
recognition and -binding site.
This region consists of a dirtier of one heavy- and one light-chain variable
domain in tight, non-covalent association.
It is in this configuration that the three CDRs of each variable domain
interact to define an antigen-binding site on
the surface of the VH-VL dimer. Collectively, the six CDRs confer antigen-
binding specificity to the antibody.
However, even a single variable domain (or half of an Fv comprising only three
CDRs specific for an antigen) has
the ability to recognize and bind antigen; although at a lower affinity than
the entire binding site:
I2I
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The Fab fragment also contains the constant domain of the light chain and the
first constant domain (CH l )
of the heavy chain. Fab fragments differ from Fab fragments by the addition of
a few residues at the carboxy
terminus of the heavy chain CHl domain including one or more cysteines from
the antibody hinge region. Fab'-SH
is the designation herein for Fab' in which the cysteine residues) of the
constant domains bear a free thiol group.
F(ab')2 antibody fragments originally were produced as pairs of Fab' fragments
which have hinge cysteines between
them. Other chemical couplings of antibody fragments are also known.
The "light chains" of antibodies (immunoglobulins) from any vertebrate species
can be assigned to one of
two clearly distinct types, called kappa and lambda, based on the amino acid
sequences of their constant domains.
Depending on the amino acid sequence of the constant domain of their heavy
chains, itnmunoglobulins can
be assigned to different classes. There are five major classes of
immunoglobulins: IgA, IgD, IgE, IgG, and IgM,
and several of these may be farther divided into subclasses (isotypes), e.g.,
IgGl, IgG2, lgG3, IgG4, IgA, and IgA2.
"Single-chain Fv" or "sFv" antibody fragments comprise the VH and VL domains
of antibody, wherein
these domains are present in a single polypeptide chain. Preferably, the Fv
polypeptide further comprises a
polypeptide linker between the VH and VL domains which enables the sFv to form
the desired structure for antigen
binding. For a review of sFv see Pluckthun in The Pharmacology of Monoclonal,
Antibodies, vol. 113, Rosenburg
and Moore eds., Springer-Verlag, New York, pp. 269-315 (1994).
The term "diabodies" refers to small antibody fragments with two antigen-
binding sites, which fragments
comprise a heavy-chain variable domain (VH) connected to a light-chain
variable domain (VL) in the same
poiypeptide chain (VH - VL). By using a linker that is too short to allow
pairing between the two domains on the
same chain, the domains are forced to pair with the complementary domains of
another chain and create two antigen-
binding sites. Diabodies are described more fully in, for example, EP 404,097;
WO 93II1161; and Hollinger et al.,
Proc. Nat!. Acad. Sci. USA, 90:b444-6448 (1993).
An "isolated" antibody is one which has been identified and separated and/or
recovered from a component
of its natwal environment. Contaminate components of its natural environment
are materials which would interfere
with diagnostic or therapeutic uses for the antibody, and may include enzymes,
hormones, and other proteinaceous
or nonproteinaceous solutes. In preferred embodiments, the anubody will be
purified (1) to greater than 95% by
weight of antibody as determined by the Lowry method, and most preferably more
than 99~ by weight, (2) to a
degree sufficient to obtain at Least 15 residues of N-terminal or internal
amino acid sequence by use of a spinning cup
sequenator, or (3) to homogeneity by SDS-PAGE under reducing or noraeducing
conditions using Coomassie blue
or, preferably, silver stain. Isolated antibody includes the antibody in situ
within recombinant veils since at least one
component of the antibody's natural environment will not be present.
Ordinarily, however, isolated antibody will
be prepared by at least one purification step.
The word "label" when used herein refers to a detectable compound or
composition which is conjugated
directly or indirectly to the antibody so as to generate a "labelled"
antibody. The label tnay be detectable by itself
(e.g. radioisotope labels or fluorescem Labels) or, in the case of an
enzymatic label, may catalyze chemical alteration
of a substrate compound or cottiposition which is detestable.
By "solid phase" is meant a non-aqueous matrix to which the antibody of the
present invention can adhere.
Exattiples of solid phases encompassed herein include those formed partially
or entirely of glass (e.g., controlled pore
glass), polysaccharides (e.g., agarose), polyacrylamides, polystyrene,
polyvinyl alcohol and silicones. In cetYairi
embodiments, depending on the context, the solid phase can comprise the well
of an assay plate; in others it is a
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purification cohlmn (e.g., an affinity chromatography column). This term also
includes a discontinuous solid phase
of discrete particles, such as those described in U.S. Patent No. 4,275,149.
A "liposome" is a small vesicle composed of various types of lipids,
phospholipids and/or surfactant which
is useful for delivery of a drug (such as the anti-ErbB2 ann'bodies disclosed
herein and, optionally, a chemotherapeutic
agent) to a mattunal. The components of the liposome are commonly arranged in
a bilayer formation, similar to the
lipid arrangement of biological membranes.
II. Com~sitions and Methods Q,f the Invention
1. Fill!-~e~gth PR0213 Polypept~~
The present invention provides newly identified and isolated nucleotide
sequences encoding polypeptides
referred to in the present application as PR0213. In particular, Applicants
have identified and isolated cDNA
encoding a PR0213 polypeptide, as disclosed in further detail in the Examples
below. Using BLAST and FastA
sequence alignment computer programs, Applicants found that a portion of the
PRO213 polypeptide has sigrtificant
homology with the human growth arrest-specific 6 {gash) protein. Accordingly,
it is presently believed thai PR0213
polypeptide disclosed in the present application may have the same or simtilar
activity as does the gash protein.
2. . Furl-length PR0274 Pwnentides
The present invention provides newly identified and isolated nucleotide
sequences encoding polypeptides
referred to in the present application as PR0274. In particular, Applicants
have identified and isolated cDNA
encoding a PR0274 polypeptide, as disclosed in further detail in the Examples
below. Using BLAST and FastA
sequence alignment computer programs, Applicants found that various portions
of the PR0274 polypeptide have
significant homology with the ? transmembrane segment receptor proteins and
FnS4 protein. Accordingly, it is
presently believed that PR0274 polypeptide disclosed in the present
application is a newly identified member of the
? transmembrane segment receptor protein andlor Fn54 protein family.
3. Full-length PR0300 Polvoep~ide
The present invention provides newly identified and isolated nucleotide
sequences encoding polypeptides
referred to in the present application as PR0300. Ln particular, Applicants
have identified and isolated cDNA
encoding a PR0300 polypeptide, as disclosed in further detail in the Examples
below. Using BLAST and FastA
sequence alignment computer programs, Applicants found that various poxtions
of the PR0300 polypeptide have
significant homology with the human Difl;' 33 protein. Accordingly, it is
presently believed that PR0300 polypeptide
disclosed in the present application is a newly identified mem>~r of the Diff
33 family.
4. Full-to h PR0284 Poly~e ti'~de~c
The present invention provides newly identified and isolated nucleotide
sequences encoding polypeptides
referred to in the present application as PR0284. In particular, Applicants
have identified and isolated cDNA
encoding a PR0284 polypeptide, as disclosed in further detail in the Examples
below, To Applicants present
Imowledge, the UNQ247 (DNA23318-1211) nucleotide sequence encodes a novel
factor; using BLAST and FastA
sequence alignment computer programs, no sequence identities to arty lmown
proteins were revealed.
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5. Full-leneth PR0296 Polypeptides
The present invention provides newly identified and isolated nucleotide
sequences encoding polypeptides
referred to in the present application as PR0296. In particular, Applicants
have identified and isolated cDNA
encoding a PR0296 polypeptide, as disclosed in further detail in the Examples
below. Using. BLAST and FastA
sequence aligntr~ent computer programs, Applicants found that the PR0296
polypeptide has significant similarity to
the sarcoma-amplified SAS protein. Accordingly, it is presently believed that
PR0296 polypeptide disclosed in the
present application is a newly identified SAS protein homolog.
6. Full-length PR0329 Polvpeptides
The present invention provides newly identified and isolated nucleotide
sequences encoding polypeptides
referred to in the present application as PR0329. in particular, Applicants
have identified and isolated cDNA
encoding a PR0329 poiypeptide, as disclosed in further detail in the Examples
below. Using BLAST and FastA
sequence alignment computer programs, Applicants found that the PR0329
polypeptide has significant similarity to
a high affuiity immunoglobulin F~ receptor. Accordingly, it is presently
believed that PR0329 polypeptide disclosed
in the present application is a newly identified F~ receptor homolog.
?. Full-length PR0362 Poiy~eptides
The present invention provides newly identified and isolated nucleotide
sequences encoding polypeptides
referred to in the present application as PR03b2. In particular, Applicants
have identified and isolated cDNA
encoding a PR0362 polypeptide, as disclosed in further detail in the Examples
below. Using BLAST and FastA
sequence alignment computer programs, Applicants found that the PR0362
polypeptide has significant similarity to
the A33 antigen protein as well as the HCAR protein and the NrCAM related cell
adhesion molecule. Accordingly,
it is presently believed that PR0362 polypeptide disclosed in the present
application is a newly A33 antigen and
HCAR protein homolog.
8. Full-length PR0363 Polypeptides
The present invention provides newly identified and isolated nucleotide
sequences encoding polypeptides
referred to in the present application as PR0363. In particular, Applicants
have identified and isolated cDNA
encoding a PR0363 polypeptide, as disclosed in further detail in the Examples
below. Using BLAST and FastA.
sequence alignment computer programs, Applicants found that the PR0363
polypeptide has significant similarity to
tl~ cell surface protein HCAR. Accordingly, it is presently believed that
PRO363 polypeptide disclosed in the present
application is a newly HCAR hotnolog.
9. dull-leneth PR0868 Polvoeotides
The present invention provides newly identified and isolated nucleotide
sequences encoding polypeptides
referred to in the present application as PR0868. In particular, Applicants
have identified and isolated cDNA
encoding a PR08b8 polypeptide, as disclosed in further detail in the Examples
below. Using BLAST and FastA
sequence aiigttinettt computer programs, Applicants found that the PR0868
polypeptide has significant similarity to
the tumor necrosis factor receptor. Accordingly, it is presently believed that
PR0868 golypeptide disclosed in the
present application is a newly identified member of the tumor necrosis factor
receptor family of proteins.
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I0. Full-le~gtjh PR0382 Poivoeotides
The present invention provides newly identified and isolated nucleotide
sequences encoding polypeptides
referred to in the present.application as PR0382. In particular, App3icants
have identified and isolated cDNA
encoding a PR0382 polypeptide, as disclosed in further detail in the Examples
below. Using BLAST and FastA
sequence alignment computer programs, Applicants found that the native PR0382
polypeptide shares significant
S homology with various serine protease proteins. Applicants have also found
that the DNA encoding the PR0382
polypeptide shares significant homology with nucleic acid encoding various
setvte protease proteins. Accordingly,
it is presently believed that PR0382 polypeptide disclosed in the present
application is a newly identified serine
protease homolog.
11. Full-ie~gth PR0545 Polvneotides
The present invention provides newly identified and isolated nucleotide
sequences encoding polypeptides
referred to in the present application as PROS4S. In particular, Applicants
have identified and isolated cDNA
encoding a PR0545 polypeptide, as disclosed in further detail in the Examples
below. Using BLAST and FastA
sequence alignnnent computer programs, Applicants found that various portions
of the PROS4S polypeptide have
significant homology with the sequences identified designated as; human
metalloproteinase ("P W01825 "), mouse
meltrin alpha ("S602S7"), mctalloprotease-disintegrin meltrin-alpha
("GENI3G9S"), ADAM 13 - Xenopeis laevis
("XLUb6003_I "), mouse meluin beta ("S60258"), rabbit metalloprotease-
disintegrin meltrin-beta, ("GEN13696"),
human meltrin S ("AF023477_1 "), human meltrin precursor ("AF023475 1 "),
human ADAM 21 ("AF029900 1 "),
and human ADAM 20 ("AF029899 1 "), thereby indicating that PROS4S may be a
novel mcltrin protein.
Accordingly, it is presently believed that the PROS4S polypeptide disclosed in
the present application is a newly
identified member of the meltrin family and possesses the cellular
adhesiveness typical of the rneltrin proteins which
comprise both metalloprotease and disintegrin domains.
12: Full-lenEth PR0617 Pojv~ptldes
The present invention provides newly identified and isolated nucleotide
sequences encoding polypeptides
referred to in the present application as PR0617. In particular, Applicants
have identified and isolated cDNA
encoding a PR0617 polypeptide, as disclosed in further detail in the Examples
below. Using BLAST and FastA
sequence alignment computer-programs, Applicants found that the PR0617
polypeptide shares significant homology
with the CD24 protein. Applicants have also found that the DNA encoding the
PR06I7 polypeptide has significant
homology with DNA encoding the CD24 protein. Accordingly, it is presently
believed that PR0617 polypeptide
disclosed in the present application is a newly identified CD2.4 homolog.
13. ly'ull-length laRQ700 Pol3y to ides
The present invention provides newly identified and isolated nucleotide
sequences encoding polypeptides
3S referred to in the present application as PRO700. In particular, Applicants
have identified and isolated cDNA
encoding a PR0700 polypeptide, as disclosed in further detail in the: Examples
below. Analysis of the amino acid
sequence of the fall-length PR0700 polypeptide using BLAST and FastA sequence
alignment computer programs,
suggests that various portions of the PR0700 polypeptide possess significant
sequence similarity to various protein
disulfide isomerases. More specifically, an analysis of the Dayhoff database
(version 35.45 SwissProt 3S) evidenced
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significant sequence similarity between the PR0700 amino acid sequence and the
following Dayhoff sequences;
polypeptide with protein disulfide isomerase activity, designated as ("P
P80664"), human FDI, designated as
("P~R51696"), human PDI, designated as (P 825297"), probable protein disulfide
isomerase er-60 precursor,
designated as ("ER60 SCHMA"), protein disulfide isomerase precursor -
Drosophila n:elanogaster, designated as
("PDI DROME"), protein disulfide-isomerase precursor - Nicotiana tabaccum,
designated as ("NTPDIGENE 1 "),
protein disulfide isomerase - Onchocerca volvulus, designated as ("OW 22440 I
"), human probable protein disulfide
isomerase p5 precursor , designated as ("ERPS HUMAN"), human protein disulfide
isomerase-related protein 5,
("HSU79278_l "), and protein disulfide isomerase precursor J prolyl 4-
hydroxy, ("PDI HUMAN"), thereby
indicating that PR0700 may be a novel protein disulfide isomerase.
Accordingly, it is presently believed that
PR0700 polypeptide disclosed in the present application is a newly identified
member of the protein disulfide
isomerase family and possesses the ability to catalyze the formation of
disulfide bonds typical of the protein disulfide
isomerase family.
14. ~u11-length PR0702 Poiypeptides
The present invention provides newly identified and isolated nucleotide
sequences encoding polypeptides
referred to in the present application as PR0702. In particular, Applicants
have identified and isolated cDNA
encoding a PR0702 polypeptide, as disclosed in further detail in the Examples
below. Using BLAST and FaSIA
sequence alignment computer programs, Applicants found that the PR0702
polypeptide has significant similarity to
the conglutinin protein. Accordingly, it is presently believed that PR0702
polypeptide disclosed in the present
application is a newly identified conglutinin homolog.
15. Full length PR0703 Polypeotides
The present invention provides newly identified and isolated nucleotide
sequences encoding polypeptides~
referred to in the present application as PRO?03. In particular, Applicants
have identified and isolated cDNA
encoding a PR0703 polypeptide, as disclosed in further detail in the Examp2es
below. Analysis of the amino acid
sequence of the full-length PR0703 polypeptide using BLAST and FastA sequence
alignment computer programs,
suggests that various portions of the PR0703 polypeptide possess significant
sequence similarity to the VLCAS
protein, thereby indicating that PR0703 may be a novel VLCAS protein. More
specifically, an analysis of the
Dayhoff database (version 35.45 SwissProt 35) evidenced significam sequence
similarity between the PR0703 amino
acid sequence and the following Dayhoff sequences, human m'RNA for very-long-
chain acyl-CoA, ("D88308"), rat
mRNA for very-long-chain aryl-CoA synthetase, ("D85100"), Mus musculus fatty
acid uansport protein,
("MMU15976"), human very-long-chain aryl-CoA synthetase, ("D88308~1 "), Mus
musculus very-long-chain
aryl-CoA synthetase, ~'AI~033031 1 "), very-long-chain acyl-CoA synthetase -
Ramcs, ("D85100~1 "), rat long-chain
fatty acid transport protein, ("FATP,-RAT"), mouse long-chain fatty acid
transport protein, ("FATP~MOUSE"),
probable long-chain fatty acid transport protein, ("FATi YEAST"), and fatty
acid transporter protein,
("CHY15839 2") , thereby indicating that PR0703 may be a novel VLCAS.
Accordingly, it is presently believed
that PR0703 polypeptide disclosed in the present application is a newly
identified member of the VLCAS family and
possesses the ability to facilitate the cellular transport of long and very
long chain fatty acids typical of the VLCAS
family.
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16. Full-IenQt~ PR0705 Polvnentides
The present invention provides newly identified and isolated nucleotide
sequences encoding polypeptides,
referred to in the present application as PRO705. In particular, Applicants
nave identified and isolated cDNA
encoding a PRO705 polypeptide, as disclosed in further detail in the Examples
below. Using BLAST and FastA
sequence atignt~nt computer programs, Applicants found that the PR0705
polypeptide has significant similarity to
the K-gIypican protein. Accordingly, it is presently believed that PR0705
polypeptide disclosed in the present
application is a newly identified member of the glypican family of
proteoglycan proteins.
l?. Full-lgngth P~Q708 Polvoeutides
The present invention provides newly identified and isolated nucleotide
sequences encoding polypeptides
referred to in the present application as PRO?08. In particular, Applicants
have identified and isolated cDNA
encoding a PR0708 polypeptide, as disclosed in further detail in the Examples
below. Using BLAST and FastA
sequence alignment computer programs, Applicants found that the PR070$
polypeptide has significant homology with
the aryl suifatase proteins. Applicants have also found that the DNA encoding
the PR0708 polypeptide has
significant homology with DNA encoding the aryl sulfatase proteins.
Accordingly, it is presently believed that
PR0708 polypeptide disclosed in the present application is a newly identified
aryl sulfatase homolog.
18. Full-le h P_R_-O~'~0 Palvaeotides
The present invention provides newly identified and isolated nucievtide
sequences encoding polypepiides
referred to in the present application as PR0320. In particular, Applicants
have identified and isolated cDNA
encoding a PR0320 polypeptide, as disclosed in further detail in the Examples
below. Analysis of the amino acid
sequence of the full-length PR0320 polypeptide using BLAST and FastA sequence
alignment computer programs,
suggests that various portions of the PR0320 polypeptide have significant
homology to the fibulin protein. More
specifically, an analysis of the Dayhoff database (version 35.45 SwissProt 35)
evidenced significant homology
between the PR0320 amino acid sequence and the following Dayhoff sequences,
human fibulin-2 precursor,
designated "FBL2,HUMAN", hutt~an fibulin-1 isofotm a precursor, designated
"FBLA HUMAN", ZK783.1
Caenorhabditis elegans, designated "CELZK783_l ", human-notch2, designated
"HSU77493 I", Nel protein
psectusor - rattus norvegicus, designated "NEL RAT", Mus musculus cefl surface
protein, designated "D32210 1 ",
mouse (fragment) Notch B protein, designated '"A49175", C50H2.3a -
Caenorhabditis elegans, designated
"CECSOH2~3", MEC-9L - Caenorhabditis elegans, designated "CEt)33933_1 ", and
Mus musculus notch 4,
designated "10 MMMHC29N7l2", thereby indicating that PRO320 may be a novel
fitiulin or fibulin-like protein.
Accordingly, it is preseniiy believed that PR0320 poIypeptide disclosed in the
present application is a newly identified
member of the fibuIin family and possesses biological activity typical of the
fibulin family.
19. Full-len~h, P]It032a Polvnentides
The present invention provides newly identified and isolated nucleotide
sequences encoding polypeptides
referred to in the present application as PR0324. In particular, Applicants
have identified and isolated eDNA
encoding a PR0324 polypeptide, as disclosed in further detail in the Examples
below. Using BLAST and FastA
seqt~ce align computer programs, Applicants found that the PR0324 polypeptide
has significant similarity to
oxidorecbuctases. Acmrdiiigly, it is presently believed that PRO324
polypeptide disclosed in the present application
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is a newly identified oxidoreductase homolog.
20. Full-length PR0351 Polypeptides
The present invention provides newly identified and isolated nucleotide
sequences encoding polypeptides
referred to in the present application as PR0351. In particular, Applicants
have identified and isolated cDNA
encoding a PRO35I polypeptide, as disclosed in further detail in the Examples
below. Analysis of the amino acid
sequence of the full-length PR0351 polypeptide using BLAST and FastA sequence
alignment computer programs,
suggests that various portions of the PR0351 polypeptide possess significant
sequence similarity to the prostasin
protein, thereby indicating that PR0351 may be a novel prostasin protein. More
specifically, an analysis of the
Dayhoff database (version 35.45 SwissProt 35) evidenced significant sequence
similarity between the PR0351 amino
acid sequence and the following Dayhoff sequences, "AC003965_1", "CELLO?Gi ?",
"GEN1291?",
°HEPS HUMAN», "GF.N14584", "MCT6~MOUSE", "HSU7532911", "PLMN ERIEU",
"TRYB HUMAN", and
"P W2298?". Accordingly, it is presently believed that FR0351 polypeptide
disclosed in the present application is
a newly identified member of the prostasin family and possesses properties and
activities typical of the prostasin
family.
21. Full-length PR0352 Polypeptides
The present invention provides newly identified and isolated nucleotide
sequences encoding polypeptides
referred to in the present application as PR0352. In particular, Applicants
have identified and isolated cDNA
encoding a PR0352 polypeptide, as disclosed in further detail in the Examples
below. Using BLAST and FastA
sequence alignment computer programs, Applicants found that the PR0352
polypeptide has significant similarity to
the buryrophilin protein. Accordingly, it is presently believed that PR0352
polypeptide disclosed in the present
application is a newly identified butyrophilin homolog.
22. dull-length PR0381 Polypeptides
The present invention provides newly identified and isolated nucleotide
sequences encoding polypeptides
referred to in the present application as PR0381. In particular, Applicants
have identified and isolated eDNA
encoding a PR0381 polypeptide, as disclosed in further detail in the Examples
below. Using BLAST and FastA
sequence alignment computer programs, Applicants found that the PR0381
polypeptide has significant similarity to
immunoghilin groteins. Accordtngiy, it is presently believed that PR0381
polypeptide disclosed in the present
application is a newly identified FKBP immunophilin homolog.
23. FuliaenQth PRO386 Polvpeptides
The present invention provides newly identified and isolated nucleotide
sequences encoding golypeptides
referred to in the present application as PR0386. In particular, Applicants
have identified and isolated cDNA
encoding a PR0386 polypeptide, as disclosed in further detail in the Examples
below. Using BLAST and FastA
seqt>etice alignment computer programs, Applicants found that the PR0386
polypeptide has significant similarity to
the beta-2 subunit of a sodium channel protein. Accordingly, it is presently
believed that PR0386 polypeptide
disclosed in the present application is homolog of a beta-2 subunit of a
sodium channel expressed in mammalian cells.
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24. Full-length PR054t1 Polvneotides
The present invention provides newly identified and isolated nucleotide
sequences encoding polypeptides
referred to in the present application as PROS40. In particular, Applicants
have identified and isolated cDNA
encoding a PR0540 polypeptide, as disclosed in further detail in the Examples
below. Analysis of the amino acid
sequence of the full-length PR0540 polypeptide using BLAST and FastA sequence
alignment computer programs,
suggests that various portions of the PR0540 polypeptide possess significant
sequence similarity to the LCAT protein,
thereby indicating that PR0540 may be a novel LCAT protein. More specifically,
an analysis of the Dayhoff
database (version 35.45 SwissProt 35) evidenced significant sequence
similarity between the PR0540 amino acid
sequence and the following Dayhoff sequences, phosphatidylcholine-sterol
acyltransferase, designated
"LCAT HUMAN", hypothetical 75.4 kd protein, designated "YN84 YEAST", Bacillus
licheniformis esterase,
designated "BLU35855_1 ", macrotetrolide resistance protein - Streptomyces,
designated "JH0555", T-cell receptor
delta chain precursor, designated "C3U583", Rhesus kringle 2, designated
"P_W07551", RAGE-1 ORFS, designated
"HSU46191 3", human Ig kappa chain VKIIi-JK3, designated "HSU0746fi_I", and
Alstroemeria inodora reverse
transcriptase, designated "AL1223606_1": Accordingly, it is presently believed
that PR0540 polypeptide disclosed
in the present application is a newly identified member of the LCAT protein
family and possesses lipid transport
capability typical of the LCAT family.
25. Full-ien$t~ PR0615 Poly eD tn ides
The present invention provides newly identified and isolated nucleotide
sequences encoding polypeptides
referred to in the present application as PR0615. In particular, Applicants
have identified and isolated cDNA
encoding a PR0615 polypeptide, as disclosed in further detail in the Examples
below. Analysis of the amino acid
sequence of the foil-length PR0615 polypeptide using BLAST and FastA sequence
alignment computer programs,
suggests that various portions of the PR0615 polypeptide possess significant
sequence similarity to the human
synaptogyrin protein, thereby indicating that PR061S tray be a novel
synaptogyrin protein. More specifically, an
analysis of the Dayhoff database (version 35.45 SwissProt 35) evidenced
significant sequence similarity between the
PR0615 amino acid sequence and the following Dayhoff sequences, "AF039085,1",
"RNU39549 1",
"CELTO8A9_8", "FSU62028_l", "S73645", "Y348 MYCPN", "AC000103_S", "", "RT12
LE1TA", and
"EBVLMP218 1". Accordingly, it is presently believed that PR0615 polypeptide
disclosed in the present application
is a newly identified member of the synaptogyrin family and possesses activity
and properties typical of the
synaptogyrin family.
26. Full-length PR0618 PolyRg tn ides .
The present invention provides newly identified and isolated rnucleotide
sequences encoding polypeptides
referred to in the present application as PR0618. In particular, AppTiicauts
have identified and isolated eDNA
encoding a PR0618 polypeptide, as disclosed in further detail in the Examples
below. Analysis of the amino acid
sequence of the full-length PR0618 polypeptide using BLAST and FastA sequence
alignment computer programs,
suggests that various portions of the PR0618 polypeptide possess significant
sequence similarity to the
enteropeptidase protein, thereby indicating that PR0618 may be a novel
enteropeptidase. More specifically, an
analysis of the Dayhoff database (version 35.45 SwissProt 35) evidenced
significant sequence similarity between the
PR0618 amino acid sequence and the following Dayhoff sequences, "P W22987",
"KAL HUMAN",
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"AC003965_I", "GEN12917", "EN'TK_HUMAN", "FA11 HUMAN", "HSU75329 1", "P
W22986", and
"PLMN HORSE". Accordingly, it is presently believed that PR0618 polypeptide
disclosed in the present application
is a newly identified member of the enteropeptidase family and possesses
catalytic activity typical of the
enteropeptidase family.
6 27. 1~ ull-len~"th ~R0719 Polvnentides
The present invention provides newly identified and isolated nucleotide
sequences encoding polypeptides
referred to in the present application as PRO?19. In particular, Applicants
have identified and isolated cDNA.
encoding a PR0719 polypeptide, as disclosed in further detail in the Examples
below. Using BLAST and FastA
sequence a3igtunent computer programs, Applicants found that the PR0719
polypeptide has significant similarity to
10' the lipoprotein lipase H protein. Accordingly, it is presently believed
that PR0719 polypeptide disclosed in the
present application is a newly identified lipoprotein lipase H homolog.
28. Full-len_eth PR0724 Polypeptides
The present invention provides newly identified and isolated nucleotide
sequences encoding polypeptides
15 referred to in the present application as PR0724. In particular, Applicants
have identified and isolated cDNA
encoding a PR0724 polypeptide, as disclosed in further detail in the Examples
below. Using BLAST and FastA
sequence aligtmnent computer programs, Applicants found that the PR0724
polypeptide has significant similarity to
the hutnatt low density lipoprotein (LDL) receptor protein. Accordingly, it is
presently believed that PR0724
polypeptide disclosed in the present application is a newly identified LDL
receptor homoiog.
29. dull IenQth PR0772 Polynentides
The present invention provides newly identified and isolated nucleotide
sequences encoding polypeptides
referred to in the present application as PR0772. In particular, Applicants
have identified and isolated cDNA
encoding a PR0772 polypeptide, as disclosed in further detail in the Examples
below. Using BLAST and FastA
sequence alignment computer programs, Applicants found that the PR0772
polypeptide has significant similarity to
the human A4 protein. Accordingly, it is presently believed that PR0772
poiypeptide disclosed in the present
application is a newly identified A4 protein homolog.
30. ~.tll lgn~th PR0852 PolvneDtides
The presept invention provides newly identified and isolated nucleotide
sequences encoding polypeptides
referred to in the present application as PR0852. In particular, Applicants
have identified and isolated cDNA
encoding a PR0852 polypeptide, as disclosed in further detail in the Examples
below. Using BLAST and FastA
sequence alignment computer programs, Applicants found that the PR0852
polypeptide has significant similarity to
various protease proteins. Accordingly, it is presently believed that PR0852
polypeptide disclosed in the present
application is a newly identified protease enzyme homolog.
31. Full-fenath PR0853 Po~,ypg t» ides
The present invention provides newly identified and isolated nucleotide
sequences encoding polypeptides
referred to in the 'present application as PR0853. In particular, Applicants
have identified and isolated cDNA
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encoding a PR0853 poIypeptide, as disclosed in further detail in the Examples
below. Analysis of the amino acid
sequence of the full-length PR0853 polypeptide using BLAST and FastA sequence
alignment computer programs,
suggests that various portions of the PR0853 polypeptide possess significant
sequence similarity to the reductase
protein, thereby indicating that PR0853 may be a novel reductase. More
specifically, an analysis of the Dayhoff
database (version 35.45 SwissProt 35) evidenced significant sequence
similarity between the PR0853 amino acid
sequence and the following Dayhoff sequences, "P W03I98", "CEC15H11 6",
"MTV030 12", "P WI5759",
"542651", "ATAC00234314", "MTV022 13", "SCU43704 I", "CELE04F6 7", and "ALFA
1". Accordingly, iL
is presently believed that PR0853 polypeptide disclosed in the present
application is a newly identified member of
the reductase family and possesses the antioxidant enzymatic activity typical
of the reductase family.
32. Full-length PR0860 Polyp~ptic~e.~
The present invention provides newly identified and isolated nucleotide
sequences encoding poiypeptides
referred to in the present application as PR0860. In particular, Applicants
have identified and isolated cDNA
encoding a PR0860 poiypeptide, as disclosed in further detail in the Examples
below. Analysis of the amino acid
sequence of the full-length PR0860 polypeptide using BLAST and FastA sequence
alignment computer programs,
suggests that various portions of the PR0860 polypeptide possess significant
sequence similarity to the neurofascin
protein, thereby indicating that PR0860 may be a novel neurofascin. More
specifically, an analysis of the Dayhoff
database (version 35.45 SwissProt 35) evidenced significant sequence
sinviarity between the PR0860 amino acid
seqtier>ce and tb:e following Dayhoff sequences, "AF040990 1 ", "AF041053_I",
"CELZK377 2", "RNU8I035 1",
"D86983_I", "S2b180", "MMBIG2A 1", "546216", and "RNU68726 1". Accordingly, it
is presently believed that
PR0860 polypeptide disclosed in the present application is a newly identified
member of the neurofascin family and
possesses the cellular adhesion properties typical of the neurofascin family.
33. Full-leng~, PR~6 Poly~e, tides
The present invention provides newly identified and isolated nucleotide
sequences encoding polypeptides
referred to in the present application as PR0846. In particular, Applicants
have identified and isolated cDNA
encoding a PR0846 polypeptide, as disclosed in further detail in the Examples
below, Analysis of the amino acid
sequence of the full-length PR0846 polypeptide using BLAST and FastA sequence
aiignrnent computer programs;
suggests that various portions of the PR0846 polypeptide possess significant
sequence similarity to the CMRF35
protein, thereby indicating that 1'R0846 may be a novel CMRF35 protein. More
specifically, an analysis of the
Dayhoff database (version 35.45 SwissProt 35) evidenced significant sequence
similarity between the PR0846 amino
acid sequence and the following Dayhoff sequences,"CM35~HUIVtAN",
"AF035963_1", "PIGR~RABTT",
"AF043724;1 ", "RNU89744 1", "A52091'1 ", "S48841", "ELK06A9 3", and "AF049588
1 ". Accordingly, it is
presently believed that PR0846 polypeptide disclosed in the present
application is a newly identified member of the
CMRF35 protein family and possesses properties typical of the CMRF35 protein
family.
34. Full-lengtl~PR0862 Polvoentldes
The present invention provides newly identified and isolated nucleotide
sequences encoding polypeptides
referred to in the present application as PR0862. In particular, Applicants
have identified and isolated cDNA
encoding a PR0862 polypepdde, as disclosed in further detail in the Examples
below. Analysis of the amino acid
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sequence of the full-length PR08b2 polypeptide using BLAST and FastA sequence
alignment computer programs,
suggests that various portions of the PR0862 polypeptide possess significant
sequence similarity to the lysoryme
protein, thereby indicating .that PR0862 may be a novel Iysoryme protein. More
specifically, an analysis of the
Dayhoff database (version 35.45 SwissProt 35) evidenced significant sequence
similarity between the PR0862 amino
acid sequence and the following Dayhoff sequences, "P P90343", and "LYC HUMAN.
Accordingly, it is presently
believed that PR0862 polypeptide disclosed in the present application is a
newly identified member of the lysozyme
family and possesses catalytic activity typical of the lysazyme family.
35. Full-length PR0~64 Polvne~tides
The present invention provides newly identified and isolated nucleotide
sequences encoding polypeptides
referred to in the present application as PR0864. In particular, Applicants
have identified and isolated cDNA
encoding a PR0864 polypeptide, as disclosed in further detail in the Examples
below. Analysis of the amino acid
sequence of the full-length PR0864 polypeptide using BLAST and FastA sequence
alignmem computer programs,
suggests that various portions of the PR0864 polypeptide possess significant
sequence similarity to the Wnt-4 protein,
thereby indicating that PR0864 may be a novel Wnt-4 protein. More
specifically, an analysis of the Dayhoff database
(version 35.45 SwissPrat 35) evidenced significant sequence similarity between
the PR0864 amino acid sequence
and the following Dayhoff sequences, "WNT4 MOUSE", "WNT3 MOUSE", "WNSA HUMAN",
"~VN7B MOUSE", "WN3A MOUSE", "XLU66288 1", "WN13_IiUMAN", "WNSB ORYLA", "WNT2
MOUSE",
and "WN7A MOUSE". Accordingly, it is presently believed that PR0864
polypeptide disclosed in the present
a~lication is a newly identified member of the Wnt-4 protein family and
possesses properties typical of the Wnt-4
protein family.
36. Full-length PR0792 Polvge ides
The present invention provides newly identified and isolated nucleotide
sequences encoding polypeptides
referred to in the present application as PR0792. In particular, Applicants
have identified and isolated cDNA
encoding a PRO792 polypeptide, as disclosed in further detail in the Examples
below. Using BLAST and FastA
sequence alignment computer programs, Applicants found that the PR0792
polypeptide has significant similarity to
the CD23 protein. Accordingly, it is presently believed that PR0792
polypeptide disclosed in the present application
is a newly identified CD23 homolog.
37. Fu! ten t~h ,Pjt0866 Poivne t~ ides
The present invention provides newly ideruified and isolated nucleotide
sequences encoding polypeptides
referred to in the present application as PR0866. In particular, Applicant's
have identified and isolated cDNA
encoding a PR0866 polypeptide, as disclosed in further detail in the Examples
below. Using BLAST and FastA
sequence alignment computer programs, Applicants found that the PR0866
golypeptide has significant similarity to
various mindin and spondin proteins. Accordingly, it is presently believed
that PR0866 polypeptide disclosed in the
present application is a newly identified mindin/spondin homolog.
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38. Full-Iengt~R0873 Poly~ptides
The present invention provides newly identified and isolated nucleotide
sequences encoding polypeptides
referred to in the present application as PR0871. In particular, Applicants
have identified and isolated cDNA
encoding a PR0871 polypeptide, as disclosed in further detail in the Examples
below. Using BLAST and FastA
sequence alignment computer programs, Applicants found that the PR0871
polypeptide has significant similarity to
the CyP-60 protein. Accordingly, it is presently believed that PRO871
polypeptide disclosed in the presem
application is a newly identified member of the cycloghilin protein family and
possesses activity typical of the
cyclophilin protein family.
39. Full-length PR0873 Poiypeptides
The present invention provides newly identified and isolated nucleotide
sequences encoding polypeptides
referred to in the present application as PR0873. In particular; Applicants
have identified and isolated cDNA
encoding a PR0873 polypeptide, as disclosed in further detail in the Examples
below. Using BLAST and FastA
sequence alignmern computer programs, Applicants found that the PR0873
polypeptide has significant similarity to
a human liver carboxylesterase. Accordingly, it is presently believed that
PR0873 polypeptide disclosed in the
present application is a newly identified tnember of the carboxylesterase
family and possesses enzymatic activity
typical of ttte carboxylesterase family.
40. FuTI-length PR09a0 Polyp~p '~c,
The present invention provides newly identified anc3 isolated nucleotide
sequences encoding polypeptides
referred to in the present application as PR0940. In particular, Applicants
have identified and isolated cDNA
encoding a PR0940 polypeptide, as disclosed in further detail in the Examples
below. Using BLAST and FastA
sequence aligntnetu computer programs, Applicants found that the PR094G
polypeptide has significant similarity to
CD33 and the OB binding protein-2. Accordingly, it is presently believed that
PR0940 polypeptide disclosed in the
present application is a newly CD33 and/or OB binding protein-2 homolog.
41. Full-length PR0941 Polvnentides
The present invention provides newly identified and isolated nucleotide
sequences encoding polypeptides
referred to in the present application as PR094I. In particular, Applicants
have identified and isolated cDNA
encoding a PR0941 polypeptide, as disclosed in further detail in the Examples
below. Using BLAST and FastA
sequence ~ computer programs, Applicants found that the PR0941 polypeptide has
significant similarity to
o~ or more cadherin proteins. Accordingly, it is presently believed that
PRO941 polypeptide disclosed in the present
application is a newly identified cadherin homolog.
42. Pttli-leng~gR0~44 Poly~gp i~
The present invention provides newly identified and isolated nucleotide
sequences encoding polypeptides
referred to in the present application as PR0944. In particular, Applicants
have identified and isolated cDNA
encoding a PR0944 polypeptide, as disclosed in further detail in the Faramples
below. Using BLAST and FastA
sequence alignment computer programs; App3icants found that the PR0944
polypepdde has significant similarity to
the CPE-R cell surface protein. Accordingly, it is presently believed that
PR0944 polypeptide disclosed in the
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present application is a newly identified CPE-R homolog.
43. Full-length PR0983 Polypeptides
The present invention provides newly identified and isolated nucleotide
sequences encoding polypeptides
referred to in the present application as PR0983. In particular, Applicants
have identified and isolated cDNA
encoding a PR0983 polypeptide, as disclosed in further detail in the Examples
below. Using BLAST and FastA
sequence alignment computer programs, Applicants found that the PR0983
polypeptide has significant similarity to
the vesicle-associated protein, VAP-33. Accordingly, it is presently believed
that PR0983 polypeptide disclosed in
the present application is a newly identified member of the vesicle-associated
membrane protein family and possesses
activity typical of vesicle-associated membrane proteins.
44. Fall-ietlgth PR01057 Polypeptides
The present invention provides newly identified and isolated nucleotide
sequences encoding polypeptides
referred to in the present application as PR01057. In particular, Applicants
have identified and isolated cDNA
encoding a PRO105? polypeptide, as disclosed in further detail in the Examples
below. Using BLAST and FastA
sequence alignment computer programs, Applicants found that the PR01057
polypeptide has significant similarity
to various protease proteins. Accordingly, it is presently believed that
PR01057 polypeptide disclosed in the present
application is a newly identified protease homolog:
45. Full-length PROltI71 Polypeptides
The present invention provides newly identified and isolated nucleotide
sequences encoding polypeptides
referred to in the present application as PR01071. In particular, Applicants
have identified and isolated cDNA
encoding a PR01071 polypeptide, as disclosed in further detail in the Examples
below. Using BLAST and FastA
sequence alignment computer programs, Applicants found that the PR01071
polypeptide has significant similarity
to the thrombospondin protein. Accordingly, it is presently believed that
PR01071 polypeptide disclosed in the
present application is a newly identified thrombospondin homolog.
46. Full-length PRO't072 Polvneotides
The present invention provides newly identified and isolated nucleotide
sequences encoding polypeptides
referred to in the present application as PR01072. In particular, Applicants
have identified and isolated cDNA
encoding a PR01072 polypeptide, as disclosed in further detail in the Examples
below. Using BLAST and FastA
sequence alignment computer programs, Applicants found that the PR01072
polypeptide has significant similarity
to various reductase proteins. Accordingly, it is presently believed that
PR01072 polypeptide disclosed in the present
application is a newly identified member of the reductase protein family.
47. Full length PR01075 Po~yoeotides
The present invention provides newly identified and isolated nucleotide
sequences encoding polypeptides
referred to in the present application as PR01075. In particular, Applicants
have identified and isolated cDNA
encoding a PR01075 polypeptide, as disclosed in futtkter detail in the
Examples below. Using BLAST and FastA
sequence aligtunent computer programs, Applicants found that the PR01075
poiypeptide has significant similarity
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to protein disulfide isomerase. Accordingly, it is presently believed that
PR01075 polypeptide disclosed in the
present application is a newly identified member of the protein disulfide
isomerase family and possesses activity
typical of that family.
48. Full- enEth PRO18I PolyQeptides
The present invention provides newly identified and isolated nucleotide
sequences encoding polypeptides
referred to in the present application as PRO/81: In particular, Applicants
have identified and isolated cDNA
encoding a PRO181 polypeptide, as disclosed in further detail in the Ex<lmples
below. Using BLAST and FastA
sequence align~nt computer programs, Applicants found that the PR018I
polypeptide has significant similarity to
the cornichon protein. Accordingly, it is presently believed that PR0181
polypeptide disclosed in the present
application is a newly identified cornichon homolog.
49. Full-left hgt PR0195 Polypgptides
The present invention provides newly identified and isolated nucleotide
sequences encoding polypeptides
referred to in the present application as PR0195. In particular, Applicants
have identified and isolated cDNA
IS et~oding a PR0195 polypeptide, as disclosed in further detail in the
Examples below. The PR0195-encoding clone
was isolated from a human fetal placenta library using a capping technique
which selects for nucleotide sequences
encoding secreted proteins. To Applicants present knowledge, the UNQl69
(DNA26847-1395) nucleotide sequence
encodes a novel factor; using BLAST and FastA sequence alignment computer
programs, no sequence identities to
any known proteins were revealed.
50. ull-teng~,t PR0865 ~ brides
The present invention provides newly identified and isolated nucleotide
sequences encoding polypeptides
referred to in the present application as PR0865. In particular, Applicants
have identified and isolated cDNA
encodipg a PR086r5 polypeptide, as disclosed in further detail in the Examples
below. The PRO865-encoding clone
was isolated from a human fetal kidney library using a trapping technique
which selects for nucleotide sequences
encoding secreted proteins. Thus, the PR0865-encoding clone may encode a
secreted factor. To Applicants present
knowledge; the UNQ434 (DNA53974-1401) nucleotide sequence encodes a novel
factor; using BLAST and FastA
sequence alignment computer programs, no sequence identities to any known
proteins were revealed.
51. Full-le0;gth PR0827 Polvuentides
The present invention provides newly identified and isolated nucleotide
sequences encoding polypeptides
referred to in the present application as PR0827. In particular, Applicants
have identified and isoiaud cDNA
encoding a PR0827 polypeptide, as disclosed in fisrttter detail in the
Examples below. Using BLAST and FastA
sequence alignment computer programs, Applicants found that the PR0827
polypeptide has significant similarity to
VIA-2 and various other itvegrin proteins. Accordingly, it is presently
believed that PR0827 polypeptide disclosed
in the present application is a novel integrin protein or splice variant
thereof.
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52. Full-len~h PR01114 Po~peptides
The present invention provides newly identified and isolated nucleotide
sequences encoding polypeptides
referred to in the present application as PRO/l I4. In particular, Applicants
have identified and isolated cDNA
encoding a PR01114 polypeptide, as disclosed in further detail in the Examples
below. Using BLAST and FasiA.
sequence alignment computer programs, Applicants found that the PRO/ I I4
polypeptide has significant similarity
to the cytokine receptor family of proteins. Accordingly, it is presently
believed that PROl 114 polypeptide disclosed
in the present application is a newly identified member of the cytoldne
receptor family of proteins and possesses
activity typical of that family.
The present invention provides newly identified and isolated nucleotide
sequences encoding polypeptides
referred to in the present application as PR01114 interferon receptor
(UNQ557). In particular, cDNA encoding a
PRO1I14 interferon receptor poiypeptide has been identified and isolated, as
disclosed in further detail in the
Examples below. It is noted that proteins produced in separate expression
rounds may be given different PRO
numbers but the UNQ number is unique for any given DNA and the encoded
protein, and will not be changed.
However, for sake of simplicity, in the present specification the protein
encoded by DNA57033-1403 as well as all
fimtter native homologues and variants included in the foregoing definition of
PRO1114 interferon receptor, will be
referred to as "PR01114 interferon receptor", regardless of their origin or
mode of preparation.
Using the WLJ-BLAST2 sequence alignment computer program, it has been found
that a full-length native
sequence PR01114 interferon receptor poIypeptide (shown in Figure 142 and SEQ
ID N0:352) has sequence identity
with the other known interferon receptors. Accordingly, it is presently
believed that PR01114 interferon receptor
possesses activity typical of other interferon receptors.
53. Full-length PR0237 Polype tn ides
The present invention provides newly identified and isolated nucleotide
sequences encoding polypeptides
referred to in the present application as PR023?. In particular, Applicants
have identified and isolated cDNA
encoding a PR023? polypeptide, as disclosed in filrther detail in the Examples
below. Using BLAST and FastA
sequence alignment computer programs, Applicants found that the PR0237
polypeptide has significant similarity to
carbonic anhydrase. Accordingly, it is presently believed that PR0237
polypeptide disclosed in the present
application is a newly identified carbonic anhydrase homolog.
5q, mil ien~rth PR0541 Poivoentides
The present invention provides newly identified and isolated nucleotide
sequences encoding polypeptides
referred to in the present application as PR0541. In particular, Applicants
have identified and isolated cDNA
encoding a PR0541 polypeptide, as disclosed in further detail in the Examples
below. Using BLAST and FastA
sequence alignment computer programs, Applicants found that the PR0541
poiypeptide has significant similarity to
a zrypsin inhibitor protein. Accordingly, it is presently believed that PR0541
polypeptide disclosed in the present
application is a newly identified member of the trypsin inhibitor protein
family.
55. fit-lenvth PRO273 Polvnentidea
The present invention provides newly identified and isolated nucleotide
sequences encoding polypeptides
referred to in the present application as PR0273. In particular, Applicants
have identified and isolated cDNA
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encoding a PR0273 poIypeptide, as disclosed in further detail in the Examples
below. Using BLAST and FastA
sequence alignment computer programs, Applicants found that various portions
of the PR02?3 polypeptide have
significant sequence identity with various chemoldnes. Accordingly; it is
presently believed that PR0273 polypepdde
disclosed in the present application is a newly identified member of the
chemokine family and possesses activity
typical of the ehemolcine family.
56. Full-lenEt~R070I Polyp_eptic~es
The present invention provides newly identified and isolated nucleotide
sequences encoding polypeptides
referred to in the present application as PRO?Ol. In particular, Applicants
have identified and isolated cDNA
encoding a PR0701 polypeptide, as disclosed in further detail in the Examples
below. Using BLAST and FastA
IO sequence alignment computer programs; Applicants found that various pardons
of the PR0701 polypeptide have
significant homology with the neuroligins 1, 2 and 3 and esterases including
carboxyesterases and
acytlcholinesterases. Accordingly, it is presently believed that PR0701
polypeptide disclosed in the present
app3ication is a newly identified member of the neuroligin family and is
involved in mediating recognution processes
between neurons andlor functions as a cell adhesin molecule as is typical of
neuroligins.
IS
5?. Full-Hen~h PR0704 PolYpe
The present invention provides newly identified and isolated nucleotide
sequences encoding polypeptides
referred to in the present application as PR0704. In particular, Applicants
have identified and isolated cDNA
encoding a PR0704 polypeptide, as disclosed in further detail in the Examples
below. Using BLAST and FastA
20 sequence alignment computer programs, Applicants found that various
portions of the PR0704 polypeptide have
. significant homology with the VIP36 and GP36b. Accordingly, it is presently
believed that PR0704 polypeptide
disclosed in the present application is a newly identified member of the
vesicular integral membrane protein family
and possesses the ability to bind to sugars and cycle between the plasma
membrane and the Golgi typical of this
family.
58. Full length PR0706 Polvneotides
The present invention provides newly identified and isolated nucleotide
sequences encoding polypeptides
referred to in the present application as PR0706. In particular, Applicants
have identified and isolated cDNA
encoding a PR0706 poIypeptide, as disclosed in further detail in the Examples
below. Using BLAST and FastA
sequence alignment computer programs; Applicants found that various portions
of the PR0706 polypepdde have
sequence identity with the human prostatic acid phosphatase precursor and the
human lysosornal acid phosphatase
precursor. Accordingly, ii is presently believed that PR0706 polypeptide
disclosed in the present application is a
newly identified member of the human prostatic acid phosphatase precursor
family and possesses phosphatase typical
of the acid phosphatase fatuity.
59. Full-ieng~h PR0707 PolvpgRt~des
The present invention provides newly identified and isolated nucleotide
sequences encoding poiypeptides
referred to in the present application as PR0707. In particular, Applicants
have identified and isolated cDNA
encoding a PR0707 polypeptide, as disclosed in further detail in the Examples
below. Using BLAST and FastA
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sequence alignment computer programs, Applicants found that various portions
of the PR0707 polypeptide have
significant homology with cadherins, particularly cadherin FIB3 found in
fibroblasts. Accordingly, it is presently
believed that PR0707 poiypeptide disclosed in the present application is a
newly identified member of the cadherin
family and possesses cell imeraction signaling typical of the cadherin family.
60. Full-length PR0322 Pojvpe~tides
The present invention provides newly identified and isolated nucleotide
sequences encoding polypeptides
referred to in the present application as PR0322. In particular, Applicants
have identified and isolated cDNA
encoding a PR0322 polypeptide, as disclosed in further detail in the Examples
below. Using BLAST and FastA
sequence alignment computer programs, Applicants found that various portions
of the PR0322 polypeptide have
significant homology with human neuropsin, serine protease, neurosin and
trypsinogen. Accordingly, it is presently
believed that PR0322 polypeptide disclosed in the present application is a
newly identified member of the serine
protease family and possesses protease activity typical of this family. It is
also believed that PRO322 is izrvoIved in
hippocampal plasticity and is associated with extracellular matrix
modifications and cell migrations.
61. Full-length PR0526 Pol~peptides
The present invention provides newly identified and isolated nucleotide
sequences encoding polypeptides
referred to in the present application as PR0526. In particular, Applicants
have identified and isolated cDNA
encoding a PR0526 polypeptide, as disclosed in further detail in the Examples
below: Using BLAST and FastA
sequence alignment computer programs, Applicants found that various portions
of the PR0526 polypeptide have
significant homology with the acid labile subunit of the insulin-like growth
factor complex (ALS), as well
carboxypeptidase, SLiT, and platelet glycoprotein V. Accordingly, it is
presently believed that PR0526 polypeptide
disclosed in the present application is a newly identified member of the
leucine-repeat rich superfamily, and possesses
protein-protein interaction capabilities typical of this family.
62. Full-le PR053I Polygeptides
The present invention provides newly identified and isolated nucleotide
sequences encoding polypeptides
referred to in the present application as PR0531. In particular, Applicants
have identified and isolated cDNA
encoding a PR0531 poIypeptide, as disclosed in fiuther detail in the Examples
below. Using BLAST and FastA
sequence alignment computer programs, Applicants found that various portions
of the PR0531 polypeptide have
significant sequence identity and similarity with members of the cadherin
superfamily, particularly, protocadherin
3. Accordingly, it is presently believed that PROS3i polypeptide disclosed in
the present application is a newly
imemiber of the cadherin superfamily, and is a protocadherin. PR0531 is a
transmembrane protein which
has extracelhtlar cadherin motifs. PRO531 is believed to be involved in cell-
cell activity, in particular, cell signaling.
63. ~utl length PRO534 Polvnentides
The present invention provides newly identified and isolated nucleotide
sequences encoding polypeptides
referred to in the present application as PR0534. In particular, Applicants
have identified and isolated cDNA
encoding a PR0534 polypeptide, as disclosed in further detail in the Examples
below. Using BLAST and FastA
sequence alignment computer programs, Applicants found that various portions
of the PR0534 polypeptide have
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significant identity or similarity with the putative disulfide isomerase erp38
precursor and thioredoxin c-3.
Accordingly, it is presently believed that PR0534 polypeptide disclosed in the
present application is a newly identified
member of the disulfide isomerase family and possesses the ability to
recognize and unscramble either intermediate
or incorrect folding patterns typical of this family.
64. F~l-IenQth PR0697 lPo~ypeptides
The present invention provides newly identified and isolated nucleotide
sequences encoding polypeptides
referred to in the present application as PR0697. In particular, Applicants
have identified and isolated cDNA
encoding a PR0697 polypeptide, as disclosed in further detail in the Examples
below. Using BLAST and FastA
sequence alignment computer programs, Applicants found that various portions
of the PR0697 polypeptide have
significant identity or similarity with sFRP-2, sFRP-1 and SARP-1, -2 and -3.
Accordingly, it is presently believed
that PR0697 palypeptide disclosed in the present application is a newly
identified member of the sFRP family and
possesses activity related to the Wnt signal pathway.
65. Full-length PRO?I7 Pol3rpe tides
The present invention provides newly identified and isolated nucleotide
sequences encoding polypeptides
referred to in the present application as PR0717. In particular, Applicants
have identified and isolated cDNA
encoding a PRO?I7 polypeptide, as disclosed in further detail in the Examples
below. To Applicants present
knowledge, the UNQ385 (DNA50988-1326} nucleotide sequence encodes a novel
factor; using BLAST and FastA
sequence alignment computer programs, no significant sequence identities to
airy known human proteins were
revealed.
66. Full-le~,glh ~PR0731 Poly~~Qtides
The present invention provides newly identified and isolated nucleotide
sequences encoding polypeptides
referred to in the present application as PR0731.. 1n particular, Applicants
have identified and isolated cDNA
encoding a PR073I polypeptide, as disclosed in further detail in the Examples
below: Using BLAST and FastA
sequence alignment computer programs, Applicants found that various portions
of the PR073I polypeptide have
significant homology with the protocadherins 4, 68, 43, 42, 3, and 5.
Accordingly, it is presently believed that
PR0731 polypeptide disclosed in the present application is a newly identi$ed
member of the protocadherin family
and possesses cell-cell aggregation or signaling activity or signal
trar>sduction involvement typical of this family.
67. Full-length PR02I8 Pnlvo~ntir~~
The present invention provides newly identified and isolated nucleotide
sequences encoding polypeptides
referred to in the present application as PR0218. In particular, Applicants
have identified and isolated cDNA
encoding a PR02I8 polypeptide, as disclosed in further detail in the Examples
below. The PR02I8-encoding clone
was isolated from a human fetal kidney library. To Applicants present
knowledge, the UNQ192 (DNA30867-1335)
nucleotide sequence encodes a novel factor; using BLAST and FastA sequence
alignment computer programs, no
significant sequence identities to any known proteins were revealed. Some
sequence identity was found with
~mbrane regulator proteins, indicating that PR0218 may function as a membrane
regulator.
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68. Full-lei3gth PR0768 Polypeptides
The present invention provides newly identified and isolated nucleotide
sequences encoding polypeptides
referred to in the present application as PR0768. In particular, Applicants
have identified and isolated cDNA
encoding a PR0768 polypeptide, as disclosed in further detail in the Examples
below. Using BLAST and FastA
seguence alignment computer programs, Applicants found that various portions
of the PR0768 polypeptide have
S significant homology with integrins, including integrin 7 and 6.
Accordingly, it is presently believed that PR0768
polypeptide disclosed in the present application is a newly identified member
of the integrin family, either a
homologue or a splice variant of integrin 7, and is involved with cell
adhesion and communication between muscle
cells and the extraceIlular matrix.
69. Full-leneth PR077I PolYpeptides
The present invention provides newly identified and isolated nucleotide
sequences encoding polypeptides
referred to in the present application as PR077I. In particular, Applicants
have identified and isolated cDNA
encoding a PR0771 polypeptide, as disclosed in further detail in the Examples
below. Using BLAST and FastA
sequence alignment computer programs, Applicants found that various portions
of the PR0771 polypeptide have
significant sequence identity and similarity with testican. Accordingly, it is
pxesently believed that PR0771
polypeptide disclosed in the present application is a newly identified member
of the testican family and possesses cell
signaling, binding, or adhesion properties, typical of this family.
70. Full-length PR0733 Polvoentides
The present invention provides newly identified and isolated nucleotide
sequences encoding polypeptides
referred to in the present application as PR0733. In particular, Applicants
have identified and isolated cDNA
encoding a PR0733 polypeptide, as disclosed in further detail in the Examples
below. Using BLAST and FastA
sequence alignment computer programs, Applicants found that various portions
of the PRO733 polypeptide have
significant sequence identity with the Tl/ST receptor binding protein.
Accordingly, it is presently believed that
PR0733 poIypeptide disclosed in the present application is a newly identified
member of the interleukin-like family
binding proteins which may be a cytokine and which may be involved in cell
signaling. It is believed that PRO733
is an ApoAIV homologue.
71. ~ 1_i lllenE"th,~'R0162 Polppeptides
The present invention provides newly identified and isolated nucleotide
sequences encoding polypeptides
referred to in the present application as PR0162. In particular., Applicants
have identified and isolated cDNA
encoding a PR0162 polypeptide, as disclosed in further detail in the Examples
below. Using BLAST and FastA
sequence alignment computer programs, Applicants found that various portions
of the PR0162 polypeptide have
sigoi$cant homology with human pancreatitis-associated protein (PAP).
Applicants have also found that the DNA
encoding the PR0162 polypeptide has significant homology with bovine
lithosiathine precursor and bovine pancreatic
thread protein (PTP). Accordingly, it is presently believed that PR0162
polypeptide disclosed in the present
application is a newly identified member of the pancreatitis-associated
protein family and possesses activity typical
of the paricreatitis-associated protein family.
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72. dull-lgnpth PR0788 Polypeptides
The present invention provides newly identified and isolated nucleotide
sequences encoding polypeptides
referred to in the present application as PR0788. In particular; Applicants
have identified and isolated cDNA
encoding a PRO788 polypeptide, as disclosed in further detail in the Examples
below. Using BLAST and FastA
sequence alignment computer programs, Applicatus found that various portions
of the PR0788 polypeptide have
significant homology with the anti-neoplastic urinary protein. Applicants have
also found that the DNA encoding
the PR0788 polypeptide has significant homology with human E48 antigen, human
component B protein, and human
prostate stem cell antigen. Accordingly, it is presently believed that PR0788
polypeptide disclosed in the present
application is a newly identified member of the anti-neoplastic urinary
protein family and possesses anti-neoplastic
activity typical of the anti-neoplastic urinary protein family.
73. Full-length PR01008 Polype ides
The present invention provides newly identified and isolated nucleotide
sequences encoding polypeptides
referred to in the present application as PR01008. In particular, Applic,~nts
have identified and isolated cDNA
encoding a PR01008 polypeptide, as disclosed in further detail in the Examples
below. Using BLAST and FastA
sequence alignment computer programs, Applicants found that various portions
of the PR01008 polypeptide have
significant sequence identity and similarity with mouse dkk-1 (mdkk-i).
Accordingly, it is presently believed that
PROI008 polypeptide disclosed in the present application is a newly identified
member of the dlck-1 family and
possesses head inducing activity typical of this family.
74. Full-length PROl0I2 Poiypeptic~es
The present invention provides newly identified and isolated nucleotide
sequences encoding polypeptides
referred to in the present application as PR01012. In particular, Applicants
have identified and isolated cDNA
encoding a PR01012 polypeptide, as disclosed in further detail in the Examples
below. Using BLAST and FastA
sequence alignment computer programs, Applicants found that various portions
of the PROl0i2 polypeptide have
sequence identity with disulfide isomerase. Accordingly, it is presently
believed that PR01012 polypeptide disclosed
in the present application is a newly identified member of the ER retained
protein family and possesses activity related
to the processing, production andior folding of polypeptides typical of the
disulfide isotr~erase family.
75. Full-lepgth P1It01014 Polype,Rti~s
The present invention provides newly identified and isolated nucleotide
sequences encoding polypeptides
referred to in the present application as PR01014. In particular, Applicants
have identified and isolated cDNA
encoding a PR01014 polypeptide, as disclosed in further detail in the Examples
below. Using BLAST and FastA
sequence alignment computer programs, Applicants found that various portions
of the PROi0i4 polypeptide have
sequence identity with reductase and dehydrogenase. Accordingly, it is
presently believed that PR01014 polypeptide
disclosed in the present application is a newly identified member of the
reductase super family and possesses
reduction capabilities typical of this family.
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76. Full-Ienglh PR01017 PoiyPeptides
The present invention provides newly identified and isolated nucleotide
sequences encoding polypeptides
referred to in the present application as PR01017. In particular, Applicants
have identified and isolated cDNA
encoding a PR01017 polypeptide, as disclosed in further detail in the Examples
below. Using BLAST and FastA
sequence alignment computer programs, Applicants found that various portions
of the PR01017 polypeptide have
sequence identity with HNK-I sulfotransferase. Accordingly, it is presently
believed that PR01017 polypeptide
disclosed in the present application is a newly identified ember of the HNK-I
sulfotransferase family and is involved
with the synthesis of HNK-1 carbohydrate epitopes typical of this family.
77. Full-length PR0474 Polypeptides
The present invention provides newly identified and isolated nucleotide
sequences encoding polypeptides
referred to in the present application as PR0474. In particular, Applicants
have identified and isolated cDNA
encoding a PR0474 polypeptide, as disclosed in further detail in the Examples
below. Using BLAST and FastA
sequence aligtunent computer programs, Applicants found that various portions
of the PR0474 polypeptide have
sequence identity with dehydrogenase, glucose dehydrogenase and
oxidoreductase. Accordingly, it is presently
believed that PRO474 polypeptide disclosed in the present application is a
newly identified member of the
dehydrogenase family and is involved in the oxidation of glucose.
78. Full-length PR01031 Polvp~ptides
The present invention provides newly identified and isolated nucleotide
sequences encoding polypeptides
referred to in the present application as PRO1031. In particular, Applicants
have identified and isolated cDNA
encoding a PR01031 polypeptide, as disclosed in further detail in the Examples
below. Using BLAST and FastA
sequence alignment computer programs, Applicants found that various portions
of the PRO1031 polypeptide have
~quenoe identity with IL 17 and CTLA-8. Accordingly, it is presently believed
that PR01031 polypeptide disclosed
in the present application is a newly identified member of the cytoldne family
and thus may be involved in
inflammation and/or the immune system.
79. ~uil-leneth PRO938 Polv~,eotides
The present invention provides newly identified and isolated nucleotide
sequences encoding polypeptides
referred to in the present application as PR0938. In particular; Applicants
have identified and isolated cDNA
encoding a PR0938 polypeptide, as disclosed in further detail in the Examples
below. Using BLAST and FastA
sequence aligz>rnent computer programs, Applicants found that the PR0938
polypeptide has significant similarity to
protein disulfide isomerase. Accordingly, it is presently believed that PR0938
polypeptide disclosed in the present
application is a newly identified member of the thioredoxin family proteins
and possesses activity typical of protein
disulfide isomerase.
gp_ ~ ~11_ip IZ,)iR 10 X82 P_oivneotides
The present invention provides newly identified and isolated nucleotide
sequences encoding polypeptides
referred to in the present application as PR01082. In particular, Applicants
have identified and isolated cDNA
encoding a PR01082 polypeptide, as disclosed in further detail in the Examples
below. Using BLAST and FastA
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sequence alignment computer programs, Applicants found that various portions
of the PR01082 polypeptide have
sequence identity with a lectin-like oxidized LDL receptor appearing in the
database as "AB010710~1 ". Accordingly,
it is presently believed shat PR01082 poiypeptide disclosed in the present
application is a newly identified member
of the LDL receptor family.
81. Full-length PR01083 PoiyRe_ptides
The present invention provides newsy identified and isolated nucleotide
sequences encoding polypeptides
referred to in the present application as PR01083. In particular, Applicants
have identified and isolated cDNA
encoding a PR01083 polypeptide, as disclosed in further detail in the Examples
below. The PROI083-encoding clone
was isolated from a human fetal kidney library using a trapping technique
which selects for nucleotide sequences
encoding secreted proteins. To Applicants present knowledge, the UNQ540
(DNA50921-1458) nucleotide sequence
encodes a novel factor; using BLAST and FastA sequence alignment computer
programs, some sequence identity with
a 7TM receptor; latrophilin related protein l and a macrophage restricted cell
surface glycoprotein was shown. The
kinase phosphorylation site and G-coupled receptor domain shown in Figure 204
indicate that PR01083 is a novel
member of the 7TM receptor superfamily.
82. Full-leng~,h PR0200 Pq,~yoentides
The present invention provides newly identified and isolated nucleotide
sequences encoding polypepiides
referred to in the present application as VEGF-E: In particular; Applicants
have identified and isolated cDNA
encoding a VEGF-E palypepGde, as disclosed in further detail in the Examples
below. Using BLAST sequence
alignment computer programs, Applicants found that the VEGF-E polypeptide has
significant homology with VEGF
arnl bone morghogenetic protein 1. In particular, the cDNA sequence of VEGF-E
exhibits 24~ amino acid similarity
with VEGF and has structural conservation. In addition, VEGF-E contains a N-
terminal half which is not present
in VEGF and that has 28~ homology to bone morphogenetic protein 1.
83. Full-iength~,~0~85 and ~~~,poJ nodes
The present invention provides newly identified and isolated nucleotide
sequences encoding polypeptides
referred to in the present application as PR0285 and PR0286 In particular,
Applicants have identified and isolated
cDNAs encoding PR0285 and. PRO286 poiypeptides, as disclosed in further detail
in the Examples below. Using
BLAST and FastA sequence alignment computer programs, Applicants found that
the coding sequences of PR0285
and PR0286 are highly homologous to DNA sequences HSU88540 I, HSU88878_l,
HSU88879 1, HSU88880 1,
and HSU88881 1 in the GenBank database.
Accordingly, it is presently believed that the PR0285 and PR0286 proteins
disclosed in the present
application are newly identified human homologues of the Drosophila protein
Toll, and are likely to play an important
role in adaptive immunity. More specifically, PR0285 and PR0286 may be
involved in inflammation, septic shock,
and response to pathogens, and play possible roles in diverse medical
conditions that are aggravated by immune
response, such as, for example, diabetes, ALS, cancer, rheumatoid arthritis,
and ulcers. The role of PR0285 and
PR0286 as pathogen pattern recognition receptors, sensing the presence of
conserved molecular structures present
on microbes, is further supported by the data disclosed in the present
application, showing that a known human Toll-
like receptor, TLR2 is a direct mediator of LPS signaling.
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84. Full-length PR0213-1. PR013 0 and PR01449 Polypeptides
The present invention provides newly identified and isolated nucleotide
sequences encoding polypeptides
referred to in the present application as PR0213-I, PR01330 and/or PR01449. In
particular, cDNA encoding a
PR0213-I, PROi330 and/or PR01449 polypeptide has been identified and isolated,
as disclosed in further detail in
the Examples below. It is noted that proteins produced in separate expression
rounds may be given different PRO
numbers but the UNQ number is unique for any given DNA and the encoded
protein, and will not be changed.
However, for sake of simplicity, in the present specification the protein
encoded by DNA30943-1 I63-1, DNA64907-
1163-1 and DNA64908-1163-1 as well as all further native homologues and
variants included in the foregoing
definition of PR0213-1, PR01330 and/or PR01449, will be referred to as "PR0213-
1, PR01330 andlor PR01449",
regardless of their origin or mode of preparation.
85. Full-length PR0298 Poivventides
The present invention provides newly identified and isolated nucleotide
sequences encoding polypeptides
referred to in the present application as FR0298. (It is noted chat PR0298 is
an arbitrary designation of a protein
encoded by the nucleic acid shown in Figure 218, SEQ ID NO: 514, and having
the amino acid sequence shown in
Figure 219, SEQ ID NO:515. Further proteins having the same amino acid
sequence but expressed in different
rounds of expression, may be given different "PRO" numbers.)
In particular, Applicants have identified and isolated cDNA encoding a PRQ298
polypeptide, as disclosed
in further detail in the Examples below. Using BLASTX 2.Oa8MP-WashU computer
program, socring parameters:
T=12, S=68, S2=36, Matrix: BLOSUM62, Applicants found that the PR0298 protein
specifically disclosed herein
shows a limited {27-38%) sequence identity with the following sequences found
in the GenBank database: S59392
(probable membrane protein YLR246w - yeast); 558154 (hypothetical protein
SPAC2F7.10 - yeast); CELF33D11 9
(F33D11.9b _- Caenorhabditis elegans); Y041 CAEEL (hypothetical 68.7 kd
protein zk75?.1); CEAC3 5 (AC3.4
Caenorhabditis elegans); 552691 (probable transmembrane protein YDR126w -
yeast); ATT12H17_14 (protein
Arabidopsis _thaliana); S55963 (probable membrane protein YNL326c - yeast);
CELC43H6 2 (C43H6.7
Caenorhabditis elegans); TM018A10 14 (A TM018A10.8 - Arabinosa thaliana).
86. Full leneth PR0337 Polvneotides
The present invention provides newly identified and isolated nucleotide
sequences encoding polypeptides
referred to in the present application as PR033?. In particular, Applicants
have identified and isolated cDNA
encoding a PR0337 polypeptide, as disclosed in further detail in the Examples
below. Using BLAST, BLAST-2 and
FastA sequence alignment computer programs, Applicants found that a full-
length native sequence PR0337 has 97 %
amino acid sequerxe identity with rat neurotrimin, 85% sequence identity with
chicken CEPU, 7386 sequence identity
with chicken G55, 59°k homology with human LAMP and 84% homology with
human OPCAM. Accordingly, it
is presently believed that PR0337 disclosed in the present application is a
newly identified member of the IgLON
sub family of the immunoglobulin superfamily and may possess neurite growth
and differentiation potentiating
properties.
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87. Full-IenEth P1,~0403 Polvoeotldes
The present invention provides newly identified and isolated nucleotide
sequences encoding polypeptides
referred to in the present application as PR0403. In particular, Applicants
have identified and isolated cDNA
encoding a PR0403 polypeptide, as disclosed in further detail in the Examples
below. Using a BLAST, BLAST-2
and FastA sequence alignment computer programs, Applicants found that a full-
length native sequence PR0403 has
94% identity to bovine ECE-2 and 64% identity to human ECE-1. Accordingly is
presently believed that PR0403
is a new member of the ECE protein family and may posses ability to catalyze
the production of active endothelin.
88. PRO Poiypgptide Variants
In addition to the full-length native sequence PRO polypep;ides described
herein, it is contemplated that PRO
polypeptide variants can be prepared. PRO polypeptide variants can be prepared
by introducing appropriate
nucleotide changes into the PRO polypeptide DNA, or by synthesis of the
desired PRO polypeptide. Those skilled
in the art will appreciate that amino acid changes may alter post
translational processes of the PRO polypeptides, such
as changing the number or position of glycosylation sites or altering the
membrane anchoring characteristics.
Variations in the native full-length sequence PRO polypeptides or in various
domains of the PRO
polypeptides described herein; can be made, for example, using any of the
techniques and guidelines for conservative
and non-conservative mutations set forth, for instance, in U,S. Patent No.
5,364,934. Variations may be a
substitution, deletion or insertion of one or more codons encoding the PltO
polypeptide that results in a change in
the amino acid sequence of the PRO polypeptide as compared with the native
sequence PRO polypeptide. Optionally
the variation is by substitution of at least one amino acid with arty other
amino acid in one or more of the domains
of the PRO polypeptide. Guidance in determining which amino acid residue may
be inserted, substituted or deleted
without adversely aff~ting the desired activity may be found by comparing 'the
sequence of the PRO poIypeptide with
that of homologous known protein molecules and minimizing the number of amino
acid sequence changes made in
regions of high homology. Amina acid substitutions can be the result of
replacing one amino acid with another amino
acid having similar sttnctural andlor chemical properties; such as the
replacement of a leucine with a serine, i.e.,
conservative amino acid replacements. Insertions or deletions may optionally
be in the range of 1 to 5 amino acids.
The variation allowed may be determined by systematically making insertions,
deletions or substitutions of amino
acids in the sequence and testing the resulting variants for activity in the
in vitro assay described in the Examples
below.
In particular embodiments, conservative substitutions of interest are shown in
Table I under the heading of
preferred substitutions. If such substitutions result in a change in
biological activity, then more substantial changes,
denominated exemplary substitutions in Table I , or as further described below
in reference to amino acid classes,
are introduced and the products screed.
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WO 99146281 PCT/US99/05028
Tab a 1
Original Exemplary Preferred
Res' a Substitutions substitutions
S Ala (A) vai; leu; ile val
Arg (R) lys; gln; asn lys
Asn (N) gln; his; lys; arg gln
Asp (D) glu gIu
Cys (C) ser ser
10Gln (Q) asn asn
Glu {E) asp asp
Gly (G) pro; ala ala
His (H) asn; gln; lys; arg arg
De (1] leu; val; met; ala; phe;
15 norleucine leu
Leu (L) norleucine; ile; val;
met; ala; phe ile
Lys (K) arg; gln; asn arg
l~let (M) leu; phe; ile leu
20Phe (F) leu; val; ile; ala; tyr leu
Pro (P) ala ala
Ser (S)
thr thr
Thr (T) ser ser
Trp t;W) tyr; phe tyr
25Tyr (1~ trp; phe; thr; ser phe
Va1 (V) ile; leu; met; phe;
aIa; norleucine leu
Substantial modifications in function or immunoIogical identity of the PRO
polypeptide are accomplished
30 by selecting substitutions that differ significantly in their effect on
maintaining (a) the structure of the polypeptide
backbone in the area of the substitution, for example, as a sheet or helical
conformation, (b) the charge or
hydrophobicity of the molecule at the target site, or (c) the bulk of the side
chain. Naturally occurring residues are
divided into groups based on common side-chain properties:
(1) hydrophobic: norleucine, met, ala, vai, leu, ile;
35 (2) neutral hydrophilic: cys, ser, thr;
(3) acidic: asp, glu;
(4) basic: asn, gin, his, lys, arg;
(5) residues that influence chain orientation: gly, pro; and
(6) aromatic: trp, tyr, phe.
40 Non-conservative substitutions will entail exchanging a member of one of
these classes for another class.
Such substituted residues also tray be introduced into the conservative
substitution sites or, more preferably, into the
remaining (non-conserved) sites.
The variations can be made using methods known in the an such as
oligonucleotide-mediated (site-directed)
mutagenesis, alanine scanning, and PCR mutagenesis. Site-directed mutagenesis
[Carter et al., Nucl. Acids Res.;
45 x:4331 (i98~; Zoller et al., Nucl. Acids Res., ~Q:6487 (1987)], cassette
mutagenesis [Wells et al., Gene, x:315
(1985)], restriction selection mutagenesis [Wells et al., Plilos. Trans. R.
Soc. London SerA, x:415 (1986)) or other
known techniques can be performed on the cloned DNA to produce the desired PRO
polypeptide variant DNA.
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Scanning amino acid analysis can also be employed to identify one or more
amino acids along a contiguous
sequence. Among the preferred scanning amino acids are relatively small,
neuual amino acids. Such amino acids
include alanine, glycine, serine, and cysteine. Alanine is typically a
preferred scanning amino acid among this group
because it eliminates the side-chain beyond the beta-carbon and is less likely
to alter the main-chain conformation of
the variant. Alanitte is also typically preferred because it is the most
common amino acid. Further, it is frequently
found in both buried and exposed positions [Creighton, 'the Protgins, (W.H.
Freeman & Co., N.Y.); Chothia, J.
of B'o , 250:1 (1976)]. If alanine substitution does not yield adequate
amounts of variant, an isoteric amino acid
can be used.
89. Modif~,ations of ]:'RO~o"~y~e tn id_es_
Covalent modifications of PRO polypeptides are included within the scope of
this invention. One type of
covalent modification includes reacting targeted amino acid residues of the
PRO polypeptide with an organic
derivatiz5ng agent that is capable of reacting with selected side chains or
the N- or C- terminal residues of the PRO
polypeptide. Derivatization with bifunctional agents is useful, for instance,
for crosslinlting a PRO polypeptide to
a vrater-insoluble support matrix or surface for use in the method for
purifying anti-PRO polypeptide antibodies, and
vice versa. Commonly used crosslinking agents include, e.g. , 1,1-
bis(diaroaceryl)-2-phenylechane, glutaraldehyde,
N-hydroxysuccinurude esters, far example, esters with 4-azidosalicylic acid,
homobifunctional imidoesters, including
disuccinimidyl esters such as 3,3'-dithiobis(succinimidylpropionate),
bifunctional maleimides such as bis-N-
maleimido-1,8-octane and agents such as methyl-3-[(p-
azidophenyl)dithio]propioimidate.
Other modifications include deamidation of glutaminyI and asparaginyl residues
to the corresponding
glutamyl and aspartyl residues, respectively, hydroxylation of proline and
lysine, phosphorylation of hydroxyl gmups
of seryl or threonyl residues, methylation of the a-amino groups of lysine,
arginine, and histidine side chains j3'.E.
Creightoa, Proteins' Strucnire and Molecular Propertiet, W.H. Freeman & Co.,
San Francisco, pp. 79-86 (1983)],
acetylation of the N-terminal amine, and amidation of any C-terminal carboxyl
group.
Another type of covalent modification of the PRO polypeptides included within
the scope of this invention
comprises altering the native gIycosyIation pattern of the polypeptide.
"A,ltering the native glycosylation pattern" is
intended for purposes herein to mean deleting one or more carbohydrate
moieties found in a native sequence PRO
polypeptide, and/or adding one or more glycosylation sites that are not
present in the native sequence PRO
polypeptide, and/or alteration of the ratio andlor composition of the sugar
residues attached to the glycosylation
site(s).
Addition of glycosylation sites to the PRO polypeptide may be accomplished by
altering the amino acid
sequence. The alteration may be made, for example, by the addition of; or
substitution by; one or more serine or
threottine residues to the r~dve sequence PRO polypeptide (for O-linked
glycosylation sites). The PRO polypeptide
amino acid sequence may optionally be altered through changes at the DNA
level, particularly by mutating the DNA
eroding the PRO polypeptide at preselected bases such that codons are
generated that will translate into the desired
amino acids.
Another means of increasing the number of carbohydrate moieties on the PRO
polypeptide polypeptide is
by chemical or enzymatic coupling of glycosides to the polypeptide. Such
methods are described in the art, e.g., in
WO 87/45330 published 11 September 1987, and in Aplin and Wriston, CRC Crit.
Rev. Biochem., pp. 259-306
(1981).
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Removal of carbohydrate moieties present on the PRO polypeptide may be
accomplished chemically or
enzymatically or by mutational substitution of codons encoding for amino acid
residues that serve as targets for
glycosylation. Chemical deglycosylation techniques are known in the art and
described, for instance, by Hakimuddin,
et al., Arch. Biochem. Bionhvs., 25:52 (1987) and by Edge et al., Anal.
Biochem., ,[ 18:131 (198I). Enzymatic
cleavage of carbohydrate moieties on poiypeptides can be achieved by the use
of a variety of endo- and exo-
glycosidases as described by Thotakura et al., Meth. Enz~ol., L8_:350 (1987).
Another type of covalent modification of PRO polypeptides of the invention
comprises linking the PRO
polypeptide to one of a variety of nonproteinaceous polymers, e.g.,
polyethylene glycol, polypropylene glycol, or
polyoxyalkylenes, in the manner set forth in U.S. Patent Nos. 4,640,835;
4,496,689; 4,301,144; 4,670,417;
4,791,192 or 4,179,337.
The PRO polypeptides of the present invention may also be modified in a way to
form a chitneric molecule
comprising a PRO polypeptide fused to another, heterologous polypeptide or
amino acid sequence. In one
embodiment, such a chimeric molecule comprises a fusion of the PRO poiypeptide
with a tag polypeptide which
provides an epitope to which an anti-tag antibody can selectively bind. The
epitope tag is generally placed at the
amino- or carboxyl- terminus of the PRO polypeptide. The presence of such.
epitope-tagged forms of the PRO
polypegtide can be detected using an antibody against the tag polypeptide:
Also, provision of the epitope tag enables
the PRO polypeptide to be readily purified by affinity purification using an
anti-tag antibody or another type of affinity
matrix that binds to the epitope tag. In an alternative embodiment, the
chimeric molecule may comprise a fusion of
the PRO polypeptide with an immunoglobulin or a particular region of an
immunoglobulin. For a bivalent form of
the chitneric molecule, such a fusion could be to the Fc region of an IgG
molecule.
Various tag polypeptides and their respective antibodies are well iatown in
the art. Examples include poly-
histidine (poly-his) or poly-histidine-glycine (poly-his-gly) tags; the flu HA
tag polypeptide and its antibody 12CA5
[Field et al., Mol. Cell. Biol., $:2159-2165 (1988)]; the c-myc tag and the
8F9, 3C7, 6E10, G4, B7 and 9E10
antibodies thereto [Evan et al., Iy olecula~ and Cellular Biology, x:3610-3b16
(1985)]; and the Herpes Simplex virus
glycoprotein D (gD) tag and its antibody [Paborsky et al., Protein
Engineering, 3_(6):547-553 (1990)]. Other tag
polypeptides include the Flag-peptide [Hopp et al., BioTechnoloQV, _6:1204-
1210 (1988)]; the KT3 epitope peptide
[Martin et al" Science, X55:192-194 (1992)]: an a tubulin epitope peptide
[Skinner et al., J. Biol. Chem., 2~6:15163-
15166 (1991)]; and the T7 gene 10 protein peptide tag [Iutz-Freyermuth et al.,
Proc. Natl Acad.Sci.USA, 87:6393-
6397 (1990)].
90. P~paration of PRO Polypentides
The description below relates primarily to production of PRO polypeptides by
culturing cells transformed
or transfected with a vector containing the desired PRO polypeptide nucleic
acid. It is, of course, contemplated that
alternative methods, which are well known in the art, may be employed to
prepare the PRO polypeptide. For
instance, the PRO poIypeptide sequence, or portions thereof, may be produced
by direct peptide synthesis using solid-
phase techniques [see, e.g., Stewart et aL, Solid-Phase Peptide Synthesis,
W.H. Fzeeman Co., San Francisco, CA
(1969); Merrifield, ~ Am. Chem. Soc., $5_:2149-2134 (1963)]. In vitro protein
synthesis may be performed using
manual techniques or by automation. Automated synthesis may be accomplished,
for instance, using an Applied
Biosystems Peptide Synthesizer (Foster City, CA) using manufacturer's
instructions. Various portions of the desired
PRO polypeptide may be chemically synthesized separately and combined using
chemical or enzymatic methods to
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produce the full-length PRO polypeptide.
A. Isolation of DATA, Encoding_PRO Poly~e~ides
DNA encoding PRO polypeptides may be obtained from a cDNA library prepared
from tissue believed to
possess the desired PRO polypeptide mRNA and to express it at a detectable
level. Accordingly, human PRO
polypeptide DNA can be conveniently obtained from a cDNA library prepared from
human tissue, such as described
in the Examples. The PRO poiypeptide-encoding gene may also be obtained from a
genotnic library or by
oligonucleotide synthesis.
Libraries can be screened with probes (such as antibodies to the desired PRO
polypeptide or oligonucleotides
of at least about 20-80 bases) designed to identify the gene of interest or
the protein encoded by it. Screening the
cDNA or genomic library with the selected probe may be conducted using
standard procedures, such as described
in Sambrook et al., Mole~ul3r ~,loning.: A I,aborator~Njgnual (New York: Cold
Spring Harbor Laboratory Press,
1989). An alternative means to isolate the gene encoding the desired PRO
polypeptide is to use PCR methodology
[Sambrook et al., supra; Dieffenbach et al., PGR Primer:A I,a~oratorv Manual
(Cold Spring Harbor Laboratory
Press, 1995)).
The Examples below describe techniques for screening a cDN.A library. The
oligonucleotide sequences
selected as probes should be of sufficient length and suiBciently unambiguous
that false positives are minimized. The
oligonucleotide is preferably labeled such that it can be detected upon
hybridization to DNA in the library being
screened. Methods of labeling are well known in the art, and include the use
of radiolabels like ~P-labeled ATP,
biotitrylation or enzyme labeling. Hybridization conditions, including
moderate stringency and high stringe~y, are
provided in Sambrook et al., supra.
Sequences identified in such library screening methods can be compared and
aligned to other known
sequences deposited and available in public databases such as GenBank or other
private sequence databases.
Sequence identity (at either the amino acid or nucleotide level) within
defined regions of the molecule or across the
full-length sequence can be determined through sequence alignment using
computer software programs such as
BLAST, ALIGN; DNAstar, and INHERIT which employ various algorithms to measure
homology.
Nucleic acid having protein coding sequence may be obtained by screening
selected cDNA or genomic
libraries using the deduced amino acid sequence disclosed herein for the first
time, and, if necessary; using
conventional primer extension procedures as descn'bed in Sambrook et al., a r
, to detect precursors and processing
intermediates of mRNA that may not have been reverse-transcribed into cDNA.
B. Sglg~ion and nsfor~~tion of Host Cells
Host cells are transfected or transformed with expression or cloning vectors
descn'bed herein for PRO
polypeptide production and cultured in conventional nutrient media modified as
appropriate for inducing promoters,
selecting uansformatus, or amplifying the genes encoding the desired
sequences. The culture conditions, such as
media, temperature, pH and the like, can be selected by the skilled artisan
without endue experimentation. In
general, principles, protocols, and practical techniques for maximizing the
productivity of cell cultures can be found
in Mammali~gCel~Biotechnology a Practical Ap rp oach, M. Butler, ed. (/R1,
Press, 1991) and Satnbrook et al:,
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WO 99!46281 PCTlUS99l05028
Methods of transfection are known to the ordinarily sidlled artisan, for
example, CaPO, and electroporation.
Depending on the host cell used, transforntation is performed using standard
techniques appropriate to such cells.
The calcium treatment employing calcium chloride, as described in Sambrook et
al., suvra, or electroparation is
generally used for prokaryotes or other cells that contain substantial cell-
waft barriers. Infection with Agrobacterium
tumefaciens is used for transformation of certain plant cells, as described by
Shaw et al., Gene, 2_x:315 (I983) and
WO 89105859 published 29 June 1989. For martunalian cells without such cell
walls, the calcium phosphate
precipitation method of Graham and van der Eb, ViroloQV, X2_:456-457 (1978)
can be employed. General aspects
of mammalian cell host system transformations have been described in U.S.
Patent No. 4,399,216. Transformations
into yeast are typically carried out according to the method of Van Solingen
et al., 3.- Bact., 130:946 (1977) and Hsiao
et al:, Proc. Nat!. Acad. Sci. (USA), 76:3829 (1979). However, other methods
for introducing DNA into cells, such
as by nuclear microinjection, electroporation, bacterial protoplast fusion
with intact cells, or polycations, e.g. ,
polybrene, polyornithine, may also be used. For various techniques for
transfonttirtg mammalian cells, see Keown
et al., Methods,~n Enzy~ology, 185:527-537 (1990) and Mansour et al., Nature,
336:348-352 (1988).
Suitable host cells for cloning or expressing the DNA in the vectors herein
include prokaryote, yeast, or
higher eukaryote cells. Suitable prokaryotes include but are not limited to
eubacteria, such as Gram-negative or
Gram-positive organisms, for example, Enterobacteriaceae such as E. coti.
Various E. coli strains are publicly
available, such as E. coli,Kl2 swain MM294 (ATCC 31,446); E. coti X1776 (ATCC
31,537); E. toll strain W31I0
(ATCC 27,325) and KS 772 (ATCC 53,635): Other suitable prokaryotic host cells
include Enterobacteriaceae such
as Escherichia, e.g., E. toll, Enterobaaer, Erwinia, Klebsietla, Proteus,
Salmonella, e.g., Salmonella typhimurium,
Serraria, e.g., Serratia marcescans, and Shigetia, as well as Bacilli such as
B. subtilis and B. licheniformis (e.g., B.
licheniformis 41P disclosed in DD 266,710 published I2 April 1989),
Pseudomonas such as P. aeruginasa, and
Streptomyces. Various E. toll strains are publicly available, such as E. toll
KI2 strain MM294 (ATCC 31,446); E.
toll XI776 (ATCC 31,537); E. toll strain W3110 (ATCC 27,325); and KS ?72 (ATCC
53,635). These examples
are illustrative rather than limiting. Strain W3110 is one particularly
preferred host or parent host because it is a
common host strain for recombinant DNA product fermentations. Preferably, the
host cell secretes minimal amounts
of proteolytic enzymes. For example, strain W3110 may be modified to effect a
genetic mutation in the genes
ettcoditlg proteins endogenous to the host, with examples of such hosts
including E. toll W3110 strain 1A2, which
has the complete genotype tonA ; E. toll W3110 swain 9E4, which has the
complete genotype tonA ptr3; E. toll
W3110 strain 27C7 (ATCC 55,244), which has the complete genotype tonA ptr3
phoA E15 (argF tac)169 degP
ompT kan ; E. toll W3110 strain 37D6, which has the complete genotype tonA
ptr3 phoA EIS (argF tac)169 degP
ompT rbs7 itvG kan'; E. toll W3110 sttaiw40B4, which is strain 37D6 with a non-
kattamycin resistant degP deletion
mutation; and an E. toll strain having mutant periplasmic protease disclosed
in U.S. Patent No. 4,946,783 issued 7
August 1990. Alternatively, in vitro methods of cloning, e.g., PCR or other
nucleic acid polymerase reactions, are
suitable.
In addition to prokaryotes, eukaryotic nvcrobes such as filamemous fungi or
yeast are suitable cloning or
expression hosts for PRO polypeptide-encoding vectors: Saccharomyces
cerevisiae is a commonly used lower
eukaryotic host microorganism. Others include Schizosaccharomyces pombe (Beach
and Nurse, Nature-, ~: 140
[1981); EP 139,383 published 2 May 1985); Kluyveromyces bolts (U.S. Patent No.
4,943,529; Fleer et at.,
BiofTechnology, Q: 968-975 (1991)) such as, e.g., K. tacos (MW98-8C, CBS683;
CBS45?4; Louvencourt et at., J
B~teriol., 737 [1983]), K.,fragitis (ATCC 12,424), K, bulgaricus (ATCC
16,045), K. wickeramii (ATCC 24,178),
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K waltii (ATCC 56,500); K. drosophilarum (ATCC 36,906; Van den Berg et u1.,
BioITechnoioev, $: 135 (1990)),
K. thennotolerans, and K. marxianus; ynrrowia (FP 402,226); Piehia paswris (EP
183,070; Sreekrishna et al., ~.
Basic 1 icrobiol., ~: 26S-278 [1988)); Candida; Trichoderma reesia (EP
244,234); Neurospora crassa (Case et al.,
Proc. Natl. Acad. Sci. USA, ?6: 5259-5263 [1979]); Schwanniomyces such as
Schwanniomyces occidentalis (EP
394,538 published 31 October 1990); and filamentous fungi such as, e.g.,
Neurospora, Penicillium, Tolypocladium
(WO 91100357 published i0 January 1991), and Aspergillus hosts such as A.
nidulans fBallance et al., Bioc a
Biophys. Res. Colruxn~~, 112: 284-289 [1983]; Tiiburn et al., a e, 26: 205-221
[1983]; Yelton et al., Prod Natl.
Aca~, Sei. USA, 81: 1470-1474 [1984]) and A. niger (Kelly and Hynes, EIvJ$O
J., 4_: 475-479 [1985]).
Methylotropic yeasts are suitable herein and include, but are not limited to,
yeast capable of growth on methanol
selected from the genera consisting of Hansenula, Candida, Kloeckera, Pichia,
Saccharomyces, Torulopsis, and
Rhodotorula. A list of specific species that are exemplary of this class of
yeasts may be found in C. Anthony, ie
Biochemistry of Mgt3ly otronhs, 269 (I982).
Suitable host cells for the expression of glycosylated PRO polypeptides are
derived from tnulticelluiar
organisms. Examples of invertebrate cells include insect cells such as
Drosophila S2 and Spodoptera Sf9, as well.
as plant cells. Exazrtples of useful mammalian host cell lines include Chinese
hamster ovary (CHO) and COS cells.
More specific examples itxlude monkey kidney CV1 line transformed by SV40 (COS-
7, ATCC CRL 1651); human
embryonic kidney line (293 or 293 cells subcloned for growth in suspension
culture, Graham et al., J. Gen Virol.,
$6_:59 (19'17)); Chinese hamster ovary cells!-DHFR (CHO, Urlaub and Chasin,
Proc. Natl. Acad. Sci. USA, ?7:4216
(1980)); fuse sertoli cells fTM4, Mather, Bioj~R, X3_:243 251 (1980)); human
lung cells (W138, ATCC CCL
75); human liver cells (Hep G2, HB 8065); and mouse mammary tumor (MMT 060562,
ATCC CCL51). The
selection of the appropriate host cell is deemed to be within the skill in the
art.
C. Selection and Use of a Realicab a Vector
The nucleic acid (e.g., cDNA or genomic DNA) encoding a desired PRO
polypeptide may be inserted into
a repHcable vector for cloning (amplification of the DNA) or for expression.
Various vectors are publicly available.
The vector may, for example, be in the form of a plasmid, cosmid, viral
particle, or phage. The appropriate nucleic
acid sequence may be izLCerted into the vector by a variety of procedures. In
general, DNA is inserted into an
appropriate restriction endonuclease sice(s) using techniques known in the
art. Vector components generally include,
but are not limited to, one or more of a signal sequence, an origin of
replication, one or more marker genes, an
enharxrer element, a promoter, and a transcription termination sequence.
Construction of suitable vectors containing
one or more of these components employs standard ligation techniques which are
latown to the skilled artisan.
The PRO polypeptide of interest may be produced recombinantly not
only~directly, but also as a fusion
polypeptide with a heterologous polypeptide, which may be a signal sequence or
other polypegtide having a specific
cleavage site at the N-terminus of the mature protein or polypeptide. In
general, the signal sequence may be a
carnponettt of the vector, or it may be a part of the PRO polypeptide DNA chat
is inserted into the vector. The signal
sequence may be a prokaryotic signal sequence selected, for example, fi om the
group of the alkaline phosphatase,
penicillittase, ipp, or heat-stable enterotoxin Il leaders. For yeast
secretian the signal sequence may be, e.g., the
yeast invenase leader, aipha factor leader (including Saccharomyces and
Kluyveromyces a-factor leaders, the latter
described in U.S. Patent No. 5,010,182), or acid phosphatase leader, the C.
albicans glucoamylase leader (EP
362,179 published 4 April 1990), or the signal described in WO 901'13646
published 15 November 1990. In
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CA 02421319 2003-03-25
WO 99!46281 PCT/US99105028
ntarttrttaIian cell expression, mammalian signal sequences may be used to
direct secretion of the protein, such as signal
sequences from secreted polypeptides of the same or related species, as well
as viral secretory leaders.
Both expression and cloning vectors contain a nucleic acid sequence that
enables the vector to replicate in
one or more selected host cells. Such sequences are well known for a variety
of bacteria, yeast, and viruses. The
origin of replication from the plasmid pBR322 is suitable for most Gram-
negative bacteria, the 2~c plasmid origin is
suitable for yeast, and various viral origins (SV40, poIyoma, adenovirus, VSV
or BPV) are useful for cloning vectors
in matttrrtalian cells.
Expression and cloning vectors will typically contain a selection gene, also
termed a selectable marker.
Typical selection genes encode proteins that (a) confer resistance to
antibiotics or other toxins, e.g" ampicillin,
neomycin, methotrexate, or tetracycline> (b) complement auxotrophic
deficiencies, or (c) supply critical nutrients not
available from complex media, e.g., the gene encoding D-alutine racemase for
Bacilli.
An example of suitable selectable markers for mammalian cells are those that
enable the identification of
cells competent to take up the PRO polypeptide nucleic acid, such as DHFR or
thymidine kinase. An appropriate
host cell when wild-type DHFR is employed is the CHO cell line deficient in
DHFR activity, prepared and
propagated as described by Urlaub et al., Proc. Nat!. Acid. Sci. USA, 77:4216
.(1980). A suitable selection gene
for use in yeast is the trill gene present in the yeast plasmid YRp7
[Stinchcomb et al., attire, 282:39 (1979);
Kingsman et al., C~eng, 7:141 (1979); Tschemper et al., Gene, 10:157 (1980)].
The trill gene provides a selection
marker for a mutant strain of yeast lacking the ability to grow in tryptophan,
for example, ATCC No. 44076 or PEP4-
1 [Jones, Genetics, $5:12 (1977)].
Expression and cloning vectors usually contain a promoter operably linked to
the PRO polypeptide nucleic
acid sequence to direct mRNA synthesis. Promoters recognized by a variety of
potential host cells are well known.
Promoters suitable for use with prokaryotic hosts include the (3-lactamase and
lactose promoter systems [Chang et
al., atur , 275:615 (1978}; Goeddel et ii., N- attire, 281:544 (1979)],
alkaline phosphatase, a tryptophan (trp)
promoter system [Goeddel, ,~l.~cke'c Acids Res., 8_:4057 (1980); EP 36,776],
and hybrid promoters such as the tic
promoter [deBoer et al., ~~ Nart. Acid. Sci. USA, 8Q:21-25 (1983)]. Promoters
for use in bacterial systems also
will contain a Shine-Dalgatno (S.D.) sequence operably linked to the DNA
encoding the desired PRO polypeptide.
Examples of suitable promoting sequences for use with yeast hosts include the
promoters for 3-
phosphoglycerate lactase [Hitzeman et al., J. Biol. Chem., X5.5:2073 (I980)]
or other gkycolytic enzymes [Hess et al.,
y, Advj~e Reg., x:149. (1968); Holland, Biochemistry, 17:4900 (1978)], such as
enolase, glyceraldehyde-3-
phosphate dehydrogenase, hexokinase, pyruvate decarboxylase,
phosphofructokinase, glucose-6-phosphate isomerase,
3-phosphoglycerate mutase, pyruvate kinase, trioseghosphate isomerase,
phosphoglucose isomerase, and glucokinase.
Other yeast promoters, which are inducible promoters having the additional
advantage of transcription
controlled by growth conditions, are the promoter regions for alcohol
dehydrogenase 2, isocytochrome C, acid
phosphatase, degradative enzymes associated with nitrogen metabolism,
metallothionein, glyceraldehyde-3-phosphate
dehydrogenase, and enzymes responsible for maltose and galactose utilization.
Suitable vectors and promoters for
use in yeast expression are further described in EP 73,657.
PRO polypeptide transcription from vectors in mammalian host cells is
controkked, for example, by
promoters obtained from the genomes of viruses such as polyoma virus, fowlpox
virus (UK 2,211,504 published 5
July 1989), adenovizus (such as Adenovirus 2), bovine papilloma virus, avian
sarcoma virus, cytomegalovirus, a
retrovirus, hepatitis-B virus and Simian Virus 40 (SV40), from heterologous
mammalian promoters, e.g., the actin
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WO 99146281 PCT/US99/05028
promoter or an immunoglobulirt promoter, and from heat-shock promoters,
provided such promoters are compatible
with the host cell systems.
Transcription of a DNA encoding the desired PRO polypeptide by higher
eukaryotes may be increased by
inserting an enhancer sequence into the vector. $nhancers are cis-acting
elements of DNA, usually about from 10
to 300 bp, that act on a promoter to increase its transcription. Many enhaneer
sequences are now known from
mammalian genes (globin, elastase, albumin, a-fetoprotein, and insulin).
Typically, however, one will use an
enhancer from a eukaryotic cell virus. Examples include the SV40 enhancer on
the late side of the replication origin
(bp 100-270), the cytomegalovirus early promoter enhancer, the polyoma
enhancer on the late side of the replication
origin, and adenovirus enhancers. The enhancer may be spliced into the vector
at a position S' or 3' to the PRO
polypeptide coding sequence, but is preferably located at a site 5' from the
promoter.
Expression vectors used in eukaryotic host cells (yeast, fungi, insect, plant,
animal, human, or nucleated
cells from other multiceliular organisms) will also contain sequences
necessary for the termination of transcription
and for stabilizing the mRNA. Such sequences are commonly available from the
5' and, occasionally 3', untranslated
regions of eukaryotic or viral DNAs or cDNAs. These regions contain nucleotide
segments transcribed as
polyadenylated fragments in the tutuanslated portion of the mRNA encoding PRO
polypeptides.
Still other methods, vectors, and host cells suitable for adaptation to the
synthesis of PRO polypeptides in
recombinant vertebrate cell culture are described in Gething et al., stance,
23_:620-625 (1981); Mantel et al.,
.Nab er , ~$~,:40-46 (1979); EP 117;060; and EP 117,U58.
D, Detecting Gene Am~liflcafionlhxnressior~
Gene amplification and/or expression may be measured in a sample directly, for
example, by conventional
Southern blotting, Northern blotting to quantitate the transcription of mRNA
[Thomas, Psoc Nat1 Acad Sue, USA,
77:5201-5205 (1980)), dot blotting (DNA analysis), or in situ hybridization,
using an appropriately labeled pmbe,
based on the sequetxes provided herein. Alternatively, antibodies may be
employed that can recognize specific
duplexes, including DNA duplexes, RNA duplexes, and DNA-RNA hybrid duplexes or
DNA-protein duplexes. The
antibodies in turn may be labeled and the assay may be carried out where the
duplex is bound to a surface, so that
upon the formation of duplex on the surface; the presence of antibody bound to
the duplex can be detected.
Gene expression, alternatively, may be measured by immunologit:al methods,
such as imtnunohistochemica!
staining of cells or tissue sections and assay of cetL culture or body fluids,
to quantitate directly the expression of gene
Product. Antibodies useful for inmtunohistocheznical staining andlor assay of
sample fluids may .be either monoclonal
or polyclonal, and may be prepared in any mammal: Conveniently, the antibodies
may be prepared against a native
sequence PRO potypeptide or against a syrnhetie peptide based on the DNA
sequences provided herein or against
exogenous sequence fused to a PRO poiypeptide DNA and encoding a specific
antibody epitope.
E. ~~a~f(_catioe g~,Polyp~ tide
Fomts of PRO polypeptides mayy be recovered from culture trtedium or from host
cell lysates. If membrane-
bound, it can be released from the membrane using a suitable detergent
solution (e.g. Triton-X 100) or by enzymatic
cleavage. Cells employed in expression of PRO polypeptides can be disrupted by
various physical or chemical
means, such as freeze-thaw cycling, sonication, mechanical disruption, or cell
lysing agents,
It may be desired to purify PRO polypeptides from recombinant cell proteins or
polypeptides. The following
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WO 99/46281 PCT/US99I05028
procedures are exemplary of suitable purification procedures: by fractionation
on an ion-exchange column; ethanol
precipitation; reverse phase HPLC; chromatography on silica or on a ration-
exchange resin such as DEAF; .
chromatofocusing; SDS-PAGE; ammonium sulfate precipitation; gel filtration
using, for example, Sephadex G-75;
protein A Sepharose columns to remove contaminants such as IgG; and metal
chelating columns to bind epitope-
tagged forms of the PRO polypeptide. Various methods of protein purification
may be employed and such methods
are known in the art and described for example in Deutscher, Methods in
Enzvmologx, 1 2 (1990); Scopes, Protein
Purification: Princitxles and Practice, Springer-Verlag, New York (1982). The
purification steps) selected will
depend, for example, on the nature of the production process used and the
particular PRO polypeptide produced.
91. Uses for PRO Polypeptides
Nucleotide sequences (or their complement) encoding the PRO polypeptides of
the present invention have
various applications in the art of molecular biology, including uses as
hybridization probes, in chromosome and gene
mapping and in the generation of anti-sense RNA and DNA. PRO polypeptide-
encoding nucleic acid will also be
useful for the preparation of PRO polypeptides by the recombinant techniques
described herein.
The full-length native sequence PRO polypeptide-encoding nucleic acid or
portions thereof, may be used
as hybridization probes for a cDNA library to isolate the full-length PRO
polypeptide gene or to isolate still other
genes (for instance, those encoding naturally-occurring variants of the PRO
polypeptide or PRO polypeptides from
other species) which have a desired sequence identity to the PRO polypeptide
nucleic acid sequences. Optionally,
the length of the probes will be about 20 to about 50 bases. The hybridization
probes may be derived from the
naicleotide sequence of any of the DNA molecules disclosed herein or from
genomic sequences including promoters,
erthancer elements and introns of native sequence PRO polypeptide encoding
DNA. By way of example, a screening
method will comprise isolating the coding region of the PRO polypeptide gene
using the known DNA sequence to
synthesize a selected probe of about 40 bases. Hybridization probes may be
labeled by a variety of labels. including
radionucleotides such as ~ZP orss S, or enzymatic labels such as alkaline
phosphatase coupled to the probe via
avidinlbiotin coupling systems. Labeled probes having a sequence complementary
to thaf of the specific PRO
polypeptide gene of the present invention can be used to screen libraries of
human cDNA, genomic DNA or mRNA
to determine which members of such libraries the probe hybridizes to.
Hybridization techniques are described in
further detail in the Examples below.
The ESTs disclosed in the present application may similarly be employed as
probes, using the methods
disclosed herein.
The probes may also be employed in PCR techniques to generate a pool of
sequences for identification of
closely related PRO polypeptide sequences.
Nucleotide sequences encoding a PRO polypeptide can also be used to construct
hybridization probes for
mapping the gene which encodes that PRO polypeptide and for the genetic
analysis of individuals with genetic
disorders. The nucleotide sequences provided herein may be mapped to a
chromosome and specific regions of a
chromosome using known techniques, such as in situ hybridization, linkage
analysis against known chromosomal
markers, and hybridization screening with libraries.
When the coding sequence for the PRO polypeptide encodes a protein which binds
to another protein, the
PRO polypeptide can be used in assays to identify its ligands. Similarly,
inhibitors of the receptor/ligand binding
interaction can be identified. Proteins involved in such binding interactions
can.also be used to screen for peptide
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WO 99/46281 PCTlUS99/05028
or small molecule inhibitors or agonists of the binding interaction. Screening
assays can be designed to find lead
compounds that mimic the biological activity of a native PRO polypeptide or a
ligand for the PRO polypeptide. Such
screening assays will include assays amenable to high-throughput screening of
chemical libraries, making them
particularly suitable for identifying small molecule drug candidates. Small
molecules contemplated include synthetic
organic or inorganic compounds. The assays can be performed in a variety of
formats, including protein-protein
binding assays, biochemical screening assays, immunoassays and cell based
assays, which are well characterized in
the art.
Nucleic acids which encode a PRO polypeptide or its modified forms can also be
used to generate either
uansgenic animals or "knock out" animals which, in turn, are useful in the
development and screening of
therapeutically useful reagents. A transgenic animal (e.g., a mouse or rat) is
an animal having cells that contain a
transgene, which transgene was introduced into the animal or an ancestor of
the animal at a prenatal, e.g., an
embryonic stage. A tran~gene is a DNA which is integrated into the genome of a
cell from which a uansgenic animal
develops. 1n one embodiment, cDNA encoding a PRO polypeptide of interest can
be used to clone genomic DNA
encoding the PRO polypeptide in accordance with established techniques and the
genomic sequences used to generate
uansgenic animals that contain cells which express DNA encoding the PR.O
polypeptide: Methods for generating
tz~ansgenic animals, particularly animals such as mice or rats, have become
conventional in the art and are described,
for example, in U,&. Patent Nos. 4,736,866 arid 4,870,009. Typically,
particular cells would be targeted for PRO
polypeptide uansgene incorporation with tissue-specific etthancers. Transgenic
animals that intbide a copy of a
transgene encoding a PRO polypeptide introduced into the germ line of the
animal at an embryonic stage can be used
to examine the effect of increased expression of DNA encoding the PRO
palypeptide. Such animals can be used as
tester animals for reagents thought to confer protection from, for example,
pathological conditions associated with
its overexpression. In accordance with this facet of the invention, an animal
is ueated with the reagent and a reduced
incidence of the pathological condition, compared to unueated animals bearing
the transgene, would indicate a
potential therapeutic intervention for the pathological condition.
Alternatively, non-human homologues of PRO polypeptides can be used to
construct a PRO polypeptide
"knock out" animal which has a defective or altered gene encoding the PRO
polypeptide of interest as a result of
homologous recombination between the endogenous gene etxoding the PRO
polypeptide a~ alters genomic DNA
encoding the PRO polypeptide introduced into an embryonic cell of the animal.
For example, cDNA encoding a PRO
polypeptide can be used to clone genomic DNA encoding the PRO polypeptide in
accordance with established
techniques. A portion of the genomic DNA encoding a PRO polypeptide can be
deleted or replaced with another
gene, such as a gene encoding a selectable marker which can be used to monitor
integration. Typically, several
kilobases of unaltered flanidng DNA (both at the 5' and 3' ends) are i>tcluded
in the vector (see e.g., Thomas and
Capecchi, ~e , _5:503 {1987) for a description of homologous recombination
vectors]. The vector is introduced into
an embryonic stem cell line (e.g., by electropora6on) and cells in which the
inuoduced DNA has homologously
recombined with the endogenous DNA are selected (see e.g., Li et al., ( ej(,
~,x:915 (1992)). The selected cells are
then injected into a blastocyst of an animal (e.g., a mouse or rat) to forma
aggregation chimeras (see e.g.. Bradley,
in Teratocarcinomas a»d Embryonic Stem Cells: A Practical Appr~clt, lc. J.
Robertson, ed. (IRL, Oxford, 1987),
pp. 113-152]. A ehimeric embryo can then be implanted into a su'ttable
pseudopregnant female foster animal and the
embryo brought to term to create a "knock out" animal. Progeny harboring the
homologously recombined DNA in
their germ cells can be identified by standard techniques and used to breed
animals in which all cells of the anima!
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CA 02421319 2003-03-25
WO 99!46281 PCT/US99/05028
contain the homologously recombined DNA. Knockout animals can be characterized
for instance, for their ability
to defend against certain pathological conditions and for their development of
pathological conditions due to absence
of the PRO poiypeptide.
When in vivo administration of a PRO polypeptide is employed, normal dosage
amounts may vary from
about 10 nglkg to up to I00 mg/kg of mammal body weight or more per day,
preferably about 1 kglkglday to 10
S mg/kglday, depending upon the route of adminisuation. Guidance as to
particular dosages and methods of delivery
is provided in the literature; see, for example, U.S. Pat. Nos. 4,657,760;
5,206,344; or 5,225,212. It is anticipated
that different formulations will be effective for different treatment
compounds and different disorders, that
administration targeting one organ or tissue, for example, may necessitate
delivery in a manner different from that
to another organ or tissue.
Where sustained-release administration of a PRO poiypeptide is desired in a
formulation with release
characteristics suitable for the ueatment of any disease or disorder requiring
administration of the PRO polypeptide,
microencapsulation of the PRO polypeptide is contemplated. Microencapsulation
of recombinant proteins for
sustained release has been successfully performed with human growth hormone
(rhGH), interferon- (rhlFN- ),
interleukin-2, and MN rgp120. Johnson et al., Nat. Med,, _2: 795-799 (1996);
Yasuda. Biomed. Ther., 27: 1221-
1223 (1993); Hora et al., Bio/Techrtologv. _8: 7SS-7S8 (1990); Cleland,
"Design and Production of Single
Immunization Vaccines Using Polylactide Polyglycolide Microsphere Systems," in
Vaccine Designi The Subunit and
Adjuvant Approach, Powell and Newman, eds, (Plenum Press: New York, 1995), pp.
439462; WO 97/03692, WO
96/40072, WO 96107399; and U.5 Pat. No. 5,654,010.
The sustained-release formulations of these proteins were developed using poly-
lactic-coglycolic acid
(PLGA) polymer due to its biocompaubiliry and wide range of biodegradable
properties. The degradation products
of PLGA, lactic and glycolic acids, can be cleared quickly within the human
body. Moreover, the degradability of
this polymer can be adjusted from months to years depending on its molecular
weight and composition. Lewis,
"Conuolled release of bioactive agents from lactidelglycolide polymer," in: M.
Chasin and R. Lattger (Eds.),
B',~egradable Polymers as Drug Delivetv Systems (Marcel Dekker: New York,
1990), pp. 1-41.
For example, for a formulation that can provide a dosing of approximately 80
g/kg/day in mammals with
a m~utimum body weight of 8S kg, the largest dosing would be approximately 6.8
mg of the PRO polypeptide per day.
In order to achieve this dosing level, a sustained- release formulation which
contains a maximum possible protein
krading (1S-20°~c w/w PRO polypeptide) with the lowest possible initial
burst (<20~) is necessary. A continuous
(zero-order) release of the PRO polypeptide from microparticles for 1-2 weeks
is also desirable. In addition, the
encapsulated protein to be released should maintain its integrity and
stability over the desired release period.
PR0213 polypeptides and portions thereof which possess .the ability to
regulate the growth induction cascade
and/or the blood coagulation cascade may also be employed for such purposes
both in vivo therapy and in vitro.
Those of ordinary skill in the art will well know how to employ PR0213
polypeptides for such uses.
PR0274 polypeptides and portions thereof which have homology to 7TM protein
and Ftt54 may also be
33 useful for in vivo therapeutic purposes, as well as for various other
applications. The identification of novel 7TM
protein and FnS4-like molecules may have relevance to a number of human
disorders which involve recognition of
ligands and the subsequent signal transduction of information contained within
those ligands in order to control
cellular processes. Thus, the identification of new 7TM protein and Fn54-like
molecules is of special importance
in that such proteins may serve as potential therapeutics for a variety of
different human disorders. Such polypeptides
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may also play important roles in biotechnological and medical research as well
as in various industrial applications.
As a result, there is particular scientific and medical interest in new
molecules, such as PR0274.
PR0300 polypeptides and portions thereof which have homology to Diff 33 may
also be useful for in vivo
therapeutic purposes, as well as for various other applications. The
identification of novel Diff 33-like molecules may
have relevance to a number of human disorders such as the physiology of
cancer. Thus, the identification of new
Diff 33-like molecules is of special importance in that such proteins may
seine as potential therapeutics for a variety
of different human disorders. Such po3ypeptides may also play important roles
in biotechnological and medical
research as well as various industrial applications. As a result, there is
particular scientific and medical interest in
new molecules, such as PR0300.
PR0296 poIypeptides of the present invention which possess biological activity
related to that of the
sarcoma-amplified SAS protein may be employed both in vivo for therapeutic
purposes and in vitro. Those of
ordinary skill in the art will well know how to employ the PR0296 polypeptides
of the present invention for such
purposes.
PR0329 polypeptides of the present invention which possess biological activity
related to that of
immunoglobulin F~ receptor protein or suburut thereof may be employed both in
vivo for therapeutic purposes and
in vitro. Those of ordinary skill in the an will well know how to employ the
PR0329 polypeptides of the present
invention for such purposes.
PR0362 polypeptides of the present invention which possess biological activity
related to that of the A33
antigen protein, HCAR protein or the NrCAM related cell adhesion molecule may
be employed both in vivo for
therapeutic purposes and in vitro.
PR03b3 polypeptides of the present invention which possess biological activity
related to that of the cell
surface HCAR prooein may be employed both in vivo for therapeutic purposes and
in vitro. Those of ordinary skill
in the art wilt well know how to employ the PR0363 polypeptides of the present
invention for such purposes.
Specifically, exuacellular domains derived from the PR0363 polypeptides may be
employed therapeutically in vivo
for lessening the effects of viral infection.
PR0868 polypeptides of the present invention which possess biological activity
related to that of the tumor
necrosis factor protein may be employed both in vivo for therapeutic purposes
and in vitro. Those of ordinary skill
in the art will well know how to employ the PR0868 poiypeptides of the present
invention for such purposes.
PR0382 polypeptides of the present invention which possess biological activity
related to that of the serine
protease proteins may be employed both in vivo for therapeutic purposes and in
vitro. Those of ordinary skill in the
art will well know how to employ the PR0382 polypeptides of the present
invention for such purposes.
PROS45 polypeptides and portions thereof which have, homology to meltrin may
also be useful for in vivo
~c purposes, as well as for various other applications. The identification of
novel molecules associated with
cellular adhesion may be relevant to a number of human disorders. Given that
the meltrin proteins may play an
important role in a number of disease processes, the identification of new
meltrin proteins and meluin-like molecules
is of special importance in that such proteins may serve as potential
thera~utics for a variety of different human
disorders. Such polypeptides may also play important roles in biotechnological
and medical research, as well as
various industrial applications. As a result, there is particular scientific
and medical interest in new molecules, such
as PR0545.
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CA 02421319 2003-03-25
WO 99/46281 PCTNS99/05028
PR0617 polypeptides of the present invention which possess biological activity
related to that of the CD24
protein may be employed both in vivo for therapeutic purposes and in vitro.
Those of ordinary skill in the art will
well know how to employ the PR0617 polypeptides of the present invention for
such purposes.
PR0700 polypeptides and portions thereof which have homology to protein
disulfide isomerase may also
be useful for in vivo therapeutic purposes, as well as for various other
applications. The identification of novel
protein disulfide isomerases and related molecules may be relevant to a number
of human disorders. Given that
formation of disulfide bonds and protein folding play important roles in a
number of biological processes, the
identification of new protein disulfide isomerases and protein disulfide
isomerase-like molecules is of special
importance in that such proteins may serve as potential therapeutics for a
variety of different human disorders. Such
polypeptides may also play important roles in biotechnological and medical
research, as well as various industrial
applications. As a result, there is particular scientific and medical interest
in new molecules, such as PR0700.
PRO702 polypeptides of the present invention which possess biological activity
related to that of the
conglutinin protein may be employed both in vivo for therapeutic purposes and
in vitro. Those of ordinary skill in
the art will well know now to employ the PR0702 polypeptides of the present
invention for such purposes. PR0702
polypeptides having conglutinin activity would be expected to be capable of
inhibiting haemagglutinin activity by
i5 influenza viruses andlor function as immunoglobulin-independent defense
molecules as a result of a complement-
mediated rtiechat>ism.
PR0703 polypeptides of the present invention which possess biological activity
related to that of the VLCAS
protein may be employed both in vivo for therapeutic purposes and in vitro.
Those of ordinary skill in the art will
well know how to employ the PR0703 polypeptides of the present invention for
such purposes.
PR0703 polypeptides and portions thereof which have homology to VLCAS may also
be useful for in vivo
therapeutic purposes, as well as for various ocher applications. The
identification of novel VLCAS proteins and
related molecules may be relevant to a number of human disorders. Thus, the
identification of new VLCAS proteins
and VLCAS protein-like molecules is of special importance in that such
proteins may serve as potential therapeutics
for a variety of different human disorders. Such polypeptides may also play
important roles in biotechnological and
medical research as well as various industrial applications. As a result,
there is particular scientific and medical
interest in new molecules, such as PR0703.
PR0705 poiypeptides of the present invention which possess biological activity
related to that of the K-
glypicatt protein may be employed both in vivo for therapeutic purposes and in
vitro. Those of ordinary skill in the
art will well know how to employ the PR0705 polypeptides of the present
invention for such purposes.
PR0708 polypeptides of the present invention which possess biological activity
related to that of the aryl
sulfatase proteins may be employed both in vivo for therapeutic purposes and
in vitro. Those of ordinary skill in the
art will well know how to employ the PR0708 polypeptides of the present
invention for such purposes.
PR0320 polypeptides of the present invention which possess biological activity
related to that of the fibulin
protein may be employed both in vivo for therapeutic purposes and in vitro.
Those of ordinary skill in the art will
well know how to employ the PRO320 polypeptides of the present invention for
such purposes.
PR0320 polypeptides and portions thereof which have homology to fibulin may
also be useful for in vivo
therapeutic purposes, as well as for various other applications. The
identification of novel fibulin proteins and related
molecules may be relevant to a number of human disorders such as cancer or
those involving connective tissue,
attachment molecules and related mechanisms. Thus, the identification of new
fibulin proteins and fibulin
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CA 02421319 2003-03-25
WO 99/x6281 PCTNS99/05028
protein-like molecules is of special importance in that such proteins may
serve as potential therapeutics for a variety
of different human disorders. Such polypeptides may also play important roles
in biotechnological and medical
research as well as various industrial applications. As a result, there is
particular scientific and medical interest in
new molecules, such as PR0320.
PRO324 polypeptides of the present invention which possess biological activity
related to that of
oxidoreductases may be employed both in vivo for therapeutic purposes and in
vitro. Those of ordinary skill in the
art will well know how to employ the PR0324 polypeptides of the present
invention for such purposes.
PR0351 poiypeptides of the present invention which possess 1>iological
activity related to that of the
prostasin protein may be employed both in vivo for therapeutic purposes and in
vitro. Those of ordinary skill in the
art will well know how to employ the PR0351 polypeptides of the present
invention for such puiposes.
PR0351 polypeptides and portions thereof which have homology to proscasin may
also be useful for in vivo
therapeutic purposes, as well as for various other applications. The
identification of novelprostasin proteins and
related molecules may be relevant to a number of human disorders. Thus; the
identification of new prostasin proteins
and prostasin -like molecules is of special importance in that such proteins
may serve as potential therapeutics for
a variety of different human disorders. Such poiypeptides may also play
important roles in biotechnological and
medical research as well as various industrial applications. As a result,
there is particular scientific and medical
interest in new molecules, such as PR0351.
PR0352 polypeptides of the present invention which possess biological activity
related to that of the
~tyrophilinprotein may be employed both in vivo for therapeutic purposes and
in vitro. Those of ordinary skill in
the an will well know how to employ the PR0352 polypeptides of the present
invention for such purposes.
PR038I polypeptides of the present invention which possess biological activity
related to that of one or more
of the FKPB immunophilin proteins may be employed both in vivo for therapeutic
purposes and in vitro, for example
for enhancing immunosuppressant activity and/or for axonal regeneration. Those
of ordinary skill in the art will well
know how to employ the PR0381 polypeptides of the present invention for such
purposes.
PR0386 polypeptides of the present invention which possess biological activity
related to that of the beta-2
subunit of a sodium channel expressed in mammalian cells may be emplayed both
in vivo for therapeutic purposes
and in vitro. Those of ordinary skill in tine art will well know how to employ
the PR038b polypeptides of the present
invention for such purposes.
PR0540 polypeptides of tire present invention which possess biological
activity related to that of the LCAT
protein may be employed both in vivo for therapeutic purposes and in vitro.
Those of ordinary skill in the art will
well know how to employ the PR0540 polypeptides of the present invention for
such purposes.
PR0615 polypeptides of the present invention which possess. biological
activity related to that of the
synaptogyrin protein may be employed both in viva for therapeutic purposes and
in vitro. Those of ordinary skill in
the art will well know how to employ the PR0615 polypeptides of the present
invention for such purposes.
PR0615 polypeptides and portions thereof which have homology to synaptogyrin
may also be useful for in
vivo therapeutic purposes, as well as for various other applications. The
identification of novel synaptogyrin proteins
and related molecules may be relevant to a cumber of human disorders. Thus,
the identification of new synaptogyrin
proteins and synaptogyrin-like molecules is of special importance in that
such. proteins may serve as potential
therapeutics 'for a variety of different human disorders. Such polypeptides
may also play important roles in
biotechnological and medical research as well as various industrial
applications. As a result, there is particular
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scientific and medical interest in new molecules, such as PR0615.
PR0618 polypeptides of the present invention which possess biological activity
related to that of an
' enteropeptidase may be employed both in vivo for therapeutic purposes and in
vitro. Those of ordinary skill in the
art will well know how to employ the PR0618 polypeptides of the present
invention for such purposes.
PR0618 polypeptides and portions thereof which have homology to
enteropeptidase may also be useful for
in vivo therapeutic purposes, as well as for various other applications. The
identification of novel enteropeptidase
proteins and related molecules may be relevant to a number of human disorders.
Thus, the identification of new
ettteropeptidase proteins and enteropeptidase-like molecules is of special
importance in that such proteins may serve
as potential therapeutics for a variety of different human disorders. Such
polypeptides may also play important roles
in biotechnological and medical research as well as various industrial
applications. As a result, there is particular
scientific and medical interest in new molecules, such as PR0618.
PR0719 polypeptides of the present invention which possess biological activity
related to that of the
lipoprotein lipase H protein may be employed both in vivo for therapeutic
purposes and in vitro. Those of ordinary
skill in the art will well know how to employ the PR0~19 polypeptides of the
present invention for such purposes.
PR0724 polypeptides of the present invention which possess biological activity
related to that of the human
LDL receptor protein tray be employed both in vivo for therapeutic purposes
and in vitro. Those of ordinary skill
in the art will well know how to employ the PR0724 polypeptides of the present
invention for such purposes.
PR0772 polypeptides of the present invention which possess biological activity
related to that of the human
A4 protein tray be employed both in vivo for therapeutic purposes and in
vitro. Those of ordinary skill in the art will
well know how to employ the PR0772 polypeptides of the present invention for
such purposes.
PR0852 polypeptides of the present invention which possess biological activity
related to that of cet~tain
protease protein tray be employed both in vivo for therapeutic purposes and in
vitro. Those of ordinary skill in the
art will well know how to employ the PR0852 polypeptides of the present
invention for such purposes.
PR0853 polypeptides of the present invention which possess biological activity
related to that of the
reductase protein may be employed both in vivo for therapeutic purposes and in
vitro. Those of ordinary skill in the
art will well know how to employ the PR0853 polypeptides of the present
invention for such purposes.
PR0853 polypeptides and portions thereof which have homology to reductase
proteins tray also be useful
for in vivo therapeutic purposes, as well as for various other applications.
Given that oxygen free radicals and
antioxidants appear to play important roles in a number of disease processes,
the identification of new reductase
proteins and reductase-like molecules is of special importance in that such
proteins may serve as potential therapeutics
for a variety of differern human disorders. Such polypeptides may also play
important roles in biotechnological and
medical research as well as various industrial applications. As a result,
there is particular scietuific and medical
interest in new molecules, such as PR0853.
PR0860 polypeptides of the present invention which possess biological activity
related to that of the
neurofasein protein may be employed both in vivo far therapeutic purposes and
in vitro. Those of ordinary skill in
the art will well know how to employ the PR0860 polypeptides of the present
invention for such purposes.
PR0860 polypeptides and portions thereof which have homology to neurofascin
may also be useful for in
vive therapeutic proposes, as well as for various other applications. The
identification of novel neurofascin proteins
arid related molecules may be relevant to a number of human disorders which
involve cellular adhesion. Thus, the
idetuification of new rteurofascin proteins and neurofascin protein-like
molecules is of special importance in that such
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proteins may serve as potential therapeutics for a variety of different human
disorders. Such polypeptides may also
play importarn roles in biotechnological and medical research as well as
various industrial applications. As a result,
there is particular scientific and medical interest in new molecules, such as
PR0860.
PR084b polypeptides of the present invention which possess biological activity
related to that of the
CMRF35 protein may be employed both in vi o for therapeutic purposes and in
vitro. Those of ordinary skill in the
art will well know how to employ the PR0846 polypeptides of the present
invention for such purposes.
PR0846 polypeptides and portions thereof which have homology to the CMRF35
protein may also be useful
for in vivo therapeutic purposes, as well as for various other applications.
The identification of novel CMRF35
protein and related molecules may be relevant to a number of human disorders.
Thus; the identification of new
CMRF35 protein and CMRF35 protein-like molecules is of special importance in
that such proteins may serve as
potential therapeutics for a variety of different human disorders. Such
polypeptides may also play important roles
in biotechnological and medical research as well as various industrial
applications. As a result, there is particular
scientific and medical interest in new molecules, such as PR0846.
PR0862 polypeptides of the present invention which possess biological activity
related to that of the
lysozyme protein may be employed both in vivo for therapeutic purposes and in
vitro. Those of ordinary skill in the
art will well know how to employ the PR0862 polypeptides of the present
invention for such purposes.
PR0862 polypeptides and portions thereof which have homology to the lysozyme
protein may also be useful
for in vivo therapeutic purposes, as well as for various other applications.
The identification of novel lysozyme
proteins and related molecules may be relevant to a number of human disorders.
Thus, the identification of new
lysozymes and lysozyme-like molecules is of special importance in that such
proteins may serve as potential
therapeutics for a variety of different human disorders. Such polypeptides may
also play important roles in
biotechnological and medical research as well as various industrial
applications. As a result, there is particular
scientific and medical interest in new molecules, such as PRO8b2.
PR0864 poiypeptides of tl~ present invention which possess biological activity
related to that of the Wnt-4
protein may be employed both in vivo for therapeutic purposes and in vitro.
Those of ordinary skill in the art will
well know how to employ the PR0864 polypeptides of the present invention for
such purposes.
PR0864 polypeptides and portions thereof which have homology to the Wnt-4
protein may also be useful
for in vivo therapeutic purposes, as well as for various other applications.
The identification of novel Wnt-4 proteins
attd related molecules may be rekvatz to a rnmnber of human disorders. Thus,
the identification of new Wnt-4 protein
and Wnt-4 protein-like molecules is of special importance in that such
proteins may serve as potential therapeutics
for a variety of different human disorders. Such polypeptides may also play
important roles in biotechnological and
medical research as well as various industrial applications. As a result,
there is particular scientific and medical
interest in new molecules, such as PR0864.
PR0792 polypeptides of the present invention which possess biological activity
related to that of the CD23
protein may be employed both in vivo for therapeutic purposes and in vitro.
Those of ordinary skill in the art will
well know how to employ the PR0792 polypeptides of the present invention for
such purposes.
PR0866 polypeptides of the present invention which possess biological activity
related to that of mindin
amllor spondin protein may be employed both in vivo for therapeutic purposes
and in vitro. Those of ordinary skill
in the art will well latow how to employ the PR0866 polypeptides of the
present invention for such purposes.
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CA 02421319 2003-03-25
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PR0871 polypeptides of the present invention which possess biological activity
related to that of the
cyclophiiin protein family may be employed both in vivo for therapeutic
purposes and in vitro. Those of ordinary
skill in the art will well lrnow how io employ the PR0871 polypeptides of the
.present invention for such purposes.
PR0873 polypegtides of the present invention which possess biological activity
related to that of
carboxylesterases may be employed both in viva for therapeutic purposes and in
vitro. For example, they be used
in conjunction with prodrugs to convert the prodrug to its active form (see
Danks et al.,sttpra). They may be used
to inhibit parasite infection (see van Pelt et al., supra). Methods for employ
the PR0873 polypeptides of the present
invention for these, and other purposes will be readily apparent to those of
ordinary skill in the art.
PR0940 polypeptides of the present invention which possess biological activity
related to that of the CD33
protein andlor OB binding protein-2 tray be employed both in vivo for
therapeutic purposes and in vitro. Those of
ordinary skill in the art will well know how to employ the PR0940 polypeptides
of the present invention for such
purposes.
PR0941 polypeptides of the present invention which possess biological activity
related to that of a cadherin
protein may be employed both in vivo for therapeutic purposes and in vitro.
Those of ordinary skill in the art will
well know how to employ the PR0941 polypeptides of the present invention for
such purposes.
PR0944 polypeptides of the present invention which possess biologicai activity
related to that of the CPE-R
protein may be employed both in vivo for therapeutic purposes and in vitro.
Those of ordinary skill in the art will
well know how to employ the PR0944 polypeptides of the present invention for
such purposes. PR0944 polypeptides
of the present invention that function to bind to Clostridium perfringens
enterotoxin (CPE) may find use for effectively
treating infection by the CPE endotoxia.
PR0983 polypeptides of the present invention which possess biological activity
related to that of the vesicle-
associated r~mbra~ protein, VAP-33, tray be employed both in vivo for
therapeutic purposes and in vitro. Those
of ordinary skill in the art will well know how to employ the PR0983
polypeptides of the present invention for such
purposes.
PR01057 polypeptides of the present invention which possess biological
activity related to that of protease
proteins may be employed both in vivo for therapeutic purposes and in vitro.
Those of ordinary skill in the art will
well know how to employ the PR01057 polypeptides of the present invention for
such purposes.
PR01071 polypeptides of tlae present invention which possess biological
activity related to that of the
thrombospondin protein may be employed both in vivo for therapeutic purposes
and in vitro. Those of ordinary skill
in the an will well know how to employ the PR01071 poIypeptides of the present
invention for such purposes.
PR01072 polypeptides of the presetu invention which possess biological
activity related to that of reductase
proteins may be employed both in vivo for therapeutic purposes.and in vitro.
Those of ordinary skill in the art will
well know how to employ the PR01072 polypeptides of the present invention for
such purposes.
PR01075 polypeptides of the present invention which possess biological
activity related to that of protein
disulfide isomerase may be employed both in vivo for therapeutic purposes and
in vitro. Those of ozdittary skill in
the art will well know how to employ the PR01075 polypeptides of the present
invention for such purposes.
PR0181 polypeptides of the present invention which possess biological activity
related to that of the
cotnichon protein may be employed both in vivo for therapeutic purposes and in
vitro. Those of ordinary skill in the
art will well know how to employ the PROl8i polypeptides of the present
invention for such purposes.
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PR0827 polypeptides of the present invention which possess biological activity
related to that of various
integrity proteins may be employed both in vivo for therapeutic purposes and
in vitro. Those of ordinary skill in the
art will well know how to employ the PR0827 polypeptides of the present
invention for such purposes.
PRO1114 polypeptides of the present invention which possess biological
activity related to that of the
cytokine receptor family of proteins may be employed both in vivo for
therapeutic purposes and in vitro. Those of
ordinary skill in the art will well know how to employ the PROl l 14
polypeptides of the present invention for such
purposes.
1n addition to the above, the PR01114 interferon receptor polypeptides may be
employed in applications,
both in vivo and in vitro, where the ability to bind to an interferon ligand
is desired. Such applications will be well
within the skill level in the an.
PR023? polypeptides of the present invention which possess biological activity
related to that of the carbonic
anhydrase protein may be employed both in vivo for therapeutic purposes and in
vitro. Those of ordinary skill in the
art will well know how to employ the PR0237 polypeptides of the present
invention for such purposes.
PR0541 polypeptides of the present invention which possess biological activity
related to that of a trypsin
iniu'bitor protein may be employed both in vivo for therapeutic purposes and
in vitro. Those of ordinary skill in the
art will well know how to employ the PR0541 polypeptides of the present
invention for such purposes.
PR0273 polypeptides can be used in assays that other chemokines would be used
in to perform comparative
assays. The results can be used accordingly.
PR0701 polypeptides of the present invention which possess biological activity
related to that of the
r~urofigin fatnt~y may be employed both in vivo for therapeutic purposes and
in vitro. Those of ordinary skill in the
art will well know how to employ the PR0701 polypeptides of the present
invention for such purposes.
PR0701 can be used in assays with neurons and its activity thereon c;an be
compared with that of neuroligins
1, 2 and 3. The results can be applied accordingly.
PR0704 polypeptides of the present invemion which possess biological activity
related to that of vesicular
integral membrane proteins may be employed both in vivo for therapeutic
purposes and in vitro. Those of ordinary
skill in tt~ art will well know how to employ the PR0704 polypeptides of the
present invention for such purposes.
PR0704 can be used in assays with the polypeptides to which they have identity
with to detertnine the
relative activities. The resuks can be applied accordingly. PR0704 can be
tagged or measured for activity to
measure endocytosis activity and thereby used to screen for agents which
effect endocytosis.
PR0706 polypeptides of the present invention which possess biological activity
related to that of the
erogenous prostatic acid phosphatase precursor may be employed both in vivo
for therapeutic purposes and. in vitro.
Those of ordinary skill in the art will well know how to employ the PRO'706
poiypeptides of the present invention
for such purposes.
PR0706 can be used in assays with human prostatic acid phosphat~ise or human
IysosomaI acid phosphatase
and its activity thereon can be compared with that of human prostatic acid
phosphatase or human lysosomal acid
phosphatase. The results can be applied at;cordiagly.
PR0707 polypeptides of the present invention which possess biological activity
related to that of cadherins
may be employed both in vivo for therapeutic purposes and in vitro. Those of
ordinary skill in the art will well know
how to employ the PR0707 polypeptides of the present invention for such
purposes.
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PR0707 can be used in assays to determine its activity in relation to other
cadherins, particularly cadherin
FIB3. The results can be applied accordingly.
PR0322 polypeptides of the present invention which possess biological activity
related to that of neuropsin
may be employed both in vivo for therapeutic purposes and in vitro. Those of
ordinary skill in the art will well know
how to employ the PR0322 polypeptides of the present invention for such
purposes.
PR0322 can be used in assays to determine its activity relative to neuropsin,
trypsinogen, serine protease
and neurosin, and the results applied accordingly.
PR0526 polypeptides of the present invention which possess biological activity
related to that of protein-
protein binding proteins may be employed both in vivo for therapeutic purposes
and in vitro. Those of ordinary skill
in the art will well know how to employ the PR0526 polypeptides of the present
invention for such purposes.
Assays can be performed with growth factors and other proteins which are known
to form complexes to
determine whether PR0526 binds thereto and whether there is increased half
life due to such binding. The results
can be used accordingly.
PR0531 polypeptides of the present invention which possess biological activity
related to that of the
protocadherins tnay be employed both in vivo for therapeutic purposes and in
vitro. Those of ordinary skill in the
an will well know how to employ the PR0531 polypeptides of the present
invention for such purposes.
PR0531 can be used in assays against protocadherin 3 and other protocadherins,
to determine their relative
activities. The results can be applied accordingly.
PR0534 polypeptides of the present invention which possess biological activity
related to that of the protein
disulfide isomerase may be employed both in vivo for therapeutic purposes and
in vitro. Those of ordinary skill in
the art will well know how to employ the PR0534 polypeptides of the present
invention for such purposes.
PR0534 can be used in assays with protein disulfide isomerase to determine the
relative activities. The
results can be applied accordingly.
PR0697 polypeptides of the present invention which possess biological activity
related to that of the sFRP
family may be employed both in vivo for therapeutic purposes and in vitro.
Those of ordinary skill in the art will well
know how to employ the PR0697 polypeptides of the present invention for such
purposes.
PR0697 can be used in assays with sFRPs and SARPs to determine the relative
activities. The results can
be applied accordingly.
PR0731 polypeptides of the present invention which possess biological activity
related to that of any
protocadtterin may be employed both in vivo for therapeutic purposes and in
vitro. Those of ordinary skill in the art
will well know how to employ the PR0731 polypeptides of the present invention
for such purposes.
PR0731 can be used in assays with the polypeptides to which they have identity
with to determine the
relative activities. The results can be applied accordingly.
PR0768 polypeptides of the present invention which possess biological activity
related to that of integrins
tray be employed both in vivo for therapeutic purposes and in vitro. Those of
ordinary skill in the art will well know
how to employ the PR0768 polypeptides of the present invention for such
purposes.
PR0768 can be used in assays with the polypeptides to which they have identity
with to determine the
relative activities. The results can be applied accordingly.
PR0771 polypeptides of the present invention which possess biological activity
related to that of the testican
protein may be employed both in vivo for therapeutic purposes and in vitro.
Those of ordinary skill in the art will
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WO 99146281 PCTNS99105028
well know how to employ the PR0771 palypeptides of the present invention for
such purposes.
PR0771 can be used in assays with the polypeptides to which they have identity
with to determine the
relative activities. The results can be applied accordingly.
PR0733 polypepddes of the present invention which possess biological activity
related to that of the proteins
which bind the T1/ST2 receptor may be employed both in vivo for therapeutic
purposes and in vitro. Those of
ordinary skill in the art will well know how to employ the PR0733 polypeptides
of the present invention for such
purposes.
PR0733 can be used in assays with the polypeptides to which they have identity
with to determine the
relative activities. The results can be applied accordingly.
PR0162 polypeptides of the present invention which possess biological activity
related to that of the
patios-associated protein may be employed both in vivo for therapeutic
purposes and in vitro. Those of ordinary
skill in the art will well know how to employ the PROl62 polypeptides of the
present invention for such purposes.
PR0162 can be used in assays with the polypeptides to which they have identity
with to determine the
relative activities. The results can be applied accordingly.
PR0788 polypeptides of the present invention which possess biological activity
related to that of the anti-
neoplastic urinary protein may be employed both in vivo for therapeutic
purposes and in vitro. Those of ordinary
skill in the art will well know how to employ the PR0788 polypeptides of the
present invention for such purposes.
PR0788 can be used in assays with the polypeptides to which they have identity
with to determine the
relative activities. The results can be applied accordingly.
PR01008 polypeptides of the presetu im~ention which possess biological
activity related to that of dkk 1 may
be empioyed both in vivo for therapeutic purposes and in vitro. Those of
ordinary skill in the art will well know how
to employ the PR01008 polypeptides of the present invention for such purposes.
PR01008 can be used in assays with the polypeptides to which they have
identity with to determine the
relative activities. The results can be applied accordingly.
PROl0I2 polypeptides of the present invention which possess biological
activity related to that of the protein
disulfide isomerase may be employed both in vivo and in vitro purposes. Those
of ordinary skill in the art will well
know how to employ the PR01012 polypeptides of the present invention for such
purposes.
PROI0I2 can be used in assays with the polypepticles to which they have
identity with to determine the
relative activities. The results. can be applied accordingly.
PROlUI4 polypeptides of the present invention which possess biological
activity related to that of reductase
may be employed both in vivo for therapeutic purposes and in vitro. Those of
ordinary skill in the art will well know
how to employ the PR01014 polypeptides of the present invention for such
purposes.
PR01014 can be used in assays with the polypeptides to which they have
identity with to determine the
relative activities. Inhibitors of PROI014 are particularly preferred. The
results can be applied accordingly.
PR01017 polypeptides of the present invention which possess biological
activity related to that of
sulfotransferase tray be employed both in vivo for therapeutic purposes and!
in vitro. Those of ordinary skill is the
art will well know how to employ the PR01017 polypeptides of the present
invention for such purses.
PR01017 can be used in assays with the polypeptides to which they have
identity wide to determine the
relative activities. The results can be applied accordingly.
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PR0474 polypeptides of the present invention which possess biological activity
related to that of
dehydrogenase may be employed both in vivo for therapeutic purposes and in
vitro. Those of ordinary skill in the
art will well know how to employ the PR0474 polypeptides of the present
invention for such purposes.
PR0474 can be used in assays with the polypeptides to which they have identity
with to determine the
relative activities. The results can be applied accordingly.
PR01031 polypeptides of the present invention which possess biological
activity related to that of IL-17 may
be employed both in vivo for therapeutic purposes and in vitro. Those of
ordinary skill in the art will well know how
to employ the PR01031 polypeptides of the present invention for such purposes.
PR01031 can be used in assays with the polypeptides to which they have
identity with to determine the
relative activities. The results can be applied accordingly.
PR0938 polypeptides of the present invention which possess biological activity
related to that of protein
disulfide isomerase may be employed both in vivo for therapeutic purposes and
in vitro. Those of ordinary skill in
the art will well know how to employ the PR0938 polypeptides of the present
invention for such purposes.
PR01082 polypeptides of the present invention which possess biological
activity related to that of the LDL
receptor may be employed both in vivo for therapeutic purposes and in vitro.
Those of ordinary skill in the art will
well know how to employ the PR01082 polypeptides of the present invention for
such purposes.
PR01082 can be used in assays with the polypeptides to which they have
identity with to determine the
relative activities. The results can be applied accordingly. PR01082 can also
be used in assays to identify candidate
agents which modulate the receptors.
PR01083 polypeptides of the present invention which possess biological
activity related to that of 7TM
receptors may be employed both in vivo for therapeutic purposes and in vitro.
Those of ordinary stall in the art will
well know how to employ the PR01083 polypeptides of the present invention for
such purposes.
In particular PR01083 can be used in assays to determine candidate agents
which control or modulate
PR01083, i.e., have an effect on the receptor.
The VEGF-E molecules herein have a number of therapeutic uses associated with
survival, proliferation
and/or differention of cells. Such uses include the treatment of umbilical
vein endothelial cells, in view of the
trated ability of VEGF-E to increase survival of human umbilical vein
endothelial cells. Treatment may be
needed if the vein were subjected to traumata, or situations wherein
artificial means are employed to enhance the
survival of the umbilical vein, for example, where it is weak, diseased, based
on an artificial matrix, or in an
artificial environment. Other physiological conditions that could be improved
based on the selective mitogenic
character of VEGF-E are also included herein. Uses also include the ueatment
of fibroblasts and myocytes, in view
of the demonstrated ability of VEGF-E to induce proliferation of fibroblasts
and hypertrophy in myocytes. In
particular, VEGF-E can be used in wound healing, tissue growth and musclc
generation and regeneration.
For the indications referred to above, the VEGF-E molecule will.be formulated
and dosed in a fashion
consistent with good medical practice taking into account the specific
disorder to be ueated, the condition of the
individual patient, the site of delivery of the VEGF-E, the method of
administration, and other factors known to
practitioners. Thus, for purposes herein, the "therapeutically effective
amount" of the VEGF-E is an amount that
is effective either to prevent, lessen the worsetW g of, alleviate, or cure
the ueated condition, in particular that amount
which is sufficient to enhance the survival, proliferation and/or
differentiation of the treated cells in vivo.
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VEGF-E amino acid variant sequences and derivatives that are itnmunologically
crossreactive with antibodies
raised against native VEGF are useful in immunoassays for VEGF-E as standards,
or, when labeled, as competitive
reagents.
The VEGF-E is prepared for storage or administration by mixing VEGF-E having
the desired degree of
purity with physiologically acceptable carriers, excipients, or stabilizers.
Such materials are non-toxic to recipients
at the dosages and concentrations employed. If the VEGF-E is water soluble, it
may be formulated in a buffer such
as phosphate or other organic acid salt preferably at a pH of about 7 to 8. if
the VEGF-E is only partially soluble
in water, it may be prepared as a microemuIsion by formulating it with a
nonionic surfactant such as Tween,
Pluronics, or PEG, e.g., Tween 80, in an amount of 0.04-0.05 % (wlv), to
increase its solubility.
Optionally other ingredients may be added such as antioxidants, e.g., ascorbic
acid; low molecular weight
(less than about ten residues) polypeptides, e.g., polyarginine or
tripeptides; proteins, such as serum albumin, gelatin,
or inununoglobulins; hydrophilic polymers such as polyvirryipyrrolidone; amino
acids, such as glycine, glutamic acid,
aspartic acid, or arginine; monosaccharides, disaccharides, and other
carbohydrates including cellulose or its
derivatives, glucose, mannose, or dexuins; chelating agents such as EDTA; and
sugar alcohols such as mannitol or
sorbitol.
The VEGF-E to be used for therapeutic administration must be sterile.
Sterility is readily accomplished by
filtration through sterile filtration membranes (e.g., 0.2 micron membranes).
The VEGF-E ordinarily will be stored
in lyophilized form or as an aqueous solution if it is highly stable to
thermal and oxidative denaturation. The pH of
the VEGF-E preparations typically will be about from 6 to 8, although 1>igher
or lower pH values may also be
appropriate in certain instances. It will be understood that use of certain of
the foregoing excipients, carriers, or
stabilizers will result in the formation of salts of the VEGF-E.
if the VEGF-E is to be used parenterally, therapeutic compositions containing
the VEGF-E generally are
placed into a container having a sterile access port, for example, an
inuavenous solution bag or vial having a stopper
pierceable by a hypodermic injection needle.
Generally, where the disorder permits, one should formulate and dose the VEGF-
E for site-specific delivery.
This is convenient in the case of wounds and ulcers.
Sustained release formulations may also be prepared, and include the formation
of microcapsular particles and
implantable articles. For preparing sustained-release VEGF-E compositions, the
VEGF-E is preferably incorporated
into a biodegradable matrix or microcapsule. A suitable rnateriai for this
purpose is a polylactide, although, other
polymers of poly-(a-hydroxycarboxylic acids), such as poly-D-{-)-3-
hydroxybutyric acid (EP 133,988A), can be used.
Other biodegradable polymers include poly{lactones), poly(acetals),
poly(orthoesters), or poly(orthocarbonates). The
initial consideration here must be that the carrier itself, or its degradation
products, is nontoxic in the target tissue
and will not further aggravate the condition. This can be determined by
rbutine screening in animal models of the
target disorder or, if such models are unavailable, in normal animals:
Numerous scientific publications document
such animal models.
For examples of sustained release compositions, see U.S. Patent No. 3,773,919,
EP 58,481A; U.S. Patent
No. 3,887,699, EP 158;277A, Canadian Patent No. 1176565, U. Sidman et al.,
Biopolymers 22, 547 [1983]; and
R. Larger et al., Chem. Tech. 12, 98 [1982).
When applied topically, the VEGF-E is suitably combined with other
ingredients, such as carriers andlor
adjuvants. There are no limitations on the nature of such other ingredients,
except that they must be pharmaceutically ..
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acceptable and efficacious for their intended administration, and catmot
degrade the activity of the active ingredients
of the composition. Examples of suitable vehicles include ointments, creams,
gels, or suspensions, with or without
purified collagen. The compositions also may be impregnated into transdermal
patches, plasters, and bandages,
preferably in liquid or semi-liquid form.
For obtaining a gel formulation, the VEGF-E formulated in a liquid composition
may be mixed with an
effective amount of a water-soluble polysaccharide or synthetic polymer such
as polyethylene glycol to form a gel
of the proper viscosity to be applied topically. The polysaccharide that may
be used includes, for example, cellulose
derivatives such as etherified cellulose derivatives, including alkyl
celluloses, hydroxyalkyl ceiluloses, and
alkylhydroxyalkyl celluloses, for example, methyiceIlulose, hydroxyethyl
cellulose, carboxymethyl ceihtIose,
hydroxypropyl methylcellulose, and hydroxypropyl cellulose; starch and
fractionated starch; agar; alginic acid and
alginates; gum arabic; pullulian; agarose; carrageenan; dextrans; dexuins;
ftuctans; inulin; mannans; xylans;
arabinans; chitosans; glycogens; glucans; and synthetic biopolymers; as well
as gums such as xanthan gum; guar
gum; locust bean gum; gum arabic; tragacanth gum; and karaya gum; and
derivatives and mixtures thereof. The
preferred gelling agent herein is one that is inert to biological systems;
nontoxic, simple to prepare, and not too runny
or viscous, and will not destabilize the VEGF-E held within it.
Preferably the polysaccharide is an etherified cellulose derivative, more
preferably one that is well defined,
purified, aid listed in USP, e.g., methylcellulose and the hydroxyalkyl
cellulose derivatives, such as hydroxypropyl
cellulose; hydroxyethyl cellulose, and hydroxypropyl methylcelhtlose. Most
preferred herein is tnethylcellulose.
The polyethylene glycol useful for gelling is typically a mixture of low and
high molecular weight
polyethylene glycols to obtain the proper viscosity. For example, a mixture of
a polyethylene glycol of molecular
weight 400-600 with one of molecular weight 1500 would be effective for this
purpose when mixed in the proper ratio
to obtain a paste.
The term "water soluble" as applied to the polysaccharides and polyethylene
giycols is meant to include
colloidal solutions and dispersions. In general, the solubility of the
cellulose derivatives is determined by the degree
of substitution of ether groups, and the stabilizing derivatives useful herein
should have a sufficient quantity of such
ether groin per anhydroglucose unit in the cellulose chant to render the
derivatives water soluble. A degree of ether
substitution of at least 0.35 ether groups per anhydroglucose unit is
generally sufficient. Additionally, the cellulose
derivatives may be in. the form of alkali metal salts, for example, the Li,
Na, K, or Cs salts.
If methylcellulose is employed in the gel, preferably it comprises about 2-5 %
, more preferably about 3 ~,
of the gel and the VEGF is present in an amount of about 300-1000 mg per ml of
gel.
The dosage to be employed is dependent upon the factors described above. As a
general proposition, the
VEGF-E is formulated and delivered to the target site or tissue at a dosage
capable of establishing in the tissue a
VEGF-E level greater than about 0.1 ng/cc up to a maximum dose that is
efficacious but not unduly toxic. This
infra-tissue concentration should be maintained if possible by continuous
infusion, sustained release, topical
application, or injection at empirically determined freque~ies.
It is within the scope hereof to combine the VEGF-E therapy with other novel
or conventional therapies
(e.g., growth factors such as VEGF, aFGF, bFGF, PDGF, IGF, NGF, anabolic
steroids, EGF or TGF-a) for
enhancing the activity of any of the growth factors, including VEGF-E, in
promoting cell proliferation, survival,
differentiation and repair. It is not necessary that such cocreatment drugs be
included per se in the compositions of
this invention, although this will be convenient where such drugs are
proteinaceous. Such admixtures are suitably
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WO 99146281 PCTIUS99/05028
administered in the same manner and for the same purposes as the VEGF-E used
alone. The useful molar ratio of
VEGF-E to such secondary growth factors is typically 1:0.1-10, with about
equimolar amounts being preferred.
The compounds of the present invention can be formulated according to known
methods to prepare
pharmaceutically useful compositions, whereby the PRO polypeptide hereof is
combined in admixture with a
pharmaceutically acceptable carrier vehicle. Suitable carrier vehicles and
their formulation, inclusive of other human
proteins, e.g., human serum albumin, are described, for example, in
Remington's Pjtaarmaceutical Sciences, 16th ed.;
1980, Mack Publishing Co., edited by Oslo et al. the disclosure of which is
hereby incorporated by reference. The
VEGF-E herein may be administered parenterally to subjects suffering from
cardiovascular diseases or conditions,
or by other methods that ensure its delivery to the bloodstream in an
effective form.
Compositions particularly well suited for the clinical administration of VEGF-
E hereof employed in the
practice of the present invention include, for example, sterile aqueous
solutions, or sterile hydratable powders such
as lyophilized protein. It is generally desirable to include further in the
formulation an appropriate amount of a
pharmaceutically acceptable salt, generally in an amount sufficient to render
the formulation isotonic. A pH regulator
such as arginine base, and phosphoric acid, are also typically included in
sufficient quantities to maintain an
appropriate pH, generally from 5.5 to 7.5. Moreover, for improvement of shelf
life or stability of aqueous
formulations, it may also be desirable to include furthez agents such as
glycerol. In this manner, variant t-PA
formulations are rendered appropriate for parenteral administration, and, in
particular, intravenous administration.
Dosages and desired drug concentrations of pharmaceutical compositions of the
present invention may vary
depending on the particular use envisioned: For example, in the treatment of
deep vein thrombosis or peripheral
vascular disease, "bolus" doses, will typically be preferred with subsequent
administrations being given to maintain
an approximately constant blood level, preferably on the order of about 3
pg;/ml.
However, for use in connection with emergency medical care facilities where
infusion capability is generally
not available and due to the generally critical nature of the underlying
disease (e.g., embolism, infarct), it will
generally be desirable to provide somewhat larger initial doses, such as an
intravenous bolus.
For the various therapeutic indications referred to for the compoumis hereof,
Lhe VEGF-E molecules will
be formulated and dosed in a fashion consistent with good medical practice
taking into account the specific disorder
to be treated, the condition of the individual patient; the site of delivery,
the method of administration and other
factors known to practitioners in the respective art. Thus, for purposes
herein, the "therapeutically effective amount"
of the VEGF-E molecules hereof is an amount that is effective either to
prevent, lessen the worsening of, alleviate,
or cure the treated condition, in particular that amount which is sufficient
to enhance the survival, proliferation or
differentiation of targeted cells in vivo. In general a dosage is employed
capable of establishing in the tissue that is
the target for the therapeutic indication being ueated a level of a VEGF-E
hereof greater than about 0.1 nglcm' up
to a maximum dose that is efficacious but not unduly toxic. It is contemplated
that infra-tissue administration may
be the choice for certain of the therapeutic indications for the compounds
hereof.
The human Toll proteins of the present invention can also be used in assays to
identify other proteins or
molecules involved in Toll-mediated signal transduaion. For example, PR0285
and PR0286 are useful in identifying
the as of yet unknown natural ligands of human Tolls, or other factors that
participate (directly or indirectly) in the
activation of and/or signaling through a human Toll receptor, such as
potential Toll receptor associated kinases. In
addition, inhibitors of the receptorlligand binding interaction can be
identified. Proteins involved in such binding
itueractions can also be used to screen for peptide or small molecule
inhibitors or agonists of the binding interaction.
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Screening assays can be designed to find lead compounds that mimic the
biological activity of a native Toll
polypeptide or a ligand for a native Toll polypeptide. Such screening assays
will include assays amenable to high-
throughput screening of chemical libraries, making them particularly suitable
for identifying small molecule drug
candidates. Small molecules contemplated include synthetic organic or
inorganic compounds. The assays can be
performed in a variety of formats, including protein-protein binding assays,
biochemical screening assays,
immunoassays and cell based assays, which are well characterized in the art.
In vitro assays employ a mixture of components including a Toll receptor
polypeptide, which may be part
of fusion product with another peptide or polypeptide, e.g., a tag for
detecting or anchoring, etc. The assay mixtures
may further comprise (for binding assays) a natural infra- or extracellular
ToII binding target (i.e. a Toll Iigand, or
another molecule known to activate andlor signal through the Toll receptor).
While native binding targets may be
used, it is frequently preferred to use portion of such native binding targets
(e.g. peptides), so long as the portion.
provides binding affinity and avidity to the subject Toll protein conveniently
measurable in the assay. The assay
mixture also contains a candidate pharmacological agent. Candidate agents
encompass numerous chemical classes,
through typically they are organic compounds, preferably small organic
compounds, ate are obtained from a wide
variety of sources, including libraries of synthetic or natural compounds. A
variety of other reagents may also be
IS included in the mixture, such as, salts, buffers, neutral proteins, e.g.
albumin, detergents, protease inhibitors,
nuclease inhibitors, antimicrobial agents, etc
In in vitro birxiirtg assays, the resultant mixture is incubated under
conditions whereby, but for the presence
of the candidate molecule, the Toll protein specifically binds the cellular
binding target, portion or analog, with a
reference binding affinity. The mixture cornponetus can be added in any order
that provides for the requisite bindings
and incubations may be performed at any temperature which facilitates optimal
binding. Incubation periods are
likewise selected for optimal binding but also minimized to facilitate rapid
high-throughput screening.
After incubation, the agent-biased binding between the Toll protein and one or
more binding targets is
detecoed by any convenient technique. For cell-free binding type assays, a
separation step is often used to separate
bound from unbound compo~nts. Separation may be effected by precipitation
(e.g. TCA precipitation,
immunoprecipitation, etc.), immobilization (e.g on a solid substrate), etc.,
followed by washing by, for example,
membrane filtration (e.g. Whattnan's P-18 ion exchange paper, Polyfiltronic's
hydrophobic GFC membrane, etc.),
gel chromatography (e.g. gel filtration, affinity, etc.), For Toll-dependent
transcription assays, binding is detected
by a change in the expression of a Toll-dependent reporter.
Detection may be effected in any convenient way. For cell-free binding assays,
one of the components
usually comprises or is coupled to a label. The label may provide for direct
detection as radioactivity, luminescence,
optical or elecuort density, etc., or indirect detection, such as, an epitope
tag, an enzyme, etc. A variety of methods
may be used to detect the label depending on the nature of the label and other
assay components, e.g. through optical
or electron density, radiative emissions, nonradiative energy transfers, etc.
or indirectly detected with antibody
conjugates, etc.
Nucleic acid encoding the Toll polypeptides disclosed herein may also be used
in gene therapy. In gene
therapy applications, genes are introduced inw cells in order to achieve in
vivo synthesis of a therapeutically effective
genetic product, for example for replacement of a defective gene. "Gene
therapy" includes both conventional gene
therapy where a lasting effect is achieved by a single treatment, and the
administration of gene therapeutic agents,
which involves the one time or repeated administration of a therapeutically
effective DNA or mRNA. Antisense
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CA 02421319 2003-03-25
WO 99146281 PCT/US991t)5018
RNAs and DNAs can be used as therapeutic agents for blocking the expression of
certain genes in vivo. It has
already been shown that short antisense oligonucleotides can be imported into
cells where they act as inhibitors,
despite their low intracellular concentrations caused by their restricted
uptake by the cell membrane. (Zamecnik er
al., Proc. Natl: Acad. Sci. USA 83, 4143-4146 [1986]). The oligonucleotides
can be modified to enhance their
uptake, e.g. by substituting their negatively charged phosphodiester groups by
uncharged groups.
There ate a variety of techniques available for introducing nucleic acids into
viable cells. The techniques
vary depending upon whether the nucleic acid is transferred into cultured
cells in vitro, or in vivo in the cells of the
intended host. Techniques suitable for the transfer of nucleic acid into
mammalian cells in vitro include the use of
liposomes, electroporation, microinjection, cell fusion, DEAF-dexuan, the
calcium phosphate precipitation method,
etc. The currently preferred in vivo gene transfer techniques include
transfection with viral (typically retrovirai)
vectors and viral coat protein-liposome mediated transfection (Dzau et al. ,
Trends in Biotechnologx ~, 205-210
[1993n. In some situations it is desirable to provide the nucleic acid source
with an agent that targets the target cells,
such as an antibody specific for a cell surface membrane protein or the target
cell, a ligand for a receptor on the target
cell, etc. Where liposomes are employed, proteins which bind to a cell surface
membrane protein associated with
endocytosis may be used for targeting andlor to facilitate uptake, e.g. capsid
proteins or fragments thereof tropic for
a particular cell type, antibodies for proteins which undergo internalization
in cycling, proteins that target intracellular
localization and enhance intracellular half life. The technique of receptor-
mediated endocytosis is described, for
example, by Wu et al., ~ Biol. Chem. ~, 4429-4432 (1987); and Wagner et al.,
Proc. Natl. Acad. Sci. USA $;Z,
3410-3414 (1990). For review of the currently lrnown gene marking and gene
therapy protocols see Anderson et al.,
'Science ~, 8U8-813 (1992).
The various uses listed in connection with the Toll proteins herein, are also
available for agonists of the
native Toll receptors, which mimic at least one biological function of a
native Toll receptor.
Neurotrimin as well as other members of the IgL.ON subfamily of the
immttnoglobulin superfamily have
been identified to have effect upon neural patterning, differentiation,
maturation and growth. As a result, PR0337
the human neurotrimin homolog polypeptides would be expected to have utility
in diseases which are characterized
by neural disfunction. For example, motoneuron disorders such as amyotrophic
lateral sclerosis (Lou Gehrig's
disease), Bell's palsy, and various condition involving spinal muscular
atrophy, or paralysis. NGF variant
formulations of the invention can be used to neat human neurodegenerative
disorders, such as Alzheimer's disease,
Parkinson's disease, epilepsy, multiple sclerosis; Huntington's chorea, Down's
Syndrome, t~rve deafr~ss; and
Meniere's disease. Moreover PR0337 polypeptide may also be used as a cognitive
enhartcer, to enhance learning
particularly in dementia or trauma, such as those associated with the above
diseases.
Further, PR0337 may be employed to treat neuropathy; and especially peripheral
neuropathy. "Peripheral
neuropathy" refers to a disorder affecting the peripheral nervous system, most
often ~tifested as one or a
combination of motor, sensory, sensorimotor, or autonomic neural dysfunction.
The wide variety of morphologies
exhibited by peripheral tieuropathies can each be attributed uniquely to an
equally wide number of causes. For
example, peripheral neuropathies can be genetically acquired, can result from
a systemic disease, or can be induced
by a toxic agent. Examples include but are not limited to diabetic peripheral
neuropathy, distal sensorimotor
neuropathy, or autonomic neuropathies such as reduced motility of the
gastrointestinal uact or stony of the urinary
bladder. Examples of nev>~athies associated with systemic disease include post-
polio syndrome or AIDS-associated
neuropathy; examples of hereditary neuropathies include Charcot-Marie-Tooth
disease, Refsum's disease,
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Abetalipoproteinemia, Tangier disease, Krabbe's disease, Metachromatic
leukodysuophy, Fabry's disease, and
Dejerine-Soaas syndrome; and examples of neuropathies caused by a toxic agent
include those caused by ueatment
with a chemotherapeutic agent such as vincristine, cisplatin, methouexace, or
3'-azido-3'-deoxythymidine.
Correspondingly, neurotrimin antagonists would be expected to have utility in
diseases characterized by excessive
neuronal activity.
Endothelia is generated from inactive intermediates, the big endothelins, by a
unique processing event
catalyzed by the zinc metalloprotease, endothelia convening enzyme (ECE). ECE
was recently cloned, and its
structure was shown to be a single pass transmembrane protein with a short
inuacellular N-tetrninal and a long
extracellular C-terminal that contains the catalytic domain and numerous N-
glycosylation sites. ECEs cleave the
endothelia propeptide between Trp73 and Va174 producing the active peptide,
ET, which appears to function as a
ZO local rather than a circulating hormone (Rubanyi, G.M. & Polokoff, M.A.,
Pharmachological Reviews 46: 325-4I5
(1994). Thus ECE activity is a potential site of regulation of endothelia
production and a possible target for
therapeutic intervention in the endothelia system. By blocking ECE activity,
it is possible stop the production of ET-I
by inhibiting the conversion of the relatively inactive precursor, big ET-1,
to the physiologically active form.
ECE 2 is 64% identical to bovine ECE-2 at the amino acid level. ECE-2 is
closely related to ECE-1 (63%
identical, 80 % conserved), neutral endopeptidase 24.11 and the Kell blood
group protein. Bovine ECE-2 is a type
II membrane-bound metalIoproteinase localized in the traps-Golgi network where
it acts as an intracellular enzyme
converting endogenous big endothelia-1 imo active endothelia (Einoto, N. and
Yanangisawa, M., J: Biol. Chem. 270:
15262-15268 (1995). The bovine ECE-2 mRNA expression is highest in parts of
the brain, cerebral cortex,
cerebellum and adrenal medulla. It is expressed at lower levels in mymetrium,
testes, ovary, and endothelial cells.
Bovine ECE-2 and ECE-1 both are more active on ET 1 as a substrate compared to
ET-2 or ET-3, Emoto and
Yanangisawa, supra.
Human ECE-2 is 736 amino acids in length with a 31 residue amino-terminal
tail, a 23 residue transmembrane helix
and a 682 carboxy-terminal domain. It is 94% identical to bovine ECE-2 and
643b identical to human ECE-1. The
predicted transmembrane domain is highly conserved between the human and
bovine ECE-2 proteins and between
human ECE-1 and human ECE 2, as are the putative N-linked glycosylation sites,
Cys residues conserved in the
neutral endopeptidase 24.11 and the Kell blood group protein family and the
putative zinc binding motif. The
sequence suggests, that like other members of the NEP-ECE-Kell family, human
ECE-2 encodes a type II
transmembrane zinc-binding metalloproteinase, which, by exuapolation from what
is known about bovine ECE-2,
is an intracellular enzyme located within the secretory pathway which
processes endogenously produced big ET-1
while it is still in the secretory vesicles. Emoto and Yanangisawa, supra.
The expression pattern of ECE-2 differs from that observed for EC&1. Northern
blot analysis of mRNA
levels indicated low levels of expression of a 3.3 kb transcript in adult
brain (highest in the cerebellum, putamen,
rrtedulla and temporal lobe, and lower in the cerebral cortex, occipital lobe
and frontal lobe), spinal cord, lung and
pancreas and higher levels of a 4.5 kb uanscript in fetal brain and kidney.
The two transcript sizes probably
represent the use of alternative polyadenylation sites as has been observed
for bovine ECE 2 (Emoto and
Yaaangisawa, supra) and ECE-I (Xu et al., Cell 78: 473-485 (1994). PCR on cDNA
libraries indicated low levels
of expression in fetal brain, fetal kidney, fetal small intestine and adult
testis. Fetal liver, fetal lung and adult
pancreas were all negative.
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The endotheiin (ET) family of peptides have potent vascular, cardiac and renal
actions which may be of
pathophysiological importance in many human disease states. ET 1 is expressed
as an inactive 212 amino acid
prepmpeptide. The prepropeptide is first cleaved at Arg52-Cys53 and Arg92-
A1a93 and then the carboxy terminal
I:ys91 and Arg92 are trimmed from the protein to generate the propeptide big
ET-1. ECEs then cleave the propeptide
between Trp73 and Va174, producing the active peptide, ET, which appears to
function as a local rather than a
circulating hormone (Rubanyi and Polokoff, Pharma. R. 46: 325-415 (1994).
Endothelins may play roles in the path~ysiology of a number of disease states
including: 1) cardiovascular
diseases (vasospasm, hypertension, myocardial ischemia; reperfusion injury and
acute myochardial infarction, stroke
(cerebral ischemia), congestive heart failure, shock, atherosclerosis,
vascular thickening); 2) kidney disease (acute
and chronic renal faihire, glometulonephritis, cirrhosis); 3) lung disease
(bronc:hial asthma, pulmonary hypertension);
4) gastrointestinal disorders (gastric ulcer, inflammatory bowel diseases); 5)
reproductive disorders (premature labor,
dysmertorhea, preeclampsia) and 6) carcinogenesis. Rubanyi & Polokoff, supra.
'
Diseases can be evaluated for the impact of ET upon them by exanuning: I)
increased production of ETs;
2) increased reactivity to ETs; andlor 3) efficacy of an ET receptor
antagonist, antibody or ECE inhibitor. Response
to the previous criteria suggest that ETs likely play roles in cerebral
vasospasm following subarachnoid hemorrhage,
hypertension (fulminant/complications), acute renal failure and congestive
heart failure. While inhibitors of ET
production or activity have not been used in models of coronary vasospasm,
acute myocardial infarction, and
attarrosclerosis, they do have elevated ET levels and increase reactivity to
ETs. Shock and pulmonary hypertension
also exhibit elevated ET levels (Rubanyi and Polokoff, supra). Inhibition .of
ECEs in these conditions may be of
therapeutic value.
The expression pattern of ECE-2 differs from that observed for ECE-1. ECE-2
was observed at low levels in the
adult brain, lung and pancreas and higher levels in feral brain and kidney by
Northern blot analysis (Fig. 8). PCR
revealed low levels of expression in additional tissues: fetal lung, fetal
small intestine and adult testis. Fetal liver was '
negative. A similar pattern was reported for bovine ECE 2 (Emoto and
Yanangisawa, supra): It is expressed in brain
tissues (cerebral cortex, cerebellum and adrenal medulla), myotnetriutn and
testis, and in low levels in ovary and very
low levels in many other tissues. Bovine ECE-1 (Xu et al, supra) is more
widely and more abundantly expressed.
It is observed in vascular endothelial cells of most organs and in sorry
parenchymal cells. With the exception for
brain, bovine ECE 2 mRNA was present at lower levels than ECE-1. Applicants
believe ECE-2 to be a particularly
good target for the therapeutic intervention for diseases such as cerebral
vasospasm following subarachnoid
hemorrhage and stroke.
92. Anti-PRO Polv~eptide Antibodies
The present invention further provides and-PRO polygeptide antibodies.
Exemplary antibodies include
polyclonal, mo~clonal, humanized, bispecific, and heteroconjugate antibodies.
A. Polyclonai Antibodies
The anti~"RO polypeptide antibodies may comprise polyclonal antibodies.
Methods of preparing polyclonal
antibodies are known to the skilled artisan. Polyclonai antibodies can be
raised in a mammal, for example, by one
or more injections of an irrnttunizing agent and, if desired, an adjuvant.
Typically, the immunizing agent andlor
adjuvant will be in~cted in the rnarmrtal by multiple subcutaneous or
intraperitoneal injections. The ~ agent
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WO 99/46281 PCT/US99105028
tray include the PRO poiypeptide or a fusion protein thereof. It may be useful
to conjugate the immunizing agent
to a protein known to be immunogenic in the mammal being immunized. Examples
of such itnmunogettic proteins
include but are not limited to keyhole limpet hemocyanin, serum albumin,
bovine thyroglobulin, and soybean trypsin
inhibitor. Examples of adjuvants which may be employed include Freund's
complete adjuvant and MPL-TDM
adjuvant (monophosphoryl Lipid A, synthetic trehalose dicorynomycolate). The
immunization protocol may be
selected by one skilled in the art without undue experimentation.
B. Monoclonal Antibodies
The anti-PRO polypeptide antibodies may, alternatively, be monoclonal
antibodies. Monoclonal antibodies
tray be prepared using hybridoma methods, such as those described by Kohler
and Milstein, Nature, 256:495 (1975).
i0 In a hybridoma method, a mouse, hamster, or other appropriate host animal,
is typically immunized with an
immunizing agent to elicit lymphocytes that produce or are capable of
producing antibodies that will specifically bind
to the immunizing agent. Alternatively, the lymphocytes may be immunized in
vitro.
The immunizing agent will typically include the PRO polypeptide of interest or
a fusion protein thereof.
Generally, either peripheral blood lymphocytes ("PBIs") are used if cells of
human origin are desired, or spleen cells
or lymph node cells are used if non-human mammalian sources are desired. The
lymphocytes are then fused with
an immortalized cell line using a suitable fusing agent, such as polyethylene
glycol, to form a hybridoma cell (coding,
Monoclonal Antibodies: Princ't~les and Practice, Academic Press, (1986) pp. 59-
103j. Immortalized cell lines are
usually transformed mammalian cells, particularly myeloma cells of rodent,
bovine and human origin. Usually, rat
or mouse myeloma cell lines are employed. The hybridoma cells may be cultured
in a suitable culture medium that
preferably contains one or more substances that inhibit the growth or survival
of the unfused, immortalized cells.
For example, if the parental cells lack the enzyme hypoxanthine guanine
phosphoribosyl uansferase (HGPRT or
HPRT), the culture medium for the hybridomas typically will include
hypoxanthine, aminopterin, and tltymidine
("HAT medium"), which substances prevent the growth of HGPRT-deficient cells.
Preferred immortalized cell lines are those that fuse efficiently, support
stable high level expression of
antibody by the selected antibody-producing cells, and are sensitive to a
medium such as HAT medium. More
preferred immortalized cell lines are marine myeloma lines, which can be
obtained, for instance, from the Salk
Institute Cell Distribution Center, San Diego, California and the American
Type Culture Collection, Rockville,
Maryland. Human myeloma and mouse-human heteromyelotna cell lines also have
been described for the production
of human monoclonal antibodies [Kozbor, J. Immunol., 133:3001 (1984); Brodeur
et al., Monoclonal Antibody
Production Techniques and Applications, Marcel Dekker, Inc., New York, (1987)
pp. 51-63].
The culture medium in which the hybridoma cells are cultured can then be
assayed for the presence of
monoclonal antibodies directed against the PRO polypeptide of interest.
Preferably, the binding specificity of
monoclonal antibodies produced by the hybridoma cells is determined by
immunoprecipitation or by an in vitro
binding assay, such as radioirntnunoassay (RIA) or enzyme-linked
immunoabsorbent assay (ELISA). Such techniques
and assays are known in the art. The binding affinity of the monoclonal
antibody can, for example, be determined
by the Scatchard analysis of Munson and Pollard, Anal. Biocltern., 107:220
(1980).
After the desired hybridoma cells are identified, the clones may be subcloned
by limiting dilution procedures
and grown by standard methods [coding, surral. Suitable culture media for this
purpose .include, for example,
Dulbecco's Modified Eagle's Medium and RPMI-1640 medium. Alternatively, the
hybtidotna cells tray be grown
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WO 99/4628T PCT/US99/05028
in vivo as ascites in a mammal.
The monoclonal antibodies secreted by the subclones may be isolated or
purified from the culture medium
or ascites fluid by conventional immunoglobulin purification procedures such
as, for example, protein A-Sepharose,
hydroxylapatite chromatography, gel electrophoresis, dialysis, or affinity
chromatography.
T'he monoclonal antibodies may also be made by recombinant DNA methods, such
as those described in
U.S. Patent No. 4,816,567. DNA encoding the monoclonal antibodies of the
invention can be readily isolated and
sequenced using conventional procedures (e.g., by using oligonucleotide probes
that are capable of binding
specifically to genes encoding the heavy and light chains of marine
antibodies). The hybridoma cells of the invention
serve as a preferred source of such DNA. Once isolated, the DNA may be placed
into expression vectors, which
are then transfected into host cells such as simian COS cells, Chinese hamster
ovary (CHO) cells, or myeloma cells
that do not otherwise produce immunoglobulin protein, to obtain the synthesis
of monoclonal antibodies in the
recombinant host cells. The DNA also may be modified, for example, by
substituting the coding sequence for human
heavy and light chain constant domains in place of the homologous marine
sequences [U.S. Patent No. 4,816,567;
Morrison et al:, su r or by covalently joining to the immunoglobulin coding
sequence all or part of the coding
sequence for a non-immunogiobulin polypeptide. Such a non-immunoglobulin
polypeptide can be substituted for the
constant domains of an antibody of the invention, or can be substituted for
the variable domains of one antigen-
combining site of an antibody of the invention to create a chimeric bivalent
antibody.
The antibodies may be monovalent antibodies. Methods for preparing manovalent
antibodies are well known
in the art. For example, one method involves recombinant expression of
immunoglobulin light chain and modified
heavy chain. The heavy chain is truncated generally at any point in the Fc
region so as to prevent heavy chain
crosslinking. Alternatively, the relevant cysteine residues are substituted
with another amino acid residue or are
deleted so as to prevent crosslinldng.
In vitro methods are also suitable for preparing monovalent antibodies.
Digestion of antibodies to produce
fragments thereof, particularly, Fab fragments, can be accomplished using
routine techniques known in the art.
C. Humanized Antibodies
The anti-PRO polypeptide ann'bodies of the invention may further comprise
humanized antibodies or human
antibodies. Humanized forms of non-human (e.g., marine) antibodies are
chimeric immunoglobulins,
immunoglobutin chains or fragments thereof (such as Fv, Fab, Fab', F{ab')z or
other antigen-binding subsequences
of antibodies) which contain minimal sequence derived from non-human
immunoglobulin. Humanized antibodies
include human immunoglobulins (recipient antibody) in which residues from a
complementary determining region
(CDR) of the recipient are replaced by residues from a CDR of a non-human
species (donor antibody) such as mouse,
rat or rabbit having the desired specificity, affinity and capacity. In some
instances, Fv framework residues of the
human immunoglobulin are replaced by corresponding non human residues.
Humanized antibodies may also
comprise residues which are found neither in the recipient antibody nor in the
imported CDR or framework
sequences. In general, the humanized antibody will comprise substantially alll
of at least one; and typically two,
variable domains, in which all or substantially all of the CDR regions
correspond to those of a non-human
itnmunoglobulin and all or substantially all of the FR regions are those of a
human immunogiobulin consensus
sequence. The humanized antibody optimally also will comprise at least a
portion of an immunoglobulin constant
region (Fc), typically that of a human imtnttnoglobulin [Jones et al., Nature,
,~: 522-525 (1986); Riechmann et al.,
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WO 99/46281 PCT/US99105028
Nature, X32:323-329 (i988): and Presta, Curr. Op. Struct. Biol., x:593-596
(1992)].
Methods for humanizing non-human antibodies are well known in the art.
Generally, a humanized antibody
has otre or more amino acid residues introduced into it from a source which is
non-human. These non-human amino
acid residues are often referred to as "import" residues, which are typically
taken from an "import" variable domain.
Humanization can be essentially performed following the method of Winter and
co-workers [3ones et al., Nature, 21:
522-525 (1986); Riechrnann et al., Nature, X32:323-327 (1988); Verhoeyen et
al., Science, x:1534-1536 (1988)],
by substituting rodent CDRs or CDR sequences for the corresponding sequences
of a human antibody. Accordingly,
such "humanized" antibodies are chimeric antibodies (U.S. Patent No.
4,816,567), wherein substantially less than
an intact human variable domain has been substituted by.the corresponding
sequence from a non-human species. In
practice, humanized antibodies are typically human antibodies in which some
CDR residues and possibly some FR
residues are substituted by residues from analogous sites in rodent
antibodies.
Human antibodies can also be produced using various techniques known in the
art, including phage display
libraries [Hoogenboom and Winter, J. Mol. Biol., 2:381 (1991); Marks et al.;
J. Mol. Biol., 222:581 (1991)]. The
techniques of Cole et al. and Boerner et al. are also available for the
preparation of human monoclonal antibodies
(Cole et al., Monoclonal Aruibodies and Cancer Therapy, Alan R. Liss, p. 77
(1985) and Boerner et al., J. Immunol.,
14111:86-95 (1991)].
D. Bis~ecific Antibodies
Bispecific antibodies are monoclonal, preferably human or humanized,
antibodies that have binding
specificities for at least two different antigens. 1n the present case, one of
the binding specificities is for the PRO
polypeptide, the other one is for any other antigen, and preferably for a cell-
surface protein or receptor or receptor
subutvt.
Methods for making bis~cific antibodies are known in the art. Traditionally,
the recombinant production
of bispecific antibodies is based on the co-expression of two immunoglobulin
heavy-chainllight-chain pairs, where
the two heavy chains have different specificities [Miistein and Cuello,
Nature, 30:537-539 (1983)]. Because of the
random assortment of immunoglobulin heavy and light chains, these hybridomas
(quadromas) produce a potential
mixture of ten different antibody molecules, of which only one has the correct
bispecific structure. The purification
of the correct molecule is usually accomplished by affinity chromatography
steps. Similar procedures are disclosed
in WO 93108829, published 13 May 1993, and in Traunecker et al., EMBD J.,
10:3655-3659 (1991).
Antibody variable domains with the desired binding specificities (antibody-
antigen combining sites) can be
fused to immunoglobutin constant domain sequences. The fusion preferably is
with an imtrnmoglobutin heavy-chain
constant domain, comprising at least part of the hinge, CH2, and CH3 regions.
It is preferred to have the first heavy
chain constant region (CH1) containing the site necessary for light-chain
binding present in at least one of the fusions.
DNAs encoding the immunoglobulin heavy-chain fusions and, if desired, the
immunoglobulin light chain, are inserted
into separate expression vectors, and are co-transfected into a suitable host
organism. For fiu~ther details of
generating bispecific antibodies see, for example, Suresh et al., Methods in
Enrymology, ,x..,21:210 (1986).
E. HeteroconiuQate Antibodies
Heteroconjugate antibodies are also within the scope of the present invention.
Heteroconjugate antibodies
are composed of two covalently joined antibodies. Such antibodies have, for
example, been proposed to target
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WO 99146281 PCT/US99105028
immune system cells to unwanted cells (U.S. Patent No. 4;676;980), and for
treatment of HIV infection (WO
91/00360; WO 921200373; EP 03089]. it is contemplated that the antibodies may
be prepared in vitro using known .
methods in synthetic protein chemistry, including those involving crosslinking
agents. For example, immunotoxins
may be constructed using a disulfide exchange reaction or by fornvng a
thioether bond. Examples of suitable reagents
for this purpose include iminothiolate and methyl-4-mercaptoburyrimidate and
those disclosed, for example, in U.S.
Patent No. 4,676;980.
93. Uses for Anti-PRO Polypeptide Antibodies
The anti-PRO polypeptide antibodies of the invention have various utilities.
For example, anti-PRO
polypeptide antibodies may be used in diagnostic assays for a PRO polypeptide,
e.g., detecting its expression in
specific cells, tissues, or serum. Various diagnostic assay techniques known
in the art may be used, such as
competitive binding assays, direct or indirect sandwich assays and
immunoprecipitation assays conducted in either
heterogeneous or homogeneous phases [Zola, Monoclonal Antibodies: A Manual of
Techniques, CRC Press, lnc.
(1987) pp. 147-158). The antibodies used in the diagnostic assays can be
labeled with a detectable moiety. The
detectable moiety should be capable of producing, either directly or
indirectly, a detectable signal. For example, the
detectable moiety may be a radioisotope, such as 'H, '° C,'2 P,'S S,
ot"~ I, a fluorescent or chemilumineseent
compound, such as fluorescein isothiocyanate, rhodamine, or luciferin, or an
enzyme, such as alkaline phosphatase,
beta-galactosidase or horseradish peroxidase. Any method known in the a.rt for
conjugating the antibody to the
detectable moiety may be employed, including those methods described by Hunter
et al., Nature, 144:945 (1962);
David et al., Biochemistry, 13:1014 (1974); Pain et al., J. Immunol. Meth.,
gQ:219 (1981); and Nygren, J.
Histochem. and Cytochem., X0_:407 (1982).
Anti-PRO polypeptide antibodies also are useful for the affinity purification
of PRO polypeptide from
recombinant cell culture or natural sources. In this process, the antibodies
against the PRO polypeptide are
immobilized on a suitable support, such a Sephadex resin or filter paper,
using methods well known in the art. The
immobilized antibody then is contacted with a sample containing the PRO
polypeptide to be purified, and thereafter
the support is washed with a suitable solvent that will remove substantially
all the material in the sample except the
PRO polypeptide, which is bound to the immobilized antibody. Finally, the
support is washed with another suitable
solvent that will release the PRO polypeptide from the antibody.
Anti-Toll receptor (i.e., anti-PR0285 and anti-PR0286 antibodies) may also be
useful in blocking the
biological activities of the respective Toll receptors. The primary function
of the family of Toll receptors is believed
to be to act as pathogen pattern recognition receptors sensing the presence of
conserved molecular pattern present
on microbes. Lipopolysaccharides (LPS, also known as endotoxins), potentially
lethal molecules produced by
various bacteria, bind to the lipopolysaccharide binding protein (LBP) in P.he
blood. The complex formed then
activates a receptor latown as CD14. There is no consensus in the art about
what happens next. According to a
hypothesis, CD14 does not directly instruct macrophages to produce cytokines,
cell adhesion proteins and enrymes
involved in the production of lower molecular weight proinflammatory
mediators, rather enables LPS to activate a
second receptor. Alternatively, it has been suggested that LPS may activate
certain receptors directly, without heap
from LBP or CD14. The data disclosed in the present application indicate that
the human toll-like receptors are
signaling receptors that are activated by LPS in an LBP and CD14 responsive
manner. As this mechanism, under
pathophysiologic conditions can lead to an often fatal syndrome called septic
shack, anti-Toll receptor antibodies (just
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WO 99146281 PCT/US99/05028
as other Toll receptor antagonists) might be useful in the treatment of septic
shock. It is foreseen that the different
Toll receptors might recognize different pathogens, e.g., various strains of
Gram-negative or Gram-positive bacteria.
Accordingly, in certain situations, combination therapy with a mixture of
antibodies specifically binding different Toll
receptors, or the use of bispecific anti-Toll antibodies may be desirable.
It is specifically demonstrated that anti-huTLlt2 antibodies are believed to
be specifically useful in blocking
the induction of this receptor by LPS. As it has been shown that LPS exposure
can lead to septic shock (Parrillo,
N. Erg!. J. Med. 328, 1471-1477 (1993]), anti-huTLR2 antibodies are
potentially useful in the ueatment of septic
shock.
The foregoing therapeutic and diagnostic uses listed in connection with the
anti-Toll receptor antibodies are
also applicable to.other Toll antagonists, i.e., other molecules (proteins,
peptides, small organic molecules, etc.) that
' block Toll receptor activation and/or signal transduction mediated by Toll
receptors.
In view of their therapeutic potentials, the Toll proteins (including variants
of the native Toll homologues),
and their agonists and antagonists (including but not limited to anti-Toll
antibodies) are incorporated in compositions
suitable for therapeutic use. Therapeutic compositions are prepared for
storage by mixing the active ingredient
having the desired degree of purity with optional physiologically acceptable
carriers, excipients or stabilizers
(Remington's Pharmaceutical Sciences 16th Edition, Osol, A. Ed. 1980) in the
form of lyophilized formulations or
aqueous solutions. Acceptable carriers, excipients or stabilizers are nontoxic
to recipients at the dosages and
concentrations employed, and include buffers such as phosphate, citrate and
other organic acids; antioxidants
including ascorbic acid; low molecular weight {less than about 10 residues)
polypeptides; proteins, such as serum
albumin, gelatin or immunoglobulins; hydrophilic polymers such as
polyvinylpyrrolidone, amino acids such as
glycir~, glutamitte, as~ragine, argini~ or lysine; monosaccharides,
disaccharides and other carbohydrates including
glucose, mannose, or dextrins; cheiating agents such as EDTA; sugar alcohols
such as manttitol or sorbitol; salt-
forming counterions such as sodium; and/or nonionic surfactants such as Tween,
Pluronics or PEG.
The active ingredients may also be entrapped in microcapsules prepared, for
example, by coacervation
techniques or by interfacial polymerization, for example,
hydroxymethylcellulose or gelatin-microcapsules and poly-
(rnethylmethacylate) microcapsules, respectively), in colloidal drug delivery
systems (for example, liposomes,
albumin microspheres, microemulsions, nano-particles and nanocapsules) or in
macroemulsions. Such techniques
are disclosed in Reming~n's Pharmaceutical Sciences, supra.
The formulations to be used for in vivo administration must be sterile. This
is readily accomplished by
filtration through sterile filtration membranes, prior to or following
lyophilization and reconstitution.
Therapeutic compositions herein generally are placed into a container having a
sterile access port, for
example, an intravenous solution bag or vial having a stopper pierceable by a
hypodermic injection needle.
The route of administration is in accord with known methods, e.g. injection or
infusion by intravenous,
intraperitoneal, intracerebral, intramuscular, intraocular, intraarterial or
intralesional routes, topical administration,
or by sustai~d release systems.
Suitable examples of sustained release preparations include semipenneable
polymer matrices in the form
of shaped articles, e.g. films, or microcapsules. Sustained. release matrices
include polyesters, hydrogels,
polylactides t',U.S. Patent 3,773,919, EP 58,481), copolymers of L-glutamic
acid and gamma ethyl-L-glutamate (L1.
Sidman gt al., ~iopolvmers 2,,~ (1): 547-556 [1983]), poly (2-hydroxyethyl-
methacrylate) (R. Larger, e,~ ~., J.
Biomed. Mates. Res. ~: 167-277 [1981] and R. Larger, Cbem. Tech. 12: 98-105
[1982]), ethylene vinyl acetate
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CA 02421319 2003-03-25
WO 99146281 PCT/US99105028
(R. Larger e_t g11., Id.) or poly-D-(-)-3-hydroxybutyric acid (EP 133,988).
Sustained release compositions also include
liposomes. Liposomes containing a molecule within the scope of the present
invention are prepared by methods
known per se: DE 3,218,121; Epstein e1 al., ProcNatl. Acad. Sci. tJSA, $2_:
3688-3692 (1985); Hwang ~ g~.,
groc. Nat,. Acad. Sci. USA 77: 4030-4034 (1980); EP 523?2; EP 36676A; EP
88046; EP 143949; EP 142641;
Japanese patent application 83-118008; U.S. patents 4,485,045 and 4,544,545;
and EP 102,324. Ordinarily the
liposomes are of the small (about 200-800 Angstroms) unilamelar eype in which
the lipid content is greater than about
30 mot. % cholesterol, the selected proportion being adjusted for the optimal
NT-4 therapy.
An effective amount of the active ingredient will depend, for example, upon
the therapeutic objectives, the
route of administration, and the condition of the patient. Accordingly, it
will be necessary for the therapist to titer
the dosage and modify the route of administration as required to obtain the
optimal therapeutic effect. A typical daily
dosage might range from about 1 ~tglkg to up to 100 mglkg or more, depending
on the factors mentioned above.
Typically, the clinician will administer a molecule of the present invention
until a dosage is reached that provides the
required biological effect. The progress of this therapy is easily monitored
by conventional assays.
The following examples are offered for illustrative purposes only, and are not
intended to limit the scope
of the present invention in any way.
All patent and literature references cited in the present specification are
hereby incorporated by reference
in their entirety.
EXAMPLES
Commercially available reagents referred to in the examples were used
according to manufacturer's
instructions tuiless otherwise indicated. The source of those cells identified
iii the following examples, and throughout
the specification, by ATCC accession numbers is the American Type Culture
Collection, Rockville, Maryland.
EXAMPLE 1: xt ell I o in H mol creenin to Ide-ti V ve Pol a 'd c A n o '
Therefor
The exu~acelltilar domain (ECD) sequences (including the secretion signal
sequence, if any) from about 950
known secreted proteins from the Swiss-Prot public database were used to
search EST databases. The EST databases
included public databases (e.g., Dayhoff, GenBank), and proprietary databases
(e.g. LIFESEQT"'', Ineyte
Pharmaceuticals, Palo Alto, CA). The search was performed using the computer
program BLAST or BLAST2
(Altschul and Gish, Methods j_n EmJrmolo>rv ~6ø: 460-480 (1996)) as a
comparison of the ECD protein sequences
to a 6 frame translation of the EST sequences. Those comparisons with a Blast
score of 70 (or in some cases 90) or
greater that did not encode known proteins were clustered and assembled into
consensus DNA sequences with the
program "phrap" (Phil Green; University of Washington, Seattle, WA;
(http:l/bowman.mbt.washington.edulphrap.docslphrap.html).
Using this exuacellular domain homology screen, consensus DNA sequences were
assembled relative to
the other identified EST sequences using phrap: in addition, the consensus DNA
sequences obtained were often (but
not always) extended using repeated cycles of BLAST and phrap to extend the
consensus sequence as far as possible
using the sources of EST sequences discussed above.
Based upon the consensus sequences obtained as described above,
oligonucleotides were then synthesized
and used to identify by PCR a cDNA library that contained the sequence of
interest and for use as probes to isolate
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a clone of the full-length coding sequence for a PRO polypeptide. Forward (.f)
and reverse (.r) PCR primers
generally range from 20 to 30 nucleotides and are often designed to give a PCR
product of about 100-1000 by in
length. The probe (.p) sequences are typically 40-55 by in length. In some
cases, additional oligonucleotides are
synthesized when the consensus sequence is greater than about 1-1.Skbp. In
order to screen several libraries for a
full-length clone, DNA from the libraries was screened by PCR amplification,
as per Ausubel et al., Current
Protocols in Molecular BioloQV, with the PCR primer pair. A positive library
was then used to isolate clones
encoding the gene of interest using the probe oligonucleotide and one of the
primer pairs.
The cDNA libraries used to isolate the cDNA clones were constructed by
standard methods using
commercially available reagents such as those from lnvitrogen, San Diego, CA.
The cDNA was primed with oligo
dT containing aNod site, linked with blunt to SaII hemikinased adaptozs,
cleaved with Notl, sized appropriately by
: gel electrophoresis, and cloned in a defined orientation into a suitable
cloning vector (such as pRKB or pRKD;
pRKSB is a precursor of pRKSD that does not contain the SftI site; see, Holmes
et al., Science, 25:1278-1280
(1991)) in the unique Xhol and Notl sites.
EXAMPLE 2: Isolation of cDNA clones by Amylase Screening
1. )'reparation of oligo dT primed cDNA library
mRNA was isolated from a human tissue of interest using reagents and protocols
from Invitrogen, San
Diego, CA (Fast Track 2). This RNA was used to generate an oligo dT primed
cDNA library in the vector pRKSD
using reagents and protocols from Life Technologies, Gaithersburg, MD (Super
Script Plasmid System). In this
procedure, the double stranded cDNA was sized to greater than 1000 by and the
SalI/Nod Tinkered cDNA was cloned
into XhoIINotI cleaved vector. pRKSD is a cloning vector that has an spb
transcription initiation site followed by
an SfiI restriction enzyme site preceding the XhoIINotI cDNA cloning sites.
2. preparation of random primed cDNA library
A secondary cDNA library was generated in order to preferentially represent
the 5' ends of the primary
cDNA clones. Sp6 RNA was generated from the primary library {described above),
and this RNA was used to
generate a random primed cDNA library in the vector pSST-AMY.O using reagents
and protocols from Life
Technologies (Super Script Plasmid System, referenced above). In this
procedure the double stranded cDNA was
sized to 500-1000 bp, Tinkered with blunt to NotI adaptors, cleaved with SfiI,
and cloned into SfiIINotl cleaved
vector. pSST-AMY.O is a cloning vector that has a yeast alcohol dehydrogenase
promoter pseceding the cDNA
cloning sites and the mouse amylase sequence (the mature sequence without the
secretion signal) followed by the yeast
alcohol dehydrogenase terminator, after the cloning sites. Thus, cDNAs cloned
into this vector that are fused in
frame with amylase sequence will lead to the secretion of amylase from
appropriately transfected yeast colonies.
3, Trancfnrmation and Detection
DNA from the library described in paragraph 2 above was chilled on ice to
which was added
electrocompetent DNIOB bacteria (Life Technologies, 20 ml). The bacteria and
vector mixture was then
electroporated as recommended by the manufacturer. Subsequently, SOC media
(Life Technologies, 1 ml) was added
and the mixture was incubated at 37°C for 30 minutes. The transformants
were then plated onto 20 standard 150
mm LB plates containing ampicillin and incubated for 16 hours {37°C).
Positive colonies were scraped off the plates
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and the DNA was isolated from the bacterial pellet using standard protocols,
e.g. CsCI-gradient. The purified DNA
was then carried on to the yeast protocols below.
The yeast methods were divided into three categories: (1) Transformation of
yeast with the plasmidlcDNA
combined vector; (2} Detection and isolation of yeast clones secreting
amylase; and (3) PCR amplification of the
insert directly from the yeast colony and purification of the DNA for
sequencing and further analysis.
The yeast swain used was HD56-SA (ATCC-90785). This strain has the following
genotype: MAT alpha,
ura3-52, leu2-3, leu2-112, his3-11, his3-15, MAL+, SUC+, GAL''. Preferably,
yeast mutants can be employed that
have deficient post-translational pathways. Such mutants may have uanslocation
deficient alleles in sec7l, sec72,
sec62, with truncated sec71 being most preferred. Ahernatively, antagonists
(including antisense nucleotides andlor
Iigands) which interfere with the normal operation of these genes, other
proteins implicated in this pcast translation
pathway (e.g., SEC6lp, SEC72p, SEC62p, SEC63p, TDl 1p or SSAlp~lp) or the
complex formation of these proteins
may also be preferably employed in combination with the amylase-expressing
yeast.
Transformation was performed based on the protocol outlined by Gietz et al.,
Nucl. Acid. Rtes., ~l;1425
(1992). Transformed cells were tlten inoculated from agar into YEPD complex
media broth (100 ml) and grown
overnight at 30°C. The YEPD broth was prepared as described in Kaiser
et al., Methods in Yeast Genetics, Cold
Spring Harbor Press, Cold Spring Harbor, NY, p. 207 (1994). The overnight
culture was then diluted to about 2
x 106 cellslml (approx. ODD=0.1) into fresh YEPD broth (500 mt) and regrown to
1 x' IO cellslml (approx.
ODD=0.4-0.S).
The cells were then harvested and prepared for transformation by transfer into
GS3 rotor bottles in a Sorval
GS3 rotor at 5,000 rpm for 5 minutes, ~ supernatant discarded, and then
resuspended into sterile water, and
centrifuged again in 50 ml falcon tubes at 3,500 rpm in a Beckman GS-6KR
centrifuge. The supernatant was
discarded and the cells were subsequently washed with LiAclT'E (10 ml, 10 mM
Tris-HCI, 1 mM EDTA pH 7.5,
100 mM LiZOOCCH~), and resuspended into iaAclTE (2.5 ml).
Transformation took place by mixing the prepared cells (100 ~c1) with freshly
denatured single stranded
salmon testes DNA (Lofstrand Labs, Gaithersburg, MD) and transforming DNA {I
fig, vol. < IO ~l) in microfuge
tubes. The mixture was mixed briefly by vortexing, then 40% PEGITE (600 1l,
40% polyethylene glycol-4000, 10
mM Tris-HCI, 1 mM EDTA, 100 nnM LizOOCCH3, pH 7.5) was added. This mixture was
gently mixed and
incubated at 30°C while agitating for 30 minutes. The cells were then
heat shocked at 42°C for 15 minutes, and the
reaction vessel centrifuged in a microfuge at 12,000 rpm for 5-10 seconds,
decanted and resuspended into TE (500
1c1, 10 mM Tris ICI, 1 mM EDTA pH 7.5) followed by recentrifugation. The cells
were then diluted into TE (I ml)
and aliquots (200 u1) were spread onto the selective media previously prepared
in 150 mm growth plates (VWR).
Alternatively, instead of multiple small reactions, the transformation was
performed using a single, large
scale reaction, wherein reagent amounts were scaled up accordingly.
The selective media used was a synthetic complete dextrose agar lacking uracil
(SCD-Ura) prepared as
descn'bed in Kaiser et aL,1_ylethcxis in~east Genetic, Cold Spring Harbor
Press, Cold Spring Harbor, NY, p. 208-
2i0 (1994). Transformants were grown at 30°C for 2-3 days.
The detection of colonies secreting amylase was performed by including red
starch is the selective growth
media. Starch was coupled to the red dye (Reactive Red-120, Sigma) as per the
procedure described by Biely et al. ,
A~l- Biochem , 1~:i76-179 (1988). The coupled starch was incorporated into the
SCD-Ura agar plates at a final
concentration of 0.15 i6 (w/v), and was buffered with potassium phosphate to a
pH of 7.0 (50-100 mM final
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concentration).
The positive colonies were picked and streaked across fresh selective media
(onto 150 tnm plates) in order
to obtain well isolated and identifiable single colonies. WeII isolated single
colonies positive for amylase secretion
were detected by direct incorporation of red starch into buffered SCD-Ura
agar. Positive colonies were determined
by their ability to break down starch resulting in a clear halo around the
positive colony visualized directly.
4. Isolation of DNA by PCR Amplification
When a positive colony was isolated, a portion of it was picked by a toothpick
and diluted into sterile water
(30 p1) in a 96 well plate. At this time, the positive colonies were either
frozen and stored for subsequent analysis
or immediately amplified. An aliquot of cells (5 ~.I) was used as a template
for the PCR reaction in a 25 gel volume
i0 containing: 0.5 p.1 Klentaq (Clontech, Palo Alto, CA); 4.0 ~d 10 mM dNTP's
(Perkin Elmer-Cecus); 2.5 uI Kentaq
buffet (Clontech); 0.25 ~1 forward oligo 1; 0.25 u1 reverse oligo 2; 12.5 u1
distilled water. The sequence of the
forward oligonucleotide 1 was:
5'-TGTAAAACGACGGCCAGTTAAATAGACCTGCAATTATTAATCT-3' {SEQ ID N0:324)
The sequence of reverse oligonucleotide 2 was:
5'-CAGGAAACAGCTATGACCACCTGCACACCTGCAAATCCATT-3' (SEQ ID N0:325)
PCR was then performed as follows:
a. Denature 92°C, 5 minutes
b. 3 cycles of: Denature 92°C, 30 seconds
Anneal 59°C, 30 seconds
Extend 72°C, 50 seconds
c. 3 cycles of: Denature 92°C, 30 seconds
Anneal 57°C, 30 seconds
Extend 72°C, 60 seconds
d. 25 cycles of: Denature 92°C, 30 seconds
Anneal S5°C, 30 seconds
Extend 72°C, b0 seconds
e. Hold 4°C
The underlined regions of the oligonucleotides annealed to the ADH promoter
region and the amylase
region, respectively, and amplified a 307 by region from vector pSST-AMY.O
when no insert was present. Typically,
the first 18 nucleotides of the 5' end of these oligonucleotides contained
annealing sites for the sequencing primers.
Thus, the total product of the PCR reaction from an empty vector was 343 bp.
However, signal sequence-fused
cDNA resulted in considerably longer nucleotide sequences.
Following the PCR, an aliquot of the reaction (5 u1) was examined by agarose
gel electrophoresis in a 1 ~
agarose gel using a Tris-Borate-EDTA (TBE) buffering system as described by
Sambrook et al., supra. Clones
resulting in a single strong PCR product larger than 400 by were further
analyzed by DNA sequencing after
purification with a 96 Qiaquick PCR clean-up column (Qiagen Inc., Chatsworrh,
CA).
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FxAMPLE 3: Isolation of cDNA Clones Encodi~g,~uman 1'Rf,~213
A consensus sequence was obtained relative to a variety of EST sequences as
described in Example 1 above,
wherein the consensus sequence obtained is herein designated DNA2873S. Based
on the DNA28735 consensus
sequence, oligonucleotides were synthesized: 1) to identify by PCR a cDNA
library that contained the sequence of
interest, and 2) for use as probes to isolate a clone of the full-length
coding sequence for PR02I3.
A pair of PCR primers (forward and reverse) were synthesized:
forward PCR Erimer 5'-TGGAGCAGCAATATGCCAGCC-3' (SHQ ID N0:3)
reverse PCR primer 5'-TTTTCCACTCCTGTCGGGTTGG-3' (SEQ ID N0:4)
Additionally, a synthetic oligonucleotide hybridization probe was constructed
from the consensus DNA28735
sequence which had the following nucleotide sequence
hy~ri~dization probe
5'-GGTGACACTTGCCAGTCAGATGTGGATGAATGCAGTGCTAGG.AGGG-3' (SEQ ID NO:S)
In order to screen several libraries for a source of a full-length clone. DNA
from the libraries was screened
by PCR amplification with the PCR primer pair idemified above. A positive
library was then used to isolate clones
encoding the PR0213 gene using the probe oligonucleotide and one of the PCR
primers. RNA for construction of
the cDNA libraries was isolated from human fetal lung tissue.
DNA sequencing of the clones isolated as described above gave the full-length
DNA sequence for PR0213
jherein designated as UNQ187 (DNA30943-1163)] (SEQ ID NO:1) and the derived
protein sequence for PR0213.
The entire nucleotide sequence of UNQ187 (DNA30943-1163) is shown in Figure 1
(SEQ ID NO:1). Clone
UNQ187 (DNA30943-1163) contains a single open reading frame with an apparent
translational initiation site at
nucleotide positions 336-338 and ending at the stop codon at nucleotide
positions 1221-1223 (Figure 1). The
predicted polypeptide precursor is 295 antino acids long (Figure 2). Clone
UNQ187 (DNA30943-1163) has been
deposited with ATCC.
Analysis of the amino acid sequence of the full-length PR0213 polypeptide
suggests that a portion of it
possesses significant homology to the human growth arrest-specific gene 6
protein. More specifically, an analysis
of the Dayhoff database (version 35.45 SwissProt 35) evidenced significant
homology between the PR0213 amino
acid sequence and the following Dayhoff sequences, HSMHC3WSA 6 and B48089.
]AMPLE 4: Isolation~f~,DNA Clones Encoding~t PR0274
A consensus sequence was obtained relative to a variety of EST sequences as
described in Example 1 above,
wherein the consensus sequence obtained is herein designated DNA36469. Based
on the DNA36469 consensus
sequence, oligonucleotides were synthesized: 1) to identify by PCR a cDNA
library that contained the sequence of
interest, and 2) for use as probes to isolate a clone of the full-length
coding sequence for PR0274. ESTs proprietary
to Genentech were employed in the consensus assembly. The ESTs are shown in
Figures 5-7 and are herein
designated DNA17873, DNA36157 and DNA28929, respectively.
Pairs of PCR primers (forward and reverse) were synthesized:
forward PCR~primer ,~6469_fl,~ 5'-CTGATCCGGTTCTTGGTGCCCCTG-3' (SEQ ID NO:11)
forward PCR pr.~'meT ~ t~~469 fZ) 5'-GCTCTGTCACTCACGCTC-3' (SEQ ID N0:12)
f~ arw td PCR prim r (3=5469 f3) 5'-TCATCTCTTCCCTCTCCC-3' (SEQ ID N0:13)
forward PCR primer 4 (~ 69 f415'-CCTTCCGCCACGGAGTTC-3' (SEQ ID N0:14)
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reverse PCR primer I 36469.r1) S'-GGCAAAGTCCACTCCGATGATGTC-3' (SEQ ID NO:15)
reverse PC~2 primer 2 136469.f~ 5'-GCCTGCTGTGGTCACAGGTCTCCG-3' (SEQ ID N0:16)
Additionally, a synthetic oligonucleotide hybridization probe was constructed
from the consensus DNA36469
sequence which had the following nucleotide sequence
hybridization vrobe t36469.n1)
5'-TCGGGGAGCAGGCCTTGAACCGGGGCATTGCTGCTGTCAAGGAGG-3' (SEQ ID N0:17)
In order to screen several libraries for a source of a full-length clone, DNA
from the libraries was screened
by PCR amplification with the PCR primer pair identified above. A positive
library was then used to isolate clones
encoding the PR0274 gene using the probe oligonucleotide and one of the PCR
primers. RNA for construction of
the cDNA libraries was isolated from human fetal liver tissue (LIB229).
. DNA sequencing of the clones isolated as described above gave the full-
length DNA sequence for PR0274
[herein designated as UNQ241 (DNA39987-1184)j (SEQ ID NO:1) and the derived
protein sequence for PR0274.
The entire nucleotide sequence of UNQ241 (DNA39987-1184) is shown in Figure 3
(SEQ ID N0:6). Clone
UNQ241 (DNA39987-1184) contains a single open reading frame with an apparent
translational initiation site at
nucleotide positions 83-85 and ending at the stop codon at nucleotide
positions 1559-1561 (Figure 3). The predicted
polypeptide precursor is 492 amino acids long (Figure 4), has an estimated
molecular weight of about 54,241 daltons
and an estimated pI of about 8.21. Clone UNQ241 (DNA39987-1184) has been
deposited with ATCC and is assigned
ATCC deposit no. 209786.
Analysis of the amino acid sequence of the full-length PR0274 polypeptide
suggests that it possesses
significant homology to the Fn54 protein. More specifically, an analysis of
the Dayhoff database (version 35.45
SwissProt 35) evidenced significant homology between the PR0274 amino acid
sequence and the following Dayhoff
sequences, MMFN54S2 1, MMFN54S1l1, CELF48C1'8, CEF38B7 6, PRP3 RAT, INL3 PIG,
MTCY07A?_13,
YNAX_KLEAE, A47234 and HME2_MOUSE.
EXAMPLE 5: lsol tion of cDNA Clones Encoding Human PRO300
A consensus sequence was obtained relative to a variety of EST sequences as
described in Example 1 above,
wherein the consensus sequence obtained is herein designated DNA35930. Based
on the DNA35930 consensus
sequence, oligonucleotides were synthesized: 1) to identify by PCR a cDNA
library that contained the sequence of
interest, and 2) for use as probes to isolate a clone of the full-length
coding sequence for PR0300.
Forward and reverse PCR primers were synthesized:
forward PCR primer 1 (35930.f1) 5'-GCCGCCTCATCTTCACGTTCTTCC-3' (SEQ ID N0:20)
forwgrd PCR primer 2 (35930,,~2~ 5'-TCATCCAGCTGGTGCTGCTC-3' (SEQ ID N0:21)
forward PCR vrimer 3 (35930.f3) 5'-CTTCTTCCACTTCTGCCTGG-3' (SEQ ID N0:22)
forward PCR grimer 4 f35930.f4) 5'-CCTGGGCAAAAATGCAAC-3' (SEQ ID N0:23)
reverse PCR primer 1 (35930.r1) 5'-CAGGAATGTAGAAGGCACCCACGG-3' (SEQ ID N0:24)
reverse PCR primer 2~35930,;~~ S'-TGGCACAGATCTTCACCCACACGG-3' (SEQ ID N0:25)
Additionally, a synthetic oligonucleotide hybridization probe was constructed
from the consensus DNA35930
sequence which had the following nucleotide sequence
hybridization probe 135930.n11
5'-TGTCCATCATTATGCTGAGCCCGGGCGTGGAGAGTCAGCTCTACAAGCTG-3' (SEQ ID N0:26)
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In order to screen several libraries for a source of a full-length clone, DNA
from the libraries was screened
by PCR amplification with the PCR primer pair identified above. A positive
library was then used to isolate clones
encoding the PR0300 gene using the probe oligonucieotide and one of the PCR
primers. RNA for construction of
the cDNA libraries was isolated from human fetal kidney tissue.
DNA sequencing of the clones isolated as described above gave the full-length
DNA sequence for PR0300
(herein designated as UNQ263 (DNA40625-1189)] (SEQ ID NO:18) and the derived
protein sequence for PR0300.
The entire, nucleotide sequence of UNQ263 (DNA40625-1189) is shown in Figure 8
(SEQ ID N0:18).
Clone UNQ263 (DNA40b25-1189) contains a single open reading frame with an
apparent translationaI initiation site
at nucleotide positions 457 and ending at the stop colon at nucleotide
positions 1416-1418 (Figure 8). The
predicted polypeptide precursor is 457 amino acids long (Figure 9). Clone
UNQ263 (DNA40625-I 189} has been
deposited with ATCC and is assigned ATCC deposit no. 209788.
Analysis of the amino acid sequence of the full-length PR0300 polypeptide
suggests 'that portions of it
possess significant homology to the Diff 33 protein. More specifically, an
analysis of the Dayhoff database (version
35.45 SwissProt 35) evidenced significant homology between the PR0300 ;amino
acid sequence and the following
Dayhoff sequence, HSU49188 1.
EXAMPLE 6: Isolation of cDNA Clones E,~coding Human PR0~84
Two cDNA sequences were isolated in the amylase screen described in Example 2
and those cDNA
sequences are,herein designated DNA12982 (see Figure 12; human placenta-
derived) and DNA15886 (see Figure
13; human salivary gland-derived). The DNA12982 and DNA15886 sequences were
then clustered and aligned,
giving rise to a consensus nucleotide sequence herein designated DNA18832.
Based on the DNA18832 consensus sequence, oligonucleotide probes were
generated and used to screen
a human placenta library (LI$89) prepared as described in paragraph I of
Example 2 above. The cloning vector was -
pRKSB (pRKSB is a precursor of pRKSD that does not contain the SfiI site; see,
Holmes et al., Sc_ fence, ~53:1278-
1280 (1991)), and the cDNA size cut was less than 2800 bp.
PCR primers (forward and reverse) were synthesized:
~rward PCR p~ em r 1 (~$l~~t",f~ 5'-TCGTACAGTTACGCTCTCCC-3' (SEQ ID N0:31)
forward PCR vrimer 2 (18832 t~ 5'-CTTGAGGAGCGTCAGAAGCG-3' (SEQ ID N0:32)
reverse PCR prZmer ~8832.rt 5'-ATAACGAATGAAGCCTCGTG-3' (SEQ ID N0:33)
Additionally, a synthetic oIigonucieotide hybridization probe was constructed
from the DNAI8$32 sequence which
had the following nucleotide sequence
hvbridizatio0 probg (18832 b1
S'-GCTAATATCTGTAAGACGGCAGCTACAGCAGGCATCATTG-3' (SEQ ID N0:34)
In order to screen several libraries for a source of a full-length clone, DNA
from the libraries was screened
by PCR amplification with the PCR primer pairs identified above: A positive
library was then used to isolate clones
encoding the PR0284 gene using the probe oligonucleotide and one of the PCR
primers.
A full length clone was identified that contained a single open reading frame
with an apparent transiational
initiation site at nucleotide positions 167-169 and ending at the stop colon
found at nucleotide positions 1022-1024
(Figure Z0; SEQ ID NO:Z?). The predicted poiypeptide precursor is 285 amino
acids long, has a calculated
molecular weight of approximately 32,190 daltons and an estimated pI of
approximately 9.03. Analysis of the full-
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WO 9914b281 PCT/US99/05028
length PR0284 sequence shown in Figure l I (SEQ ID N0:28) evidences the
presence of the following: a signal
peptide from about amino acid 1 to about amino acid 24, transmembrane domains
from about amino acid 76 to about
amino acid 96 and from about amino acid I71 to about amino acid 195 and a
potential N-glycosylation site from about
amino acid 153 to about amino acid 156. Clone UNQ247 (DNA23318-1211) has been
deposited with ATCC on April
21, 1998 and is assigned ATCC deposit no. 209787.
Analysis of the amino acid sequence of the full-length PR0284 polypeptide
suggests that it possesses no
significant sequence similarity to any known protein. However, an analysis of
the Dayhoff database (version 35.45
SwissProt 35) evidenced some degree of homology between the PR0284 amino acid
sequence and the following
Dayhoff sequences, JQ0124, CELE04A4 5, AB006451~1, AF030162_l, IM23 YEAST,
571194, NIA_CUCMA,
IM17 YEAST, I50479 and HUMZFIiP'1.
EXAMPLE 7: Isolation of cDNA Clones Encoding HurJnan PR0296
A cDNA sequence isolated in the amylase screen as described in Example 2 above
was found, by BLAST
and FastA sequence alignment, to have sequence homology to a nucleotide
sequence encoding sarcoma-associated
protein SAS. This cDNA sequence is herein designated DNA23020 (see Figure 16).
The DNA23020 sequence was
IS then compared to a variety of expressed sequence tag (EST) databases which
included public EST databases (e.g.,
GenBank) and a proprietary EST DNA database (LIFESEQT"', Incyte
Pharmaceuticals, Palo Alto, CA) to identify
existing homologies. The homology search was performed using the computer
program BLAST or BLAST2 (Altshul
et al., Methods in Enz~tmoiogy 2~"ø6 :460-480 (1996)). Those comparisons
resulting in a BLAST score of 70 (or in
some cases 90) or greater that did not encode known proteins were clustered
and assembled into a consensus DNA
sequence with the program "phrap" (Phil Green, University of Washington,
Seattle, Washington;
http:llbozeman.mbt.Washington.edulphrap.docslphrap.html). The consensus
sequence obtained therefrom is herein
designated DNA35858. Two proprietary Genentech ESTs were employed in the
assembly wherein those EST
sequences are herein identified as DNA2I97I (Figure 17; SEQ ID N0:38) and
DNA29037 (Figure 18; SEQ ID
N0:39).
Based on the DNA35858 consensus sequence, oligonucleotide probes were
generated and used to screen
a human kidney library (LIB228) library prepared as described in paragraph 1
of Example 2 above. The cloning
vector was pRKSB (pRKSB is a precursor of pRKSD that does not contain the SfiI
site; see, Holines et al., Sci" epee,
25:1278-1280 (1991)), and the cDNA size cut was less than 2800 bp.
PCR primers (forward and reverse) were synthesized:
~rward PCR primer 1 135858.f115'-ACCCACGTCTGCGTTGCTGCC-3' (SEQ ID N0:40)
forward PCR primer 2 t35858.f1) 5'-GAGAATATGCTGGAGAGG-3' (SEQ ID N0:41)
reverse PCR~rimerS33858.r115'-AGGAATGCACTAGGATTCGCGCGG-3' (SEQ ID N0:42)
Additionally, a synthetic oligonucleotide hybridization probe was constructed
from the consensus DNA35858
sequence which had the following nucleotide sequence
hybridization probe f35858.v~1
5'-GGCCCCAAAGGCAAGGACAAAGCAGCTGTCAGGGAACCTCCGCCG-3' (SEQ ID N0:43)
In order to screen several libraries for a source of a full-length clone, DNA
from the libraries was screened
by PCR amplification with the PCR primer pair identified above. A positive
library was then used to isolate clones
encoding the PR0296 gene using the probe -oligonucleotide and one of the PCR
primers.
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A full length clone was identified that contained a single open reading frame
with an apparent translational
initiation site at nucleotide positions 174-176 and ending at the stop colon
found at nucleotide positions 786-788
(Figure 14; SEQ ID N0:35). The predicted polypeptide precursor is 204 amino
acids Long, has a calculated
molecular weight of approximately 22,147 daltons and an estimated pI of
approximately 8.37. Analysis of the full-
length PR0296 sequence shown in Figure 15 (SEQ ID N0:36) evidences the
presence of the following: a signal
peptide from about amino acid I to about amino acid 34 and transmembrathe
domains from about amino acid 47 to
about amino acid 63, from about amino acid ?2 to about amino acid 95 and from
about amino acid 162 to about amino
acid 182. Clone UNQ260 (DNA39979-1213) has been deposited with A'TCC on April
21, 1998 and is assigned
ATCC deposit no. 209789.
Analysis of the amino acid sequence of the full-length PR0296 polypeptide
suggests that it possesses
significant sequence similarity to the sarcoma-amplifed SAS protein, thereby
indicating that PR0296 may be a novel
SAS homolog. More specifically, an analysis of the Dayhoff database (version
35.45 SwissProt 3S) evidenced
significant homology between the PR0296 amino acid sequence and the following
Dayhoff sequences, I5839i,
GEN11,061, SSC2B04 1, HSU81031 2, CD63 RAT, CD63 MOUSE, CD63 HUMAN,
AF022813_I,
CD63 RABIT and C002 HUMAN.
IS
EXAMPLE 8: Isolation of cDN,A Clqnes Encoding Human PR0329
A consensus sequence was obtai~d relative to a variety of EST sequences as
described in Example 1 above,
wherein the consensus sequence obtained is herein designated DNA3S612. Based
on the DNA3S6I2 consensus
sequence, oligonucIeotides were synthesized: 1) to identify by PCR a cDNA
library that contained the sequence of
interest, and 2) for use as probes to isolate a clone of the full-length
coding sequence for PR0329.
PCR primers (forward and reverse) were synthesized:
forward PCR rp ime~l (35612 f1) 5'-TGGGCTGTGTCCTCATGG-3' (SEQ ID N0:46)
forward PCR primer 2 135612 f215'-TTTCCAGCGCCAATTCTC-3' (SEQ ID N0:47)
reverse PCR vrimer I 13,6,12 r() 5'-AGTTCTTGGACTGTGATAGCCAC-3' (SEQ ID N0:48)
reverse PCR primed 21356I2 r2) 5'-AAACTTGGTTGTCCTCAGTGGCTG-3' (SEQ ID N0:49)
Additionally, a synthetic oligoaucleotide hybridization probe was constructed
from the.consensus DNA3S612
sequence which had the following nucleotide sequence
hvbridi~ation_probe (3S6I2 nIl
S'-GTGAGGGACCTGTCTGCACTGAGGAGAGCAGCTGCCACACGGAGG-3' (SEQ ID NO:50)
In order to screen several libraries for a source of a full-length clone, DNA
from the libraries was screened
by PCR amplification with the PCR primer pairs identified above: A positive
library was then used to isolate clones
etxoding the PR0329 gene using the probe oIigonucleotide and one of the PCR
primers. RNA for constntction of
the cDNA libraries was isolated from (roman fetal liver tissue (LIB6).
DNA sof the clones isolated as described above gave the full-length DNA
sequence for PR0329
[herein designated as UNQ29I (DNA40594-1233)) (SEQ ID N0:44) and the derived
protein sequence for PR0329.
The entire nucleotide sequence of UNQ291 (DNA40594-1233) is shown in Figure 19
(SEQ ID N0:44).
Clone UNQ291 t'DNA40594-1233) contains a single open reading frame with an
apparent translational initiation site
at nucleotide positions 9-11 and ending at the stop colon at nucleotide
positions 1086-10$8 (Figure 19). The
predicted polypeptide precursor is 359 amino acids song (Figure 20). The full-
length PR0329 protein shown in
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Figure 20 has an estimated molecular weight of about 38,899 daltons and a pI
of about 5.21. Clone UNQ291
(DNA40594-1233) has been deposited with ATCC on February S, 1998 and is
assigned ATCC deposit no. 209617.
Analysis of the amino acid sequence of the full-length PR0329 polypeptide
suggests that it possesses
significant sequence similarity to a high affinity immunoglobulin F~ receptor
protein. More specifically, an analysis
of the Dayhoff database (version 35.45 SwissProt 35) evidenced significant
homology between the PR0329 amino
acid sequence and the following Dayhoff sequences, FCG1 HUMAN, FCGO HUMAN, P
891439, P 822549,
P R9I438, P W00859, P~R20811, P 822550, HUMCD6406 1 and FCG1 MOUSE.
EXAMPLE 9: Isolation of cDNA Clones Encoding Human PR0362
A consensus sequence was obtained relative to a variety of EST sequences as
described in Example 1 above,
wherein the consensus sequence obtained is herein designated DNA42257. Based
on the DNA42257 consensus .
sequence, oIigonucleotides were synthesized: 1 ) to identify by PCR a cDNA
library that contained the sequence of
interest, and 2) for use as probes to isolate a clone of the full-length
coding sequence for PR0362.
PCR primers (forward and reverse) were synthesized:
forward primerf42257.f1)5'-TATCCCTCCAATTGAGCACCCTGG-3' (SEQ
PCR 1 ID N0:53}
~rward primer142~57.t215'-GTCGGAAGACATCCCAACAAG-3' (SEQ ID
PCR 2 N0:54)
reverse timer 142257.r1)5'-CTTCACAATGTCGCTGTGCTGCTC-3' (SEQ
PCR 1 ID N0:55)
reverse primer(42257.r2)5'-AGCCAAATCCAGCAGCTGGCTTAC-3' (SEQ
PC~t_ 2 1D N0:56)
Additionally, a synthetic oligonucleotide hybridization probe was constructed
from the consensus DNA42257
sequence which had the following nucleotide sequence
hybridization probe 142257.n1)
5'-TGGATGACCGGAGCCACTACACGTGTGAAGTCACCTGGCAGACTCCTGAT-3' (SEQ ID N0:57)
In order to screen several libraries for a source of a full-length clone, DNA
from the libraries was screened'
by PCR amplification with the PCR primer pairs identified above. A positive
library was then used to isolate clones
encoding the PR0362 gene using the probe oligonucleotide and one of the PCR
primers. RNA for construction of
the cDNA libraries was isolated from human fetal brain tissue (LIB153).
DNA sequencing of the clones isolated as described above gave the full-length
DNA sequence for PR0362
[herein designated as UNQ317 (DNA45416-1251)] (SEQ ID N0:51) and the derived
protein sequence for. P80362.
The entire nucleotide sequence of UNQ317 (DNA45416-1251} is shown in Figure 21
(SEQ ID N0:51).
Clone UNQ317 (DNA45416-1251) contains a single open reading frame with an
apparent uanslational initiation site
at nucleotide positions 119-121 and ending at the stop codon at nucleotide
positions 1082-1084 (Figure 21}. The
predicted polypeptide precursor is 321 amino acids long (Figure 22). The full-
length PR0362 protein shown in
Figure 2 has an estimated molecular weight of about 35,544 dahons and a p1 of
about 8.51. Analysis of the full-length
PR0362 polypeptide as shown in Figure 22 evidences the presence of a
glycosaminoglycan attachment site at about
amino acid 149 to about amino acid 152 and a transmembratte domain from about
amino acid 276 to about amino acid
306. Clone UNQ317 (DNA45416-1251) has been deposited with ATCC on February 5,
1998 and is assigned ATCC
deposit no. 209620.
Analysis of the amino acid sequence of the full-length PR0362 polypeptide
suggests that it possesses
significant sequence similarity to the A33 antigen protein and the HCAR
protein. More specifically, an analysis of
the Dayhoff database (version 35.45 SwissProt 35) evidenced significant
homology between the PRO362 amino acid
1$$
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sequence and the following I?ayhoff seguences, AB00234I_l, HSU55258.-1,
HSC?NRCAM~1, RNU81037~1,
A33 HUMAN, P WI4158, NMNCAMRI_l, HST1TINN2_t, S71824_1 and HSU63041_l.
)jXAMPLE 14: Isolation of cDNA Clones Encoding Human PR03b3
A consensus sequence was obtained relative to a variety of EST sequences as
described in Example 1 above,
wherein the consensus sequence obtained is hereindesignated DNA42828. Based on
the DNA42828 consensus
sequence, oligonucleotides were synthesized: 1) to identify by PCR a cDNA
library that contained the sequence of
interest, and 2) for use as probes to isolate a clone of the full-length
coding sequence for PR0363.
A pair of PCR primers (forward and reverse) were synthesized:
forward PCR,pi'imer ~42828.f_l~ 5'-CCAGTGCACAGCAGGCAACGAAGC-3' (SEQ ID N0:60)
reverse PCR primer (428~8.r1) 5'-ACTAGGCTGTATGCCTGGGTGGGC-3' (SEQ ID N0:61)
Additionally, a synthetic oligonucieotide hybridization probe was constructed
from the consensus DNA42828
sequence which had the following nucleotide sequence
hybridization probe (42828 n!)
5'-GTATGTACAAAGCATCGGCATGGTTGCAGGAGCAGTGACAGGC-3' (SEQ ID N0:62)
In order to screen several libraries for a source of a full-length clone, DNA
from the libraries was screened
by PCR amplification with the FCR primer pair identified above. A positive
library was then used to isolate clones
encoding the PR03b3 gene using the probe oligonucleotide and one of the PCR
primers. RNA for construction of
the cDNA libraries was isolated from human fetal kidney tissue (LiB227).
DNA sequencing of the clones isolated as described above gave the full-length
DNA sequence for PR0363
[herein designated as UNQ318 (DNA45419-1252)] (SEQ ID N0:58) and tire derived
protein sequence for PR03b3.
The entire nucleotide sequence of UNQ318 (DNA45419-1252) us shown in Figure 23
(SEQ ID N0:58).
Clone UNQ318 (DNA45419-1252) contains a single open reading frame with an
apparent translational initiation site
at nucleotide positions 190-192 and ending at the stop codon at nucleotide
positions 1309-131 I (Figure 23). The
predicted polypeptide precursor is 373 amino acids long (Figure 24). The futi-
length PR0363 protein shown in
Figure 24 has an estimated molecular weight of about 41,281 daItons and a p1
of about 8.33. A transmernbrane
domain exists at amino acids 221 to 254 of the amino acid sequence shown in
Figure 24 (SEQ ID N0:59). The
PR0363 polypeptide also possesses at least two myelin PO protein domains from
about amino acids 15 to 56 and from
about amino acids 87 to l I6. . Clone UNQ318 (DNA45419-1252) has been
deposited with ATCC on February 5,
1998 and is assigned ATCC deposit no. 209616.
Analysis of the amino acid sequence of the full-length PR0353 polypeptide
suggests that it possesses
significant sequence similarity to the cell surface protein HCAR, thereby
indicating that PR0363 may be a novel
HCAR homolog. More specifically, an analysis of the Dayhoff database (version
35.45 SwissProt 35) evidenced
significant homology between the PR0363 atnino acid sequence and the following
Dayhoff sequences, HS46KDA_1,
HSU90716 1, MMCARH_l, MMCARHOM_l, MMU90715_l, A33 HUMAN, P_W14146, P_W14158,
A42632
and B42632.
FXA~1 ,: Isolation of cDNg,~Clonec Enco~l'~t Human PR"~868
A consensus sequence was obtaica~d relative to a variety of EST sequences as
described in Example 1 above,
wherein the consensus sequence obtained is herein designated DNA38133. Based
on the DNA38133 consensus
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sequence, oligonucleotides were synthesized: 1 ) to identify by PCR a cDNA
library that contained the sequence of
interest, and 2) for use as probes to isolate a clone of the full-length
coding sequence for PR0868.
A pair of PCR primers (forward and reverse) were synthesized:
forward PCR~rimer (38133.f115'-GTAGCAGTGCACATGGGGTGTTGG-3' (SEQ 1D N0:65)
reverse PCR primer (38133.r1) 5'-ACCGCACATCCTCAGTCTCTGTCC-3' (SEQ ID NO:b6)
Additionally, a synthetic oligonucleotide hybridization probe was constructed
from the consensus DNA38I33
sequence which had the following nucleotide sequence
hybridization probe 138133.x11
5'-ACGATGATCGCGGGCTCCCTTCTCCTGCTTGGATTCCTTAGCACCACCAC-3' (SEQ ID N0:67)
In order to screen several libraries for a source of a full-length clone, DNA
from the libraries was screened
by PCR amplification with the PCR primer pair identified above. A positive
library was then used to isolate clones
encoding the PR0868 gene using the probe oligonucleotide and one of the PCR
primers. RNA for construction of
the cDNA libraries was isolated from human fetal kidney tissue (LIB227).
DNA sequencing of the clones isolated as described above gave the full-length
DNA sequence for PRO868
(herein designated as UNQ437 (DNA52594-1270)) (SEQ ID N0:63) and the derived
protein sequence for PR0868.
The entire nucleotide sequence of UNQ437 (DNA52594-1270) is shown in Figure 25
(SEQ ID N0:63).
Clone UNQ437 (DNA52594-1270) contains a single open reading frame with an
apparent translational initiation site
at nucleotide positions 325-327 and ending at the stop codon at nucleotide
positions 2290-2292 (Figure 25). The
predicted polypeptide precursor is 655 amino acids long (Figure 26). The full-
length PR0868 protein shown in
Figure 26 has an estimated molecular weight of about 71,845 daltons and a pI
of about 8.22. Analysis of the full-
length PR0868 polypeptide sequence demonstrates the presence of conserved
cysteine-containing domains from about
amino acid 66 to about amino acid 78 and from about amino acid 123 to about
amino acid 134 of the sequence shown
in Figure 26 (SEQ ID N0:3), a TNFR death domain from about amino acid 85 to
about amino acid 110, a
FASA mouse death domain block from about amino acid 159 to about amino acid
175 and a transmembrane domain
from about amino acid 347 to about amino acid 375. Clone UNQ437 (DNA52594-
1270) has been deposited with
ATCC on March 17, 1998 and is assigned ATCC deposit no. 209679
Analysis of the amino acid sequence of the full-length PR0868 polypeptide
suggests that it possesses
significant sequence similarity to the tumor necrosis factor receptor protein,
thereby indicating that PR0868 may be
a novel meraber of the tutnor necr~is factor receptor family. More
specifically, an analysis of the Dayhoff database
(version 35.45 SwissProt 35) evidenced significant homology between the PR0868
amino acid sequence and the
following Dayhoff sequences, RNU94330_l, P 899933, P 899945, P 899950,
HSU94332 1, CD40 HUMAN,
S63368_l, T'NRZ HUMAN, MVU87844 1 AND CVU87837_l.
EX~iPLE 12: Isolation of cDNA Clones Encoding Human PR0382
A consensus sequence was obtained relative to a variety of EST sequences as
described in Example 1 above,
wherein the consensus sequence obtained is herein designated DNA30892. Based
on the DNA30892 consensus
sequence, oligonucleotides were synthesized: 1) to identify by PCR a cDNA
library that contained the sequence of
interest, and 2) for use as probes to isolate a clone of the full-length
coding sequence for PR0382.
A pair of PCR primers (forward and reverse) were synthesized:
forward PCR primer 5'-TGACATCGCCCTTATGAAGCTGGC-3' (SEQ ID N0:70)
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reverse PCR primer 5'-TACACGTCCCTGTGGTTGCAGATC-3' (SEQ ID N0:71)
Additionally, a synthetic oligonucleotide hybridization probe was constructed
from the consensus DNA30892
sequence which had the following nucleotide sequence
bKt~idization probe
5'-CGTTCAATGCAGAAATGATCCAGCCTGTGTGCCTGCCCAACTCTGAAGAG-3' (SEQ ID N0:72)
In order to screen several libraries for a source of a foil-length clone; DNA
from the libraries was screened
by PCR amplification with the PCR primer pair identified above. A positive
library was then used to isolate clones
encoding the PR03$2 gene using the probe oligonucleotide and one of the PCR
primers. RNA for construction of
the cDNA libraries was isolated from human fetal kidney tissue (LIB227).
DNA sequencing of the clones isolated as described above gave the ifull-length
DNA sequence for PR0382
jherein designated as UNQ323 (DNA45234-1277)] (SEQ ID N0:68) and the derived
protein sequence for PR0382.
The entire nucleotide sequence of UNQ323 (DNA45234-I277) is .shown in Figure
27 (SEQ ID N0:68).
Clone UNQ323 (DNA45234-1277) contains a single open reading frame with an
apparent translational initiation site
at nucleotide positions 126-128 and ending at the stop codon at nucleotide
positions 1485-1487 (Figure 27). The
predicted polypeptide precursor is 453 amino acids long {Figure 28). The full-
length PR0382 protein shown in
Figure 28 has an estimated molecular weight of about 49,334 daltons and a pI
of about 6.32. Analysis of the native
PR0382 amino acid sequence shown in Figure 28 (SEQ ID N0:69) indicates the
presence of a putative
transmembrane domain from about amino acid 240 to about amino acid 284, a
putative signal peptide at about amino
acid 1 to about amino acid 20, a putative apple domain at about amino acid 386
to about amino acid 4I9, a putative
Kringle domain at about amino acid 394 to about amino acid 406 and a histidine-
containing protease active site at
about amino acid 253 to about amino acid 258. Clone UNQ323 (DNA45234-1277) has
been deposited with ATCC
on March 5, 1998 and is assigned ATCC deposit no. 209654.
Analysis of the amino acid sequence of the full-length PR0382 polypeptide
suggests that it possess
significant homology to serine protease proteins, thereby indicating that
PR0382 may be a novel serine protease.
Specifically, an analysis of the Dayhoff database (version 35.45 SwissProt 35)
evidenced significant homology
ZS between the PR0382 amino acid sequence and the following Dayhoff sequences,
HSU75329 I, ENTK_MOUSE,
HEPS HUMAN, AF030065_l, HEPS RAT, PLMN PIG, P 889430, P 889935, PLMN HORSE;
PLMN BOVIN
and P 883959.
EXAMPLE i3: Isolation of cDNA Tones Encoding Human PR0545
A consensus sequence was obtained relative to a variety of EST seqltences as
described in Example I above,
wherein the consensus sequence obtained is herein designated DNA44706. An EST
proprietary to Genentech was
employed in the consensus assembly and is herein designated DNA13217 (Figure
31; SEQ ID N0:75). Based on
the DNA44706 consensus sequence, oligonucleotides were synthesized: 1) to
identify by PCR a cDNA library that
contained the sequence of interest, and 2) for use as probes to isolate a
clone of the full-length coding sequence for
PR0545.
Forward and reverse PCR primers were synthesized:
forward PCR primers 5'-GTCTCAGCACGTGTTCTGGTCTCAGGG-3' (SEQ ID N0:7fi)
f~vyrard pCR primer 2 5'-CATGAGCATGTGCACGGC-3' (SEQ ID NO:77)
forward PCR primer 3 5'-TACCTGCACGATGGGCAC-3' (SEQ ID NO:78)
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forward PCR primer 4 5'-CACTGGGCACCTCCCTTC-3' (SEQ ID N0:79)
reverse PCR~rirr~,er I 5'-CTCCAGGCTGGTCTCCAAGTCCTTCC-3' (SEQ ID N0:80)
reverse PCR primer 2 5'-TCCCTGTTGGACTCTGCAGCTTCC-3' (SEQ ID N0:8I)
reverse PCR,primer 3 5'-CTTCGCTGGGAAGAGTTTG-3' (SEQ ID N0:82)
Additionally, a synthetic oligonucleotide hybridization probe was constructed
from the consensus DNA44706
sequence which had the following nucleotide sequence
hybridization pro,~e
S'-GTGCAACCAACAGATACAAACTCTTCCCAGCGAAGAAGCTGAAAAGCGTC-3'
(SEQ ID N0:83)
In order to screen several libraries, for a source of a full-length clone, DNA
from the libraries was screened
by PCR amplification with one of the PCR primer pairs identified above. A
positive library was then used to isolate
clones encoding the PR0545 gene using the probe oligonucleotide and one of the
PCR primers. RNA for
construction of the cDNA libxaries was isolated from human placenta tissue
(L1B90).
DNA sequencing of the clones isolated as described above gave the full-length
DNA sequence for PR0545
[herein designated as UNQ346 (DNA49624-1279)] (SEQ ID NO:?3) and the derived
protein sequence for PR0545.
The entire nucleotide sequence of UNQ346 (DNA49624-1279) is shown in Figure 29
(SEQ 1D N0:73).
Clone UNQ346 (DNA49624-1279) contains a single open reading frame with an
apparent uanslational initiation site
at nucleotide positions 3I1-313 and ending at the stop codon at nucleotide
positions 2516 2518 (Figure 29). The
predicted polypeptide precursor is 735 amino acids long (Figure 30): The full-
length PR0545 protein shown in
Figure 30 has an estimated molecular weight of about 80,177 daltons and a pI
of about 7.08. Important regions of
the PR0545 amino acid sequence include the signal peptide, corresponding to
amino acids 1-28, five potential
N-glycosylation sites, from about amino acid 111-114, amino acids 146-149,
amino acids 348-351, amino acids 449-
452, and amino acids 648-651, and a neutral zinc metallopeptidase, zinc-
binding region signature sequence, from
about amino acids 344-353. Clone UNQ346 (DNA49624-1279) has been deposited
with ATCC and is assigned
ATCC deposit no. 209655.
EXAMPLE 14: Isolation o~cD~C~ne~Encoding Human PR0617
A consensus sequence was obtained relative to a variety of EST sequences as
described in Example 1 above,
wherein the consensus sequence obtained is herein designated DNA42798. Based
on the DNA42798 sequence,
oligorntcleotides were synthesized: 1) to identify by PCR a cDNA library that
contained the sequence of interest, and
2) for use as probes to isolate a clone of the full-length coding sequence for
PR0617.
A pair of PCR primers (forward and reverse) were synthesized:
forward PCR prim,.gr 5'-ACGGGCACACTGGATCCCAAATG-3' (SEQ ID N0:86)
Feverse PCR primer 5'-GGTAGAGATGTAGAAGGGCAAGCAAGACC-3' (SEQ ID NO:8?)
Additionally, a synthetic oligoztucleotide hybridization probe was constructed
from the consensus DNA42798
sequence which had the following nucleotide sequence
hybridization ro ~
5'-GCTCCCTACCCGTGCAGGTTTCTTCATTTGTTCCTTTAACCAGTATGCCG-3' (SEQ ID NO:88)
In order to screen several libraries for a source of a full-length clone, DNA
from the libraries was screened
by PCR amplification with the PCR primer pair identified above. A positive
library was then used to isolate clones
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encoding the PR0617 gene using the probe oligonucleotide and one of the PCR
primers. RNA for construction of
the cDNA libraries was isolated from human fetal kidney tissue (LIB227).
DNA sequencing of the clones isolated as described above gave the full-length
DNA sequence for PR0617
(herein designated as UNQ353 (DNA48309-1280)} (SEQ ID NO:1) and the derived
protein sequence for PR0617.
The entire nucleotide sequence of UNQ353 (DNA48309-1280) is shown in Figure 32
(SEQ ID N0:84).
Clone UNQ353 (DNA48309-1280) contains a single open reading frame wiith an
apparent uanslational initiation site
at nucleotide positions ?23-725 and ending at the stop codon at nucleotide
positions 924-926 (Figure 32). The
predicted polypeptide precursor is 67 amino acids long (Figure 33). The ful>I-
length PR0617 protein shown in Figure
33 has an estimated molecular weight of about 6,981 daltons and a pI of about
7.47. Analy~js of the PR0617 amino
acid sequence also evidences the existence of a putative signal peptide from
about amino acid 15 to about amino acid
27 and a putative protein kinase C phosphorylation site from about amino acid
41 to about amino acid 43. Clone
UNQ353 t',DNA48309-1280) has been deposited on March 5, 1998 with .ATCC and is
assigned ATCC deposit no.
209656.
Analysis of the amino acid sequence of the full-length PR0617 polypeptide
suggests that it possesses
significant homology to the CD24 protein, thereby indicating that PR0617 may
be a novel CD24 homolog. More
specifically, an analysis of the Dayhoff database (version 35.45 SwissProt 35)
evidenced significant homology
between the PR0617 amino acid sequence and the following Dayhoff sequences,
CD24 HUMAN, CD24 MOUSE,
S15785, CD24 RAT, VGE BPG4, MSES HUMAN, HSMHC3W36A 2, MLUI51$4 8, P 885075,
SEPL HUMAN
and MTCY63 13.
EXAMPLE 15: Isolation of cDNA Clones EneodingHuman PR0700
A consensus sequence was obtained relative tn a variety of EST sequences as
described in Example 1 above,
wherein the consensus sequence obtained is herein designated DNA30837. Based
on the DNA30837 consensus
sequence, oligonucleotides were synthesized: 1) to identify by PCR a cDNA
library that contained the sequence of
interest, and 2) for use as prolxs to isolate a clone of the full-length
coding sequence for PR0700.
Fotvvard and reverse PCR primers were synthesized:
forwa PCR nFimer 1 5'-ATGTTCTTCGCGCCCTGGTG-3' (SEQ ID N0:91)
forward PCR pn'me_r 2? 5'-CCAAGCCAACACACTCTACAG-3' (SEQ ID N0:92)
averse PCR"p~mer 1 5 -AAGTGGTCGCCTTGTGCAACGTGC-3
' (SEQ ID N0:93)
averse PC rimer 5'-GGTCAAAGGGGATATATCGCCAC-3' (SEQ ID NO~.94)
Additionally, a synthetic oligonucleotide hybridization probe was constructed
from the consensus DNA30837
sequence which had the following mtcleotide sequence
hybridiz tion probe
5'-GCATGGAAGATGCCAAAGTCTATGTGGCTAAAGTGGACTGCACGGCCCA-3'
(SEQ ID N0:95)
In order to screen several libraries for a source of a full-length clone, DNA
from the libraries was screened
by PCR amplification with one of the PCR primer pairs identified above. A
positive library was then used to isolate
clones encoding the PR07D0 gene using the probe oligonucleotide and one of the
PCR primers. RNA for
constrnction of the cDNA libraries was isolated from human fetal kidney tissue
(L,IB227).
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DNA sequencing of the clones isolated as described above gave the full-length
DNA sequence for PR0700
[herein designated as UNQ364 (DNA46776-1284)] (SEQ ID N0:89) and the derived
protein sequence for PR0700.
The entire nucleotide sequence of UNQ364 (DNA46776-1284) is shown in Figure 34
(SEQ ID N0:89).
Clone UNQ3b4 (DNA46776-1284) contains a single open reading frame with an
apparent translational initiation site
at nucleotide positions 33-35 and ending at the stop codon at nucleotide
positions 1329-1331 (Figure 34). The
predicted polypeptide precursor is 432 amino acids long (Figure 35). The full-
length PR0700 protein shown in
Figure 35 has an estimated molecular weight of about 47,629 daltons and a pI
of about 5.90. Important regions of
the amino acid sequence of PR0700 include the signal peptide, corresponding to
amino acids from about 1 to 33,
regions homologous to disulfide isomerase, corresponding to amino acids from
about 82-99, 210-255, and 345-360,
a tyrosine kinase phosphorylation site, corresponding to amino acids from
about 143-151, and an endoplasmic
reticulum targeting sequence, corresponding to amino acids from about 429-432.
Clone UNQ364 (DNA46776-1284)
has been deposited with ATCC and is assigned ATCC Deposit No. 209721.
EXAMP~.E 16: T~nlaric,n of cDNA Clones Encoding Human PR0702
A consensus sequence was obtained relative to a variety of EST sequences as
described in Example 1 above,
wherein the consensus sequence obtained is herein designated DNA36623. Based
on the DNA36623 consensus
sequence, oligonucleotides were synthesized: 1) to identify by PCR a cDNA
library that contained the sequence of
interest, and 2) for use as probes to isolate a clone of the full-length
coding sequence for PRO?02.
A pair of PCR primers (forward and reverse) were synthesized:
forward PCR~rimer (36623.f1) 5'-CGCTGACTATGTTGCCAAGAGTGG-3' (SEQ ID N0:98)
reverse PCR printer 136623.r1) 5'-GATGATGGAGGCTCCATACCTCAG-3' (SEQ ID N0:99)
Additionally, a synthetic oligorntcleotide hybridization probe was constructed
from the consensus DNA36623
sequence which had the following nucleotide sequence
~,tvbridization probe 136623.n1)
5'-GTGTTCATTGGCGTGAATGACCTTGAAAGGGAGGGACAGTACATGTTCAC-3' (SEQ ID N0:100)
1n order to screen several libraries for a source of a full-length clone, DNA
from the libraries was screened
by PCR amplification with the PCR primer pair identified above. A positive
library was then used to isolate clones
encoding the PR0702 gene using the probe oligonucleotide and one of the PCR
primers. RNA for construction of
the cDNA libraries was isolated from human fetal liver tissue (L.IB229).
DNA sequencing of the clones isolated as described above gave the full-length
DNA sequence for PR0702
[herein designated as UNQ366 tDNA50980-1286)] (SEQ ID N0:96) and the derived
protein sequence for PR0702.
The entire nucleotide sequence of UNQ366 (DNA50980-1286) is shown in Figure 36
(SEQ ID N0:96).
C1one UNQ366 (DNA50980-1286) contains a single open reading frame with an
apparent uanslational initiation site
at nucleotide positions 22-24 and ending at the stop codon at nucleotide
positions 853-855 (Figure 36). The predicted
polypeptide precursor is 277 amino acids long (Figure 37). The full-length
PR0702 protein shown in Figure 3? has
an estimated molecular weight of about 30,645 daltons and a pI of about 7.47.
Analysis of the full-length native
PR0702 amino acid sequence evidences the presence of a putative signal peptide
from about amino acid 1 to about
amino acid 25, potential N-glycosylation sites from about amino acid 230 to
about amino acid 233 and from about
amino acid 258 to about amino acid 261 and a C-type lectin domain signature
sequence from about amino acid 248
to about amino acid 270. Glove UNQ366 (DNA50980-1286) has been deposited with
ATCC on March 31, 1998 and
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is assigned ATCC deposit no. 209717.
Analysis of the amino acid sequence of the full-length PR0702 polypeptide
suggests that it possesses
significant sequence similarity to the conglutinin protein, thereby indicating
that PR0702 may be a novel conglutinin
homolog. More specifically, an analysis of the Dayhoff database (version 35.45
SwissProt 35) evidenced significant
homology between the PR0702 amino acid sequence and the following Dayhoff
sequences, S32436, P 875642, P
WI8780, P W18781, A53330, AC002528 1, HSPPA2IC0 1; CA21 HUMAN, CA14 HUMAN and
A61262.
EXAMPLE I7: Isolation of cDNA Clones Fncodin~l-I,t~trna~ PRQZ,O~
A consensus sequence was obtained relative to a variety of EST sequences as
described in Example 1 above,
wherein the consensus sequence obtained is herein designated DNA43047. Based
on the DNA43047 consensus
sequence, oIigonucleotides were synthesized: 1) to identify by PCR a cDNA
library that contained the sequence of
interest, and 2) far use as probes to isolate a clone of the full-length
coding sequence for PR0703.
Forward and reverse PCR primers were synthesized:
forward PCR ,primer 5'-GAGAGCCATGGGGCTCCACCTG-3' (SEQ fD N0:103)
reve se CR pritrter 1 5'-GGAGAATGTGGCCACAAC-3' (SEQ ID N0:104)
reverse PCR primer 2 5'-GCCCTGGCACAGTGACTCCATAGACG-3' ($EQ ID NO:105)
reverse PCR rimer 3 5'-ATCCACTTCAGCGGACAC-3' (SEQ ID N0:106)
Additionally, a synthetic oligonucleotide hybridization probe was constructed
from the consensus DNA40654
sequence which had the following nucleotide sequence
hybridi~,alion probe
5'-CCAGTGCCAGGATACCTCTCTTCCCCCCAGAGCATAACAGACACG-3'
(SEQ ID N0:107)
1n order to screen several libraries far a source of a full-length clone, DNA
from the libraries was screened
by PCR amplification with one of the PCR primer pairs identified above. A
positive library was then used to isolate
clones encoding the PR0703 gene using the probe oligonucieotide and one of the
PCR primers. RNA for
construction of the cDNA libraries was isolated from human fetal kidney tissue
(LIB227).
DNA sequencing of tla; clones isolated as described above gave the fitIl-
length DNA sequence for PR0703
[herein designated as UNQ367 (DNA50913-1287)] (SEQ ID NO:I01 ) and the derived
protein sequence for PR0703.
The entire nucleotide sequence of UNQ357 (DNA50913-1287) is shown in Figure 38
(SEQ ID NO:101).
Clone UNQ367 (DNA50913-1287) contains a single open reading frame with an
apparent translational initiation site
at nucleotide positions I IS-I17 and ending at the stop codon at nucleotide
positions 2305-2307 (Figure 38). The
predicted polypeptide precursor is 730 amino acids long (Figure 39). The full-
length PR0703 protein shown in
Figure 39 has an estimated molecular weight of about 78,644 daltons, and a p1
of about: 7.b5. Impor~taant regions of
the PR0703 amino acid sequence include the signal peptide, a CAMP- and cGMP-
dependent protein kinase
phosphorylation site, a CUB domain protein motif, N-glycosyIation sites and a
putative AMP-binding domain
signature. Clone UNQ367 (DNAS0913-1287) has been deposited with ATCC and is
assigned ATCC deposit no.
20971b.
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EXAMPLE I8: Isolation of cDNA Clones Encoding Human FR0745
A consensus sequence was obtained relative to a variety of EST sequences as
described in Example 1 above,
wherein the consensus sequence obtained is herein designated DNA43437. Based
on the DNA43437 consensus
sequence, oligonucleotides were synthesized: 1) to identify by PCR a cDNA
library that contained the sequence of
interest, and 2) for use as probes to isolate a clone of the full-length
coding sequence for PR0705.
A pair of PCR primers (forward and reverse) were synthesized:
forward PCR~rimer 5'-AAGCGTGACAGCGGGCACGTC-3' (SEQ ID NO:110)
reverse PCR primer 5'-TGCACAGTCTCTGCAGTGCCCAGG-3' (SEQ ID NO:111)
Additionally, a synthetic oligonucleotide hybridization probe was constructed
from the consensus DNA43437
sequence which had the following nucleotide sequence
. h~rbridization probe 143437.n11
5'-GAATGCTGGAACGGGCACAGCAAAGCCAGATACTTGCCTG-3' (SEQ ID N0:112)
In order to screen several libraries for a source of a full-length clone, DNA
from the libraries was screened
by PCR amplification with the PCR primer pair identified above. A positive
library was then used to isolate clones
encoding the PRO705 gene using the probe oligonucleotide and one of the PCR
primers. RNA for construction of
the cDNA libraries was isolated from human fetal kidney tissue (LIB22?).
DNA sequencing of the clones isolated as described above gave the full-length
DNA sequence for PRO?05
[herein designated as UNQ369 (DNA50914-1289)] (SEQ ID N0:108) and the derived
protein sequence for PR0705.
The entire nucleotide sequence of UNQ369 (DNA50914-1289) is shown in Figure 40
(SEQ ID N0:108).
Clone UNQ369 fDNA50914-1289) contains a single open reading frame with an
apparent translational initiation site
at nucleotide positions 566-568 and ending at the stop codon at nucleotide
positions 2231-2233 (Figure 40). The
predicted polypeptide precursor is 555 amino acids long (Figure 41). The full-
length PR0705 protein shown in
Figure 41 has an estimated molecular weight of about 62,736 daltons and a pI
of about 5.36. Analysis of the full-
length PR0705 sequence as shown in Figure 41 evidences the presence of the
following: a signal peptide from about
amino acid 1 to about amino acid 23, a eukaryotic DNA topoisomerase I active
site from about amino acid 418 to
about amino acid 436, and various regions that show homology to various
glypican proteins from about amino acid
237 to about amino acid 279, about amino acid 421 to about amino acid 458,
about amino acid 53 to about amino acid
74, about amino acid 466 to about amino acid 504, about amino acid 308 to
about amino acid 355, about amino acid
104 to about amino acid 156 and about amino acid 379 to about amino acid 410.
Clone UNQ369 (DNA50914-1289)
has been deposited with ATCC on March 31, 1998 and is assigned ATCC deposit
no.209722.
Analysis of the amino acid sequence of the full-length PR0705 poIypeptide
suggests that it possesses
significant sequence similarity to the K-glypican protein, thereby indicating
that PR0705 may be a novel glypican
protein family member. More specifically, an analysis of the Dayhoff database
(version 35.45 SwissProt 35)
evidenced significant homology between the PR0705 amino acid sequence and the
following Dayhoff sequences,
GFCK _ - -MOUSE, GLYP CHICK, GLYP RA?, GLYP HUMAN, GPC2 RAT, GPCS HUMAN, GPC3
HUMAN,
GPC3 RAT, P 830168 and CEC03H12 2.
E~MPLE 19: Zs_olation of cDNA Clones Encoding, Human PR0708
A consensus sequence was obtained relative to a variety of EST sequences as
described in bxample 1 above,
wherein the consensus sequence obtained is herein designated DNA34024. Based
on the DNA34024 consensus
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sequence, oligonucleotides were synthesized: 1) to identify by PCR a cDNA
library that contained the sequence of
interest, and 2) for use as probes to isolate a clone of the full-length
coding sequence for PR0708.
A'pair of PCR primers (forward and reverse) were synthesized:
forward PCR primer 5'-CCCAACCCAACTGTTTACCTCTGG-3' (SEQ ID NO:115)
reverse PAR primer S '-CTCTCTGAGTGTACATCTGTGTGG-3' (SEQ ID NO:I I6)
Additionally, a synthetic oligonucleotide hybridization probe was constructed
from the consensus DNA34024
sequence which had the following nucleotide sequence
hybridization pro>2e
5'-GCCACCCTACCTCAGAAACTGAAGGAGGTTGGNTATTCAACGCATATGGTCGG-3' (SEQ ID NO:137)
In order to screen several libraries for a source of a full-length clone, DNA
from the libraries was screened
by PCR amplification with the PCR primer pair identified above. A positive
library was then used to isolate clones
encoding the PR0708 gene using the probe oligonucleotide and one of the PCR
primers. RNA for construction of
the cDNA libraries was isolated from human bone marrow tissue (LIB255).
DNA sequencing of the clones isolated as described above gave the full-length
DNA sequence for PRO?08
[herein designated as UNQ372 (DNA48296-1292)] (SEQ ID NO: l I3) and the
derived protein sequence for PR0708.
The entire nucleotide sequence of UNQ372 (DNA48296-1292) is shown in Figures
42A-B (SEQ ID
NO:I 13). Clone UNQ372 (DNA48296-1292) contains a single open reading frame
with an apparent. translational
initiation site at nucleotide positions 891-893 and ending at the stop codon
at nucleotide positions 2436-2438 (Figures
42A-B). The predicted polypeptide precursor is 515 amino acids long (Figure
43). The full-length PR0708 protein
shown in Figure 43 has an estimated molecular weight of about 56,885 daltons
and a pI of about 6.49. Analysis of
ZO the PR0708 amino acid sequence shown in Figure 43 (SEQ ID N0:114) evidences
the existence of a putative signal
peptide at about amine acid 1 to about amino acid 37, putative sulfatase
signature sequences at about amino acid 120
to about amino acid 332 and about amino acid 168 to about amino acid I77, a
putative tyrosine ldnase phosphorylation
site from about amino acid 163 to about amino acid 169 and potential N-
glycosylation sites from about amino acid
357 to about amino acid 160, about amino acid 306 to about amino acid 309 and
about amino acid 318 to about amino
acid 321. Clone UNQ372 (DNA48296-1292) has been deposited with ATCC on March i
i , 1998 and is assigned
ATCC deposit no. 209668.
Analysis of the amino acid sequence of the full-length PR0708 polypeptide
suggests that it possesses
significant homology to the aryl sulfatase proteins, thereby indicating that
PR0708 may be a novel aryl sulfatase
homolog. More specifically, an analysis of the Dayhoff database (version 35.45
SwissProt 35) evidenced significant
homology between the PR0708 amino acid sequence and the following Dayhoff
sequences, ARSB_HUMAN,
CELC54D2 2, 602857, STS HUMAN, I37186, I37187, GEN12648, CELD1014 ?, GAGS
HUMAN and
SPHM HUMAN.
EXAMPLE 20: Isolation of cDNA Clones Enco_~Og HLma~PRCt~~
A consensus sequence was obtained relative to a variety of EST sequences as
described in Example 1 above,
wherein the consensus sequence obtained is herein designated DNA2f739. Based
on the DNA28739 consensus
sequence, oligonucleotides were synthesized: i) to identify by PCR a cDNA
library that contained the sequence of
interest, and 2) for use as probes to isolate a clone of the full-length
coding equence for PR0320.
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A pair of PCR primers (forward and reverse) were synthesized:
forward PCR~r~mer 5'-CCTCAGTGGCCACATGCTCATG-3' (SEQ ID N0:120)
:reverse PCR primer 5'-GGCTGCACGTATGGCTATCCATAG-3' (SEQ ID N0:121)
Additionally, a synthetic oligonucleotide hybridization probe was constructed
from the consensus DNA28739
sequence which had the following nucleotide sequence
S hvbridization probe
5'-GATAAACTGTCAGTACAGCTGTGAAGACACAGAAGAAGGGCCACAGTGCC-3' (SEQ ID N0:122)
In order to screen several libraries for a source of a full-length clone, DNA
from the libraries was screened
by PCR amplification with the PCR primer pair identified above. A positive
library was then used to isolate clones
encoding the PR0320 gene using the probe oligonucleotide and one of the PCR
primers. RNA for construction of
the cDNA libraries was isolated from human fetal lung tissue (LIB25).
DNA sequencing of the clones isolated as described above gave the fall-length
DNA sequence for PR0320
[herein designated as UNQ281 (DNA32284-1307)] (SEQ ID N0:118) and the derived
protein sequence for PR0320.
The entire nucleotide sequence of UNQ281 {DNA32284-1307) is shown in Figure 44
(SEQ ID N0:118).
Clone UNQ281 (DNA32284-1307) contains a single open reading frame with an
apparent translational initiation site
at nucleotide positions 135-137 and ending at the stop codon at nucleotide
positions 1149-i 151 (Figure 44). The
predicted polypeptide precursor is 338 amino acids long (Figure 45). The full-
length PR0320 protein shown in
Figure 45 has an estimated molecular weight of about 37,143 daltons and a p1
of about 8.92. Important regions of
the PR0320 amirm acid sequence include the signal peptide, corresponding to
amino acids 1-21, an EGF-like domain
cysteine pattern signature, corresponding to amino acids 80-91, and three
calcium-binding EGF-like domains,
corresponding to amino acids 103-124. 230-151 and 185-206, respectively. Clone
UNQ281 (DNA32284-I307) has
been deposited with ATCC and is assigned ATCC deposit no. 209670.
EXAMPLE 21: Isolation of cDN ~ Clones Encoding Human PR0324
A consensus sequence was obtained relative to a variety of EST sequences as
described in Example 1 above,
wherein the consensus sequence obtained is herein designated DNA34347. Based
on the DNA34347 consensus
sequence, oligonucleotides were synthesized: 1) to identify by PCR a cDNA
library that contained the sequence of
interest, and 2) for use as probes to isolate a clone of the full-length
coding sequence for PR0324.
PCR primers (forward and reverse) were synthesized:
forward~CR Qrimer 1 5'-GCAATGAACTGGGAGCTGC-3' (SEQ ID N0:125)
fear PAR primer 2 5'-CTGTGAATAGCATCCTGGG-3' (SEQ ID N0:126)
fo arv~R primer 3 5'-C1T1~TCAAGCCACTGGAGGG-3' . (SEQ ID N0:127)
rgyerse PCR primer 1 5'-CTGTAGACATCCAAGCTGGTATCC-3' (SEQ ID N0:128)
reverse PCR primer 2 S'-AAGAGTCTGCATCCACACCACTC-3' (SEQ ID N0:129)
Additionally, a synthetic oligonucleotide hybridization probe was constructed
from the consensus DNA34347
sequence which had the following nucleotide sequence
I?yrridizati~n ,probe
5'-ACCTGACGCTACTATGGGCCGAGTGGCAGGGACGACGCCCAGAATG-3' (SEQ ID N0:130)
In order to screen several libraries for a source of a full-length clone, DNA
from the libraries was screened
by PCR amplification with one of the PCR primer pairs identified above. A
positive library was then used to isolate
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clones encoding the PR0324 gene using the probe oligonucleotide and one of the
PCR primers. RNA for
construction of the cDNA libraries was isolated from human fetal liver tissue
(LIB6).
DNA sequencing of the clones isolated as described above gave the full-length
DNA sequence for PRO324
(herein designated as UNQ285 (DNA36343-1310)] (SI:Q ID N0:123) and the derived
protein sequence for PR0324.
The entire nucleotide sequence of UNQ285 (DNA36343-1310) is shown in Figure 46
(SEQ ID NO:123).
Clone UNQ285 (DNA36343-I3I0) contains a single open reading frame with an
apparent translational initiation site
at nucleotide positions 144-14b and ending at the stop colon at nucleotide
positions 1011-1013 (Figure 46). The
predicted poIypeptide precursor is 289 amino acids long (Figure 47). The full-
length PR0324 protein shown in
Figure 47 has an estimated molecular weight of about 32,268 dahons and a pI of
about 9.21. Analysis of the PR0324
polypeptide sequence shown in Figure 47 (SEQ ID N0:124) evidence the presence
of the following: a signal peptide
IO from about amino acid I to about amino acid 31, a transmembrane domain from
about amino acid 136 to about amino
acid 157, tyrosine Iattase phosphorylation sites from about amino acid 106 or
about amino acid 107 to about amino
acid 113 and regions that are homologous to short-chain alcohol dehydrogenase
regions from about amino acid 80
to about amino acid 90, from about amino acid I3l to about amino acid 168,
from about amino acid 1 to about amino
acid 13 and from about amino acid 176 to about amino acid I85. Clone UNQ285
(DNA36343-1310) has been
deposited with ATCC on March 30, 1998 and is assigned ATCC deposit no. 209718.
Analysis of the amino acid sequence of the full-length PR0324 polypeptide
suggests that it possesses
significant sequence similarity to oxidoreductases; thereby indicating that
PR0324 may be a novel oxidoreduetase
homolog. More specifically, an analysis of the Dayhoff database (version 35.45
SwissProt 35) evidenced significant
homology between the PR0324 amino acid sequence and the following Dayhoff
sequences, 861209, A69965,
YQJQ BACSU, D69930, 576124, FABG ECOLI, C70023, 577280, FABG VIBHA and MTV013
6.
EXAMPLE 22: Isolation of ~DN~Clortes Er~coding",~Iuman PRO351
A consensus sequence was obtained relative to a variety of EST sequences as
described in Example I above,
wherein the consensus sequence obtained is herein designated DNA35950. Based
on the DNA3S950 consensus
sequence, oligonucleotides were synthesized: 1) to identify by PCR a cDNA
library that contained the sequence of
interest, and 2) for use as probes to isolate a clone of the full-length
coding sequence for PR0351.
Forward and reverse PCR primers were synthesized:
forward PCR primer 5'-CCTGTGCTGTGCCTCGAGCCTGAC-3' (SEI~ ID N0:133)
reverse PCR primer 5'-GTGGGCAGCAGTTAGCACCGCCTC-3' (SEQ ID N0:134)
Additionally, a synthetic oligonucleotide hybridization probe was constructed
from the consensus DNA35950
sequence which had the following nucleotide sequence
~,tybridi2ation
5'-GGCTGGCATCATCAGCTTTGCATCAAGCTGTGCCCAGGAGGACGC-3'
(SEQ ID N0:135)
In order to screen several libraries for a source of a full-length clone, DNA
from the libraries was screened
by PCR amplification with one of the PCR primer pairs identified above. A
positive library was then used to isolate
clones encoding the PR0351 gene using the probe oligonucleotide and one of the
PCR primers. RNA for
construction of the cDNA libraries was isolated from human fetal liver tissue
(Li8230).
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DNA sequencing of the clones isolated as described above gave the full-length
DNA sequence for PR0351
[herein designated as UNQ308 (DNA40571-1315)) (SEQ ID NO: I31 ) and the
derived protein sequence for PR0351.
The entire nucleotide sequence of UNQ308 (DNA40571-1315) is shown in Figure 48
(SEQ ID N0:131}.
Clone UNQ308 tDNA4057I-1315) contains two open reading frames with an apparent
translational initiation site at
nucleotide positions 189-191 and a second open reading frame beginning at
nucleotide 4?0, with the two open reading
frames ending at the stop codons at nucleotide positions 363-365 and 2009-
2011, respectively (Figure 48). The
predicted polypeptide precursor is 571 amino acids long (Figure 49). Important
regions of the amino acid sequence
of PR0351 include the signal peptide, regions having sequence similarity to
serine proteases of the trypsin family,
two N-glycosylation sites, and three Kringle domains. Clone UNQ308 (DNA40571-
1315) has been deposited with
ATCC and is assigned ATCC deposit no. 209784.
EX,QMPLE 23: Isolation of cDNA Clones Encoding~Human PR0352
A consensus sequence was obtained relative to a variety of EST sequences as
described in Example 1 above,
wherein the consensus sequence obtained is herein designated DNA36950. Based
on the DNA36950 consensus
sequence, oligonucleotides were synthesized: 1) to identify by PCR a cDNA
library that contained the sequence of
interest, and 2) for use as probes to isolate a clone of the full-length
coding sequence for PR0352.
PCR primers (forward and reverse) were synthesized:
forward primer I 5'-CTGGCACAGCTCAACCTCATCTGG-3'
PCR (SEQ ID . :I38)
forward primer 2 5'-GCTGTCTGTCTGTCTCATTG-3' (SEQ
PCR ID N0:139)
forward rimer 3 5'-GGACACAGTATACTGACCAC-3' (SEQ
PCR~ ID N0:140)
reverse rimer 1 5'-TGCGAACCAGGCAGCTGTAAGTGC-3' (SEQ
PCRp ID N0:141)
reverse rimer 2 5'-TGGAAGAAGAGGGTGGTGATGTGG-3' (SEQ
PQR~ ID N0:142)
Additionally, a synthetic oligonucleotide hybridization probe was constructed
from the consensus DNA36950
sequence which had the following nucleotide sequence
~,rbridization probe
5'-CAGCTGACAGACACCAAACAGCTGGTGCACAGTTTCACCGAAGGC-3' (SEQ ID NO:143)
In order to screen several libraries for a source of a full-length clone, DNA
from the libraries was screened
by PCR amplification with the PCR primer pair identified above. A positive
library was then used to isolate clones
erioodi<tg the PR0352 gene using the probe oligonucleotide and one of the PCR
primers. RNA for construction of
the cDNA libraries was isolated from human fetal kidneytissue (LIB227).
DNA sequencing of the clones isolated as described above gave the full-length
DNA sequence for PR0352
(herein designated as UNQ309 (DNA41386-1316)] (SEQ ID N0:136) and the derived
protein sequence for PR0352.
The entire nucleotide sequence of UNQ309 (DNA41386-1316) is shown in Figure 50
(SEQ ID N0:136).
Clone UNQ309 (DNA41386-1316) contains a single open reading frame with an
apparern translational initiation site
at nucleotide positions 152-154 and ending at the stop codon at nucleotide
positions 1100-1102 (Figure 50). The
predicted polypeptide precursor is 316 amino acids long (Figure 51). The full-
length PR0352 protein shown in
Figure 2 has an estimated pI of about 4.62. Analysis of the full-length PR0352
sequence evidences the presence of
a signal peptide from about amino acid 1 to about amino acid 28, a
transmembrane domain from about amino acid
251 to about amino acid 270, potential N-glycosylation sites from about amino
acid 91 to about amino acid 94, about
amino acid 104 to about amino acid 107, about amino acid 189 to about amino
acid 192 and about amino acid 215
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to about amino acid 218 and a region having homology to immunoglobulins and
MHC from about amine acid 217
to about amino acid 234. Clone UNQ309 (DNA41386-1316) has been deposited with
ATCC on March 26, 1998 and
is assigned ATCC deposit tio. 209703.
Analysis of the amino acid sequence of the full-length PR0352 polypeptide
suggests that it possesses
significant sequence similarity to the buryrophilin protein, thereby
indicating that PR0352 is a novel buryrophilin
homolog. More specifically, an analysis of the Dayhoff database (version 35.45
SwissProt 35) evidenced significant
homology between the PR0352 amino acid sequence and the following Dayhoff
sequences, BUTY HUMAN,
HSB73~1, GGCD80_l, 146690, A33 HUMAN, P 867988, CD86 MOUSE, P R713b0, B39371
and D50558~1.
EXAMPLE 24: Isolation of cDNA CZ~s Encoding Humarl ~R0381
A consensus sequence was obtained relative'to a variety of EST sequences as
described in Example I above,
wherein the consensus sequence obtained is herein designated DNA396S1. Based
on the DNA39651 consensus
sequence, oligonucleotides were synthesized: 1) to identify by PCR a cDNA
library that contained the sequence of
interest, and 2) for use as probes to isolate a clone of the full-length
coding sequence for PR0381.
A pair of PCR primers (forward and reverse) were synthesized:
IS forward PC~,~rimer 5'-CTTTCCTTGCTTCAGCAACATGAGGC-3' (SEQ ID N0:146)
reverse SCR primer 5'-GCCCAGAGCAGGAGGAATGATGAGC-3' (SEQ ID NO:I47)
Additionally, a synthetic oligonucleotide hybridization probe was constructed
from the consensus DNA39651
sequence which had the following nucleotide sequence
hybridization probe
5'-GTGGAACGCGGTCTTGACTCTGTTCGTCACTTCTTTGATTGGGGCTTTG-3' (SEQ ID N0:148)
1n order to screen several libraries for a source of a full-length clone, DNA
from the libraries was screened
by PCR amplification with the PCR primer pair identified above. A positive
library was then used to isolate clones
encoding the PR038I gene using the probe oligonucleatide and one of flue PCR
primers. RNA for construction of
the cDNA libraries was isolated from human fetal kidney tissue (L1B22'7).
DNA sequencing of the clones isolated as described above gave the full-length
DNA sequence for PR0381
ll~rein designated as UNQ322 (DNA44194-1317) (SEQ ID NO: I44) and the derived
protein sequence for PR0381.
The entire nucleotide sequence of UNQ3z2 (DNA44194-1317) is shown in Figure 52
(SEQ ID N0:144).
Clone UNQ322 (DNA44194-I3I7) contains a single open reading frame with an
apparent translational initiation site
at nucleotide positions 174-176 and ending at the stop codon at nucleotide
positions 807-809 (Figure 52). The
predicted polypeptide precursor is ZI1 amino acids long (Figure 53). The foil-
length PR0381 protein shown in
Figure 53 has an estimated molecular weight of about 24,I72 daltons arid a p1
of about 5.99. Analysis of the full-
length PR0381 polypeptide shown in Figure 53 (SEQ ID N0:145) evidences the
presence of the following; a signal
peptide from about amino acid 1 to about amino acid 20, a potential N-
glycosylation site from about amino acid 1?6
to about amino acid 179, potential casein kinase II phosphoryladon sites from
about amino acid 143 to about amino
acid 146, from about amino acid 156 to about amino acid I59, from about amino
acid I78 to about amino acid 181,
and from about amino acid 200 to about amino acid 203, an endoplasmic
reticulum targeting sequence from about
amino aad 208 to about amino acid 211, FKBP-type pegtidyl-prolyl cis~trans
isomerase sites from about amino acid
~8 to about amino acid 114 and from about amino acid 118 to about amino acid
131, EF-hand calcium binding
'vies from about amino acid 191 to about amino acid 203, from about amino acid
184 to about amino acid 203
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and from about amino acid 140 to about anvno acid 159> and an S-100/ICaBP type
calcium binding domain from
about amino acid 183 to about amino acid 203. Clone UNQ322 (DNA44194-1317) has
been deposited with ATCC
on April 28, 1998 and is assigned ATCC deposit no. 209808.
Analysis of the amino acid sequence of the full-length PR0381 polypeptide
suggests that it possesses
significant sequence similarity to FKBP immunophilin proteins, thereby
indicating that PR0381 may be a novel FKBP
itnnaunophilin homolog. More specifically, an analysis of the Dayhoff database
(version 35.45 SwissProt 35)
evidenced significant homology between the PR0381 amino acid sequence and the
following Dayhoff sequences,
AF040252_l, 149669, P 893551, S71238, CELCOSC8_l, CEU27353_l, MIP TRYCR,
CEZC455 3,
FKB4 HUMAN and 140718.
EXAMPLE 25: Isolation of cDNA Clones Encoding Human PR0386
A consensus sequence was obtained relative to a variety of EST sequences as
described in Example 1 above,
wherein the consensus sequence obtained is herein designated DNA406?4. Two
proprietary Genentech EST
sequences were employed in the consensus sequence assembly, wherein those EST
sequences are herein designated
DNA23350 (Figure 56; SEQ ID NO:151) and DNA23536 (Figure 5?; SEQ ID N0:152).
Based on the DNA40674
consensus sequence, oligonucleotides were synthesized: 1) to identify by PCR a
cDNA library that contained the
sequence of interest, and 2) for use as probes to isolate a clone of the full-
length coding sequence for PR0386.
A pair of PCR primers (forward and reverse) were synthesized:
forward PCR.primer 5'-ACGGAGCATGGAGGTCCACAGTAC-3' (SEQ ID N0:153)
revere PC n~mer 5'-GCACGTTTCTCAGCATCACCGAC-3' (SEQ ID N0:154)
Additionally, a synthetic oligonucleotide hybridization probe was constructed
from the consensus DNA40674
sequence which had the following nucleotide sequence
h~r~ridization probe
5'-CGCCTGCCCTGCACCTTCAACTCCTGCTACACAGTGAACCACAAACAGTT-3' (SEQ ID N0:155)
In order to screen several libraries for a source of a full-length clone, DNA
from the libraries was screened
by PCR amplification with the PCR primer pair identified above. A positive
library was then used to isolate clones
encoding the PR0386 gene using the probe oligonucleotide and one of the PCR
primers. RNA for construction of
the cDNA libraries was isolated from human fetal brain tissue (LIBI53).
DNA sequencing of the.clones isolated as described above gave the full-length
DNA sequence for PR0386
[herein designated as UNQ326 (DNA45415-1318)] (SEQ ID N0:149) and the derived
protein sequence for PR0386.
The entire nucleotide sequence of UNQ326 (DNA45415-1318) is shown in Figure 54
(SEQ ID N0:149).
Clone UNQ326 (DNA45415-1318) contains a single open reading frame with an
apparent translational initiation site
at nucleotide positions 146-148 and ending at the stop codon at nucleotide
positions 791-793 (Figure 54). The
predicted polypeptide precursor is 215 amino acids long (Figure 55). The full-
length PR038ti protein shown in
Figure 55 has an estimated molecular weight of about 24,326 daltons and a p1
of about 6.32. Analysis of the full-
length PR0386 sequence shown in Figure 55 (SEQ ID NO:150) evidences the
presence of the following: a signal
peptide from about amino acid 1 to about amino acid 20, a transmembrane domain
from about amino acid 161 to
about amino acid 179, an itnmunoglobulin-like fold from about amino acid 83 to
about amino acid 127 and potential
N-glycosylation sites from about amino acid 42 to about amino acid 45, from
about amino acid 66 to about amino
acid 69 and from about amino acid 74 to about amino acid 77. Clone UNQ326
(DNA45415-1318) bas been deposited
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with ATCC on April 28, 1998 and is assigned ATCC deposit no. 209810.
Analysis of the amino acid sequence of the full-length PR0386 polypeptide
suggests that it possesses
significant sequence similarity to the sodium channel beta-2 subunit, thereiby
indicating that PR0386 is a novel
homolog thereof. More specifically, an analysis of the Dayhoff database
(version 35.45 Swissfrot 35) evidenced
significant homology between the PR0386 amitw acid sequence and the following
Dayhoff sequences; A57843,
MYPO HUMAN, GEN14531, X4024, I-iS46ICDA 1, HSU90716 t, D86996 2, MUSIGLVD I,
DMU42768 1
and S19247.
EXAMPLE ,~6_: Isolation of cDNA Clones Encoding Human PR0540
A consensus sequence was obtained relative to a variety of EST sequences as
described in Example 1 above,
wherein the consensus sequence obtained is herein designated DNA39631. Based
on the DNA39631 consensus
sequence, oligonucleotides were synthesized: 1) to identify by PCR a cDNA;
library that contained the sequence of
interest, and 2) for use as probes to isolate a clone of the full-length
coding sequence for PR0540.
Forward and reverse PCR primers were synthesized:
forward PCR primer 5'-CTGGGGCTACACACGGGGTGAGG-3' (SEQ ID NO:158)
IS reverse PCR Drimer 5'-GGTGCCGCTGCAGAAAGTAGAGCG-3' (SEQ ID N0:159)
Additionally, a synthetic oligonucleoiide hybridization probe was constructed
from the consensus DNA40b54
sequence which had the following nucleotide sequence
l~,vbridization probe
5'-GCCCCAAATGAAAACGGGCCCTACTTCCTGGCCCTCCGCGAGATG-3'
2l? (SEQ ID N0:160)
In order to screen several libraries for a source of a fesll-length clone, DNA
from the libraries was screened
by PCR amplification with one of the PCR primer pairs identified above. A
positive library was then used to isolate
clones encoding the PR0540 gene using the probe oligonucleotide and one of the
PCR primers. RNA for
construction of the cDNA libraries was isolated from human fetal kidney tissue
(LIB227).
25 DNA sequencing of the clotres isolated as described above gave the full-
length DNA sequence for PR0540
[herein designated as UNQ341 (DNA44189-1322)] {SEQ ID N0:156) and the derived
protein sequence for PR0540.
The entire nucleotide sequence of UNQ341 (DNA44189-1322) is shown in Figure 58
(SEQ ID N0:156).
Clone UNQ341 (DNA44189-1322) contains a single open reading frame with an
apparent uanslational initiation site
at nucleotide positions 21-23 and ending at the stop codon at nucleotide
positions 1257-1259 (Figure 58). The
30 predicted polypeptide precursor is 412 amino acids long {Figure 59). Tike
full-length PR0540 protein shown in
Figure 59 has an estimated molecular weight of about 46,658 daltons and a pi
of about 6.65. Important regions of
the amino acid sequence of PR0540 include the signal peptide, potential N-
glycosylation sites, a potential lipid
substrate binding site, a sequence typical of lipases and serine proteins, and
a beta-uansducin family Trp-Asp repeat.
Clone UNQ341 (DNA44189-1322) has been deposited with ATCC and is assigned ATCC
deposit no. 209699.
FXAMPhE 27: Isolation of cDN~,Clonr"s En_coy~g~uma_n PRCW15
A consensus sequence was obtained relative to a variety of ES1' sequences as
described in Example 1 above,
wherein the consensus sequence obtained is herein designated DNA42240. Based
on the DNA42240 consensus
sequence, oligonucleotides were synthesized: 1) to identify by PCR a cDNA
library that contained the sequence of
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interest, and 2} for use as probes to isolate a clone of the full-length
coding sequence for PR0615.
Forward and reverse PCR primers were synthesized:
forward PCR primer 5'-TGGTCTTCGCCTTGATCGTGTTCT-3' (SEQ ID N0;163)
forward PCR primer 5'-GTGTACTGAGCGGCGGTTAG-3' (SEQ ID N0:154)
reverse PCR primer 5'-CTGAAGGTGATGGCTGCCCTCAC-3' (SEQ ID N0:165)
reverse PCR primer 5'-CCAGGAGGCTCATGGGAAAGTCC-3' (SEQ ID NO:I66)
Additionally, a synthetic oligonucleotide hybridization probe was constructed
from the consensus DNA42240
sequence which had the following nucleotide sequence:
hybridization vrobe
5'-CCACGAGTCTAAGCAGATGTACTGCGTGTTCAACCGCAACGAGGATGCCT-3'
(SEQ ID NO:lb7)
In order to screen several Fibraries for a source of a full-length clone, DNA
from the libraries was screened
by PCR amplification with one of the PCR primer pairs identified above. A
positive library was then used to isolate
clones encoding the PR0615 gene using the probe oligonucleotide~ and one of
the PCR primers. RNA for
construction of the cDNA libraries was isolated from human bone marrow tissue
(LIB255).
DNA sequencing of the clones isolated as described above gave the full-length
DNA sequence for PR0615
[herein designated as UNQ352 (DNA48304-1323)) (SEQ ID N0:161) and the derived
protein sequence for PR0615.
The entire nucleotide sequence of UNQ352 (DNA48304-1323) is shown in Figure 60
(SEQ ID NO:161).
Clone UNQ352 (DNA48304-1323) contains a single open reading frame with an
apparent translational initiation site
at nucleotide positions 51-53 and ending at the stop codon at nucleotide
positions ?23-725 (Figure 60). The predicted
polypeptide precursor 'ss 224 amino acids long (Figure 61). The full-length
PR0615 protein shown in Figure 6I has
an estimated molecular weight of about 24,810 daltons and a p1 of about 4.75.
Important regions of the amino acid
sequence of PR0615 include a type II transmembrane domain, corresponding to
about amino acids 24-43, other
transmembrane domains, corresponding to about amino acids 74-90, 108-126, and
145-161, respectively, and a
potential N-glycosylation site, corresponding to about amino acids 97-100.
Clone UNQ352 (DNA48304-1323) has
been deposited with ATCC and is assigned ATCC deposit no. 209811.
EXAMPLE 28: Isolation of cDNA Clones Encoding Human PR0618
A consensus sequence was obtained relative to a variety of EST sequences as
described in Example 1 above,
wherein the consensus sequence obtained is herein designated DNA30900. Based
on the DNA30900 consensus
sequence, oligonucleotides were synthesized: 1) to identify by PCR a cDNA
library that contained the sequence of
interest, and 2) for use as probes to isolate a clone of the full-length
coding sequence for PR0618.
Forward and reverse PCR primers were synthesized:
~rward PCR yritner 5'-TAACAGCTGCCCACTGCTTCCAGG-3' (SEQ ID N0:171)
reverse PCR primer 5'-TAATCCAGCAGTGCAGGCCGGG-3' (SEQ ID N0:172)
Additionally, a synthetic oligonucleotide hybridization probe was constructed
from the consensus DNA30900
sequence which had the following nucleotide sequence
hybridization probe
5'-ATGGCCTCCACGGTGCTGTGGACCGTGTTCCTGGGCAAGGTGTGGCAGAA-3'
(SEQ ID N0:173)
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Screening of the above described library gave rise to the partial cDNA clone
designated herein DNA35597
(SEQ )D N0:170). Extension of this sequence using repeated cycles of BLAST and
phrap gave rise to a nucleotide
sequence designated herein as DNA43335. Primers based upon the DNA43335
consensus sequence were then
prepared as follows.
forward PQ,R ,primer 5'-TGCCTATGCACTGAGGAGGCAGAAG-3' (SEQ ID N0:174)
reverge PCR primer 5'-AGGCAGGGACACAGAGTCCATTCAC-3' (SEQ ID N0:175)
Additionally; a synthetic oligorntcleotide hybridization probe was constructed
from the consensus DNA43335
sequence which had the following nucleotide sequence
~bridization ,probe
5'-AGTATGATTTGCCGTGCACCCAGGGCCAGTGGACGATCCAGAACAGGAGG-3'
(SEQ ID N0:176)
In order to screen several libraries for a source of a full-length clone, DNA
from the libraries was screened
by PCR amplification with one of the PCR primer pairs identified above. A
positive Library was then used to isolate
foil length clones encoding the PR0618 gene using the second probe
oligonucleotide and one of the second set of PCR
primers. RNA for construction of the cDNA libraries was isolated from human
fetal liver tissue (LIB229).
DNA sequetxittg of the clones isolated as described above gave the tall-length
DNA sequence for PR0618
[herein designated as UNQ354 (DNA49152-1324)] (SEQ ID NO:I68) and the derived
protein sequence for PR0618.
The entire nucleotide sequence of UNQ354 (DNA49152-1324) is shown in Figure 62
(SEQ ID N0:168).
Clone UNQ354 (DNA49152-I324) contains a single open reading frame with an
apparent transIational initiation site
at nucleotide positions 73-75 and ending at the stop codon at nucleotide
positions 2479-2481 (Figure fi2). The
predicted polypeptide precursor is 802 amino acids long (Figure 63). The full-
length PR0618 protein shown in
Figure 63 has an estimated molecular weight of about 88,846 daltons and a pI
of about 6.41. Important regions of
the amino acid sequence of PR0618 include type II uansmembrane domain, a
sequence typical of a protease, trypsin
family, histidine active site, multiple N-glycosylation sites, two sequences
typical of a Kringle domain, two regions
having sequence similarity to Kallilaein light chain, and a region having
sequence similarity to low-density lipoprotein
receptor. Clone UNQ354 (DNA49152-1324) has been deposited with ATCC and is
assigned ATCC deposit no.
209813.
F~MPLE 29: Is9lation of cDNA Cloneq Encodinsr Huna~R0719
A consensus sequence was obtained relative to a variety of EST sequences as
described in Example 1 above,
wherein the consensus sequence obtained is herein designated DNA44851. Based
on the DNA448S1 consensus
sequence, oligonucleotides were synthesized: 1) to identify by PCR a cDNA
library that captained the sequence of
interest, and 2) for use as probes to isolate a clone of the full-length
coding sequence for PR0719.
A pair of PCR primers (forward and reverse) vrere synthesized:
5'-GTGAGCATGAGCGAGCCGTCCAC-3' (SEQ ID N0:179)
reverse P,~~,pri_~rter 5'-GCTATTACAACGGTTC'iTGCGGCAGC-3' (SEQ ID NO:I80)
Additionally, a synthetic oligonucleotide hybridization probe was constructed
from the cotuensus DNA448S 1
sequence which had the following nucleotide sequence
h~:>~~~~
5 =TTGACTCTCTGGTGAATCAGGACAAGCCGAGTTTTGCCTTCCAG-3' (SEQ ID N0:181)
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In order to screen several libraries for a source of a full-length clone, DNA
from the libraries was screened
by PCR amplification with the PCR primer pair identified above. A positive
library was then used to isolate clones
encoding the PR0719 gene using the probe oligonucleotide and one of the PCR
primers. RNA for construction of
the cDNA libraries was isolated from human placenta tissue (LIB90).
DNA sequencing of the clones isolated as described above gave the full-length
DNA sequence for PR0719
therein designated as UNQ387 (DNA49646-1327)] (SEQ 1D N0:177) and the derived
protein sequence for PR0719.
The entire nucleotide sequence of UNQ387 (DNA49646-1327) is shown in Figure 65
(SEQ ID N0:177).
Clone UNQ387 (DNA49646-1327) contains a single open reading frame with an
apparent translational initiation site
at nucleotide positions 223-225 and ending at the stop codon at nucleotide
positions 1285-1287 (Figure 65). The
predicted polypeptide precursor is 354 amino acids long (Figure 66). The full-
length PR0719 protein shown in
Figure 66 has an estimated molecular weight of about 39,362 daltons and a pI
of about 8.35. Analysis of the full
length PR0719 sequence evidences the presence of a signal peptide from about
amino acid 1 to about amino acid 16
as shown in Figure 66 (SEQ ID N0:178), a lipase-associated serine-containing
active site at about amino acid 163
to about amino acid 172, and two potential N-glycosylation sites from about
amino acid 80 to about amino acid 83
and about amino acid 136 to about amino acid 139. Clone UNQ387 (DNA49646-1327)
has been deposited with
ATCC on March 26, 1998 and is assigned ATCC deposit no. 209705.
Analysis of the amino acid sequence of the full-length PR0719 polypeptide
suggests that it possesses
significant sequence similarity to the lipoprotein lipase H protein, thereby
indicating that PR0719 may be a navel
lipoprotein lipase homolog. More specifically, an analysis of the Dayhoff
database (version 35.45 SwissProt 35)
evidenced significant homology between the PR0719 amino acid sequence and the
following Dayhoff sequences,
LIPL HUMAN, LIPH HUMAN, D83548_l, A24059l1, P 830740, D88666_l, A43357,
A46696, B43357 and
A49488.
EXAMPLE 30: Violation of cDNA Clones Encoding Human PR0724
A consensus sequence was obtained relative to a variety of EST sequences as
described in Example 1 above,
wherein the consensus sequence obtained is herein designated DNA35603. Based
on the DNA35603 consensus
sequence, oligonueleotides were synthesized: 1) to identify by PCR a cDNA
library that contained the sequence of
interest, and 2) for use as probes to isolate a clone of the full-length
coding sequence for PR0724.
Pairs of PCR primers (forward and reverse) were synthesized:
forward PCR~rimer 1 5'-GGCTGTGACTGTGGAGACAC-3' (SEQ ID N0:184)
forward PCRp itir mer 2 5'-GCAAGGTCATTACAGCTG-3' (SEQ ID N0:185)
reverse PCR orimer1 5'-AGAACATAGGAGCAGTCCCACTC-3' (SEQ ID NO:186)
reverse PCR Timer 2 5'-TGCCTGCTGCTGCACAATCTCAG-3' (SEQ ID NO:I87)
Additionally, a synthetic oligonucleotide hybridization probe was constructed
from the consensus DNA35603
sequence which had the following nucleotide sequence
hybridization r8 obe
5'-GGCTATTGCTTGCCTTGGGACAGACCCTGTGGCTTAGGCTCTGGC-3' (SEQ ID NO:I88)
In order to screen several libraries for a source of a full-length clone, DNA
from the libraries was screened
by PCR amplification with the PCR primer pairs identified above. A positive
library was then used to isolate clones
erxoding the PR0724 gene using the probe oligonucleotide and one of the PCR
primers. RNA for construction of
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the eDNA libraries was isolated from human fetal lung tissue (LIB26).
DNA sequencing of the clones isolated as described above gave the full-length
DNA sequence for PRO?24
[herein designated as UNQ389 (DNA49631-1328)] (SEQ ID N0:182) and the derived
protein sequence for PRO724.
The entire nucleotide sequence of UNQ389 (DNA49631-1328) is shown in Figure 67
(SEQ iD N0:182).
Clone UNQ389 (DNA49631-1328) contains a single open reading frame with an
apparent translational initiation site
at nucleotide positions 546-S48 and ending at the stop codon at nucleotide
positions 2685-2687 (Figure 67). The
predicted polypeptide precursor is 713 amino acids long (Figure 68): The full-
length PR0724 protein shown in
Figure 68 has an estimated molecular weight of about 76;193 daltons and a pI
of about 5.42. Analysis of the full-
length PR0724 amino acid sequence shown in Figure b8 (SEQ ID N0:183) evidences
the presence of the following:
a signal peptide from about amino acid 1 to about amino acid 16, a
uansme:mbrane domain from about amino acid
IO 442 to about amino acid 462 and LDL receptor class A domain regions from
about amino acid 1S2 to about amino
acid 171, about amino acid 331 to about amino acid 350, about amino acid 3?4
to about amino acid 393 and about
amino acid 411 to about amino acid 430: Clone UNQ389 (DNA49631-1328) has been
deposited with ATCC on April
28, 1998 and is assigned ATCC deposit no. 209806
Analysis of the amino acid sequence of the fall-length PR0724 polypeptide
suggests that it possesses
1S significant sequence similarity to the human LDL receptor protein, thereby
indicating that PR0724 may be a novel
LDL receptor homolog. More specifically, an analysis of the Dayhoff database
(version 35.45 SwissProt 35)
evidenced significant homology between the PR0724 amino acid sequence and the
following Dayhoff sequences,
P R48S47, MMAM2R 1, LRP2 RAT, P RbOSl7, P 847861, P R05S33, A44513~1, A30363,
P 874692 and
LMLIPOPHO l.
E~,AMPLE 3I: Isolation of cDNA Clones Encoding Human PR0772
One cDNA sequence was isolated in the amylase screen described in Example 2,
wherein that cDNA
sequence is herein designated DNA43S09 (see Figure 71). Based on the DNA43S09
sequence, oligonucleotide probes
were generated and used to screen a human fetal lung library (LI825) prepared
as described in paragraph 1 of
Example 2 above. The cloning vector was pRKSB (pRKSB is a precursor of pRKSD
that does not contain the Sfil
site; see, Hohnes et al., 'e ce, 25:1278-1280 (1991)), and the cDNA size cut
was less than 2800 bp.
A pair of PCR primers (forward and reverse) were synthesized based on the
DNA43S09 sequence:
forward PCR arimer S'-CGTTTTGCAGAACCTACTCAGGCAG-3' (SEQ ID N0:192)
reverse PCR~rimer S'-CCTCCACCAACTGTCAATGTTGTGG-3' (S1EQ ID N0:193)
Additionally, a synthetic oligotutcleotide hybridization probe was constructed
from the consensus DNA43S09
sequence which had the following nucleotide sequence
hybridization rp obe
S'-AAAGTGCTGCTGCTGGGTCTGCAGACGCGATGGATAACGT-3' (SEQ ID N0:194)
Using the above described primers and library, a full length clone was
identified that contained a single open
3S reading frame with an apparent translational initiation site at nucleotide
positions 131-133 and ending at the stop
colon found at nucleotide positions 587-S89 (Figure 69; SEQ ID N0:189). The
predicted polypeptide precursor is
152 amino acids long, has a calculated molecular weight of approximately
17,170 daltons and an estimated pI of
approximately 9.62. Analysis of the full-length PR0772 sequence shown in
Figure 70 (SEQ iD N0:19U) evidences
the presence of the following: a potential type II aansmembrane domain from
about amino acid 26 to about amino
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acid 42, other potential transmembrane domains from about amino acid 44 to
about amino acid 65, from about amino
acid 81 to about amino acid 101 and from about amino acid 109 to about amino
acid 129, leucine zipper pattern
sequences from about amino. acid 78 to about amino acid 99 and from about
amino acid 85 to about amino acid 106.
Clone UNQ410 (DNA49645-1347) has been deposited with ATCC on April 28, 1998
and is assigned ATCC deposit
no. 209809.
Analysis of the amino acid sequence of the full-length PR0772 polypeptide
suggests that it possesses
significant sequence similarity to the human A4 protein, thereby indicating
that PR0772 tray be a novel A4 protein
homolog. More specifically, an analysis of the Dayhoff database (version 35.45
SwissProt 35) evidenced significant
homology between the PR0772 amino acid sequence and the following Dayhoff
sequences, HSU9330S_l,
A4P HUMAN, CELB0454 2, VPU 1SRV, CELC12D12 2, OCCM AGRTl, LBPHIG1E 50, YIGK
ECOLI,
S76245 and P 850807.
EXAMPLE 32: Isolation of cDNA Clones Encoding Human PR0852
A consensus sequence was obtained relative to a variety of EST sequences as
described in Example 1 above,
wherein the consensus sequence obtained is herein designated DNA34364. Based
on the DNA34364 consensus
sequence; oligonucleotides were synthesized: 1) to identify by PCR a cDNA
library that contained the sequence of
interest, and 2) for use as probes to isolate a clone of the full-length
coding sequence for PR0852.
PCR primers (forward and reverse) were synthesized:
forward PCR nrit~,er 1 5'-CGCAGAAGCTACAGATTCTCG-3' (SEQ ID N0:197)
forward PCI~primer 2 5'-GGAAATTGGAGGCCAAAGC-3' (SEQ ID N0:198)
forward primer 3 5'-GGATGTAGCCAGCAACTGTG-3' (SEQ ID N0:199)
PCR
fotwvard primer 4 5'-GCCTTGGCTCGTTCTCTTC-3' (SEQ ID N0:200)
PCR
forward PCR primer 5 5'-GGTCCTGTGCCTGGATGG-3' (SEQ ID N0:201)
reverse PCR,primer 1 5'-GACAAGACTACCTCCGTTGGTC-3' (SEQ ID N0:202)
reverse PCR rimer 2 5'-TGATGCACAGTTCAGCACCTGTTG-3' (SEQ ID N0:203)
p
Additionally,a synthetic oligonucleodde hybridization probe was constructed
from the consensus DNA34364
sequence which had the following rntcleotide sequence
hvbridization~robe
5'-CGCTCCAAGGGCTTTGACGTCACAGTGAAGTACACACAAGGAAGCTG-3' (SEQ ID N0:204)
In order to screen several libraries for a source of a full-length clone, DNA
from the libraries was screened
by PCR amplification with the PCR primer pair identified above. A positive
library was then used to isolate clones
encoding the PR0852 gene using the probe oligonucleotide and one of the PCR
primers. RNA for construction of
the cDNA libraries was isolated from human fetal kidney tissue (LIB228).
DNA sequencing of the clones isolated as described above gave the full-length
DNA sequence for PR0852
[herein designated as UNQ418 {DNA45493-I349)) (SEQ ID N0:195) and the derived
protein sequence for PR0852.
The entire nucleotide sequence of UNQ418 (DNA45493-1349) is shown in Figure 72
(SEQ ID N0:195).
Clone UNQ418 (DNA45493-1349) contains a single open reading frame with an
apparent translational initiation site
at nucleotide positions 94-96 and ending at the stop codon at nucleotide
positions 16748-1650 (Figure 72). The
predicted polypeptide precursor is 518 amino acids long (Figure ?3). The full-
length PRO852 protein shown in
Figure 73 has an estimated molecular weight of about 56.180 daltons and a pI
of about 5.08. Analysis of the full-
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length PRO852 sequence shown in Figure 73 (SEQ ID N0:196) evidences the
presence of the following: a signal
peptide from about amino acid I to about amino acid 20, a transmembrane domain
from about amino acid 466 to
about amino acid 494, potential N-glycosylation sites from about amino acid
170 to about amino acid 173 and about
amino acid 366 to about amino acid 369, leucine zipper sequence pattern blocks
from about amino acid 10 to about
amino acid 3I and from about amino acid 197 to about amino acid 2I8 and blocks
of amino acids having sequence
homology to eukaryotic and viral aspartyl proteases from about amino acid 109
to about amino acid 1 I8; from about
amino acid 252 to about amino acid 26I and from about amino acid 298 to about
amino acid 310: Clone UNQ418
(DNA45493-1349) has been deposited with ATCC on April 28, 1998 and is assigned
ATCC deposit no: 209805.
Analysis of the amino acid sequence of the full-length PR0852 polypeptide
suggests that it possesses
significant sequence similarity to various protease proteins, thereby
indicating that PR0852 may be a novel protease
protein or homolog thereof. More specifically, an analysis of the Dayhoff
database (version 35.45 SwissProt 35)
evidenced signifecant homology between the PRO852 amino acid sequence and the
following Dayhoff sequences,
PEPC HUMAN> S66516, S66517, PEPS CHICK, CATD HUMAN, P 874207, CARP YEAST,
PEP2_RABIT,
CATS HUMAN and RENI MOUSE.
IS EXAMPLE 33: Isolation of cpNA Clones Encoding Hqman PR0853
A consensus sequetxe was obtained relative to a variety of EST sequences as
described in Example l above,
wherein the consensus sequence obtained is herein designated DNA43050: Based
on the DNA43050 consensus
sequence, oligonucieotides were synthesized: 1) to identify by PCR a cDNA
library that contained the sequence of
irnerest, and 2) for use as probes to isolate a clone of the full-length
coding sequence for PR0853.
Forward and reverse PCR primers were synthesized:
forward PCR Diner 5'-CTTCATGGCCTTGGACTTGGCCAG-3' (SEQ ID N0:207)
reverse PCR primer 5'-ACGCCAGTGGCCTCAAGCTGGTTG-3' (SEQ ID N0:208)
Additionally, a synthetic oligonucleotide hybridization probe was constructed
from the consensus DNA43050
sequence which had the following nucleotide sequence
hv,~~ridization probe
5'-CTTTCTGAGCTCTGAGCCACGGTTGGACATCCTCATCCACAATGC-3' (SEQ ID N0209)
In order to screen several Hbraties for a source of a full-length clone; DNA
from the libraries was screened
by PCR atnplificarion with otte of the PCR primer pairs identified above. A
positive library was then used to isolate
clones encoding the PR0853 gene using the probe oiigonucleotide and one of the
PCR primers. RNA for construction
of the cDNA libraries was isolated from human fetal Iddney tissue (LIB228).
DNA sequencing of the clones isolated as described above gave the full-length
DNA sequence for PR0853
(herein desigmted as UNQ419 (DNA48227-1350)] (SEQ ID N0:205) and ttie derived
protein sequence for PR0853.
The entire nucleotide sequence of UNQ419 (DNA48227-1350) is shown in Figure 74
(SEQ ID N0:205).
Clone UNQ419 (DNA48227-1350) oorttaains a single open reading frame with an
apparent translational initiation site
at nucleotide positions I28-130 and ending at the stop codon at nucleotide
positions 1259-1261 (Figure 74). The
predicted polypeptide precursor is 377 amino acids long (Figure 75). 'Ihe full-
length PR0853 protein shown in
Figure 75 has an estimated molecular weight of about 40,849 daltons and a pI
of about 7.98. Important regions of
the amino acid sof PR0853 include tht signal peptide, corresponding to amino
acids from about 1 to about
I6 of SEQ >D N0:206, the glycosaminoglycan attachment site, corresponding to
amino acids from about 46 to about
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49 of SEQ ID N0;206, and two sequences typical of the short-chain alcohol
dehydrogenase family, corresponding
to amino acids from about 37 to about 49 and about 114 to about 124 of SEQ ID
N0:206, respectively. Clone
UNQ419 (DNA48227-1350) has been deposited with ATCC and is assigned ATCC
deposit no. 209812.
EJ,~AMPLE 34: Isolation of cDNA Clones Encoding Human PR0860
S A consensus sequence was obtained relative to a variety of EST sequences as
described in Example I above,
wherein the consensus sequence obtained is herein designated DNA38137. Based
on the DNA38137 consensu
sequence, oligonucleotides were synthesized: I) to identify by PCR a cDNA
library that contained the sequence of
interest, and 2) for use as probes to isolate a clone of the full-length
coding sequence for PR0860.
Forward and reverse PCR primers were synthesized:
. forward~CR primer 5'-GAAGGGACCTACATGTGTGTGGCC-3' (SEQ ID N0:212)
reverse PCR gt~imer S'-ACTGACCTTCCAGCTGAGCCACAC-3' (SEQ ID N0:213)
Additionally, a synthetic oligonucleotide hybridization probe was constructed
from the consensus DNA406S4
sequence which had the following nucleotide sequence
hybridization rya obe
S'-AGGACTACACGGAGCCTGTGGAGCTTCTGGCTGTGCGAATTCAGCTGGAA-3'
(SEQ ID N0:214)
In order to screen several libraries for a source of a full-length clone, DNA
from the libraries was screened
by PCR amplification with one of the PCR primer pairs identified above. A
positive library was then used to isolate
clones encoding the PR0860 gene using the probe oligonucleotide and one of the
PCR primers. RNA for
construction of the cDNA libraries was isolated from human fetal lung tissue
(LIB26).
DNA sequencing of the clones isolated as described above gave the full-length
DNA sequence for PR0860
[herein designated as UNQ421 (DNA41404-1352)J (SEQ ID N0:210) and the derived
protein sequence for PR0860.
The entire nucleotide sequence of UNQ421 (DNA47404-1352) is shown in Figures
76A-B (SEQ ID
N0:210). Clone UNQ421 (DNA4I404-1352) contains a single open reading frame
with an apparent translatianal
2S initiation site at nucleotide positions S8-60 and ending at the stop codon
at nucleotide positions 3013-30iS (Figures
76A-8). The predicted polypeptide precursor is 985 amino acids tong (Figure
77). The full-length PR0860 protein
shown in Figure 77 has an estimated molecular weight of about 10S,336 daltons
and a p1 of about 6.5S. Important
regions of the amino acid sequence. of PR0860 include the transmembrane region
corresponding to about amino acids
448-467, the extracellular domain, corresponding to amino acids about 1-447,
several N-glycosylation sites, numerous
N-myristoylation sites and a sequence typical of phosphotyrosine interaction
domain proteins.. Clone UNQ421
(DNA41404-1352) has been deposited with ATCC and is assigned ATCC deposit no.
209844.
SAMPLE 35: Isolation of cDNA Clones ncoding Human PR0846
A consensus sequence was obtained relative to a variety of EST sequences as
described in Example 1 above,
wherein the consensus sequence obtained is herein designated DNA39949. Based
on the DNA39949 consensus
sequence, oligonucleotides were synthesized: 1) to identify by PCR a cDNA
library that contained the sequence of
interest, and 2) for use as probes to isolate a clone of the full-length
coding sequence for PR0846.
Forward and reverse PCR primers were synthesized:
forward PCR timer S'-CCCTGCAGTGCACCTACAGGGAAG-3' (SEQ ID NO:2I7)
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averse P ~t primer 5'-CTGTCTTCCCCTGCTTGGCTGTGG-3' (SEQ 1D N0:218)
Additionally, a synthetic oligonucleotide hybridization probe was constructed
from the consensus DNA39949
sequence which had the following nucleotide sequence
hybridization probe
S '-GGTGCAGGAAGGGTGGGATCCTCTTCTCTCGCTGCTCTGGCCACATC-3'
S (SEQ ID N0:219)
In order to streets several libraries for a source of a full-length clone, DNA
from the libraries was screened
by PCR amplification with one of the PCR~ primer pairs identified above. A
positive library was then used to isolate
clones encoding the PR0846 gene using the probe oligonucleotide and one of the
PCR primers. RNA for
construction of the cDNA libraries was isolated from human fetal kidney
itissue (LIB227).
DNA sequencing of the clones isolated as described above gave the full-length
DNA sequence for PR0846
(herein designated as UNQ422 {DNA44196-1353)] (SEQ 1D N0:215) and the derived
protein scqtience for PR0846.
The entire nucleotide sequence of UNQ422 (DNA44196-1353) is shown in Figure 78
(SEQm N0:2I5).
Clone UNQ422 (DNA4419tr1353) contains a single open reading frame with an
apparent translational initiation site
at nucleotide positions 25-27 and ending at the stop codon at nucleotide
positions 1021-1023 (Figure 78). The
predicted poiypeptide precursor is 332 amino acids long (Figure 79). The full-
length PR0846 protein shown in
Figure 79 has an estimated molecular weight of about 36,143 daltons and a pI
of about 5.89. Important regions of
the amino acid sequence of PR0846 include the signal peptide, the
transmetnbrane domain, an N-glycosylation site,
a atypical of fibrinogen beta and gamma chains C-terminal domain, and a
sequence typical of Ig like V-type
dwmain as shown in Fgure 79. Clone UNQ422 t'DNA44196-1353) rtes been deposited
with ATCC and is assigned
ATCC deposit no.-209847_
EXAMPLE 36. Lsolation of cDNA Clones Encoding Human PR0862
A consensus sequence was obtained relative to a variety of EST sequences as
described in Example 1 above,
wherein the consensus sequence obtained is herein designated DNA47370. Based
on the DNA47370 consensus
sequence, oligonucleotides were synthesized: I) to identify by PCR a cDNA
library that contained the sequence of
intcrcst, and 2) for use as probe to isolate a clone of the full-length
codittl; sequence for PR0862.
Forward and teversc pCR primers were synthesized:
forward PCR~,rriLn~r 5'GGGATCATG'TTGT'I'GGCCCTGGTC-3' (SEQ ID N0:222)
reverse PCR primer 5'-GCAAGGCAGACCCAGTCAGCCAG-3' (SEQ ID N0:223)
Additionally, a synthcac oligonucleotide~ hybridization probe was constructed
from the consensus DNA47370
sequetxc which had the following nucleotide sequence
ltyt~n~dization nrobc
5'-CTGCCTGCTACCCTCCAAGTGAGGCCAAGCTCTACGGTCGTTGTG-3'
(SEQ ID N0:22~
in order to screen severer h'braries for a soutrx of a full-length clone, DNA
from the libraries was screened
by PCR ampIifi~rion with one of dbe PCR primer parts identified above. A
positive library was then used to isolate
clones encoding the PR08b2 gene using site probe oligonuclcotidc and orre of
the PCR primers. RNA for
cotzsuuaroa of the cDNA Libraries was isolated from human pa~reas tissue (I
IB55).
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DNA sequencing of the clones isolated as described above gave the full-length
DNA sequence for PR0862
(herein designated as UNQ424 (DNA52187-1354)] (SEQ ID N0:220) and the derived
protein sequence for PRO862.
The entire nucleotide sequence of UNQ424 (DNA52187-1354) is shown in Figure 80
(SEQ 1D N0:220},
Clone UNQ424 (DNA52I87-1354) contains a single open reading frame with an
apparent translational initiation site
at nucleotide positions 41012 and ending at the stop codon at nucleotide
positions 848-850 (Figure 80). The
predicted polypeptide precursor is 146 amino acids long (Figure 81). The full-
length PR0862 protein shown in
Figure 81 has an estimated molecular weight of about 15,430 daltons and a pI
of about 5.05. Important regions of
the amino acid sequence of PR0862 include the signal peptide, an N-
myristoylation site, and sequences having
similarity to region to Alpha-lactalbuminllysozyme C proteins as shown in
Figure 81. Clone UNQ424 (DNA52I87-
1354) has been deposited with the ATCC and is assigned ATCC deposit no.
209845.
10:
EXAMPLE 37: Isolation of cDNA Clones Encodi~ Human PR0864
A consensus sequence was obtained relative to a variety of EST sequences as
described in Example 1 above,
wherein the consensus sequence obtained is herein designated DNA40666. Based
on the DNA40666 consensus
sequence, oligonucleotides were synthesized: l) to identify by PCR a cDNA
library that contained the sequence of
IS interest, and 2) for use as probes to isolate a clone of the full-length
coding sequence for PR08b4.
Forward and reverse PCR-primers were synthesized:
Forward PCR primer 5'-GCTGCAGCTGCAAATTCCACTGG-3' (SEQ ID N0:22?)
zgverse PCR_primer 5'-TGGTGGGAGACTGTTTAAATTATCGGCC-3' (SEQ ID N0:228)
Additionally, a synthetic oligonucleotide hybridization probe was constructed
from the consensus DNA40666
20 sequence which had the following nucleotide sequence
hybridization pmbe
5'-TGCTTCGTCAAGTGCCGGCAGTGCCAGCGGCTCGTGGAGTT-3'
(SEQ ID N0:229)
1n order to screen several libraries for a source of a full-length clone, DNA
from the Libraries was screened
25 by PCR amplification with orte of the PCR primer pairs identified above. A
positive library was then used to isolate
clones encoding the PR0864 gene using the probe oligonucleotide and one of the
PCR primers. RNA for
construction of the cDNA libraries was isolated from human fetal brain tissue
(L1B153).
DNA sequencing of ttte clones isolated as described above gave the full-length
DNA sequence for PR0864
[herein designated as UNQ426 (DNA48328-1355)] (SEQ ID N0:225) and the derived
protein sequence for PR0864.
30 The entire nucleotide sequence of UNQ426 (DNA48328-1355) is shown in Figure
82 (SEQ ID N0:225).
Clone UNQ426 (DNA48328-1355) contains a single open reading frame with an
apparent translational initiation site
at nucleotide positions 37-39 and ending at the stop codon at nucleotide
positions 1090-1092 (Figure 82). The
predicted poLypeptide precursor is 351 amino acids long (Figure 83). The full-
length PR0864 protein shown in
Figure 83 has an estimated molecular weight of about 39,052 and a p1 of about
8.97. Important regions of the amino
35 acid sequence of PR0864 include the signal peptide, two N-glycosylation
sites, a Wnt-1 family signature sequence,
and sequence regions homologous to Wnt-1 family proteins as shown in Figure
83. Clone UNQ426 (DNA48328-
1355) has been deposited with ATCC and is assigned ATCC deposit no. 209843.
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EXAMPLE 38: Isolation of cDNA Clones Eneod~pg Human PR0792
A conserzsus sequence was obtained relative to a variety of EST sequences as
described in Example 1 above,
wherein the consensus sequence obtained is herein designated DNA3810ti. Based
on the DNA38106 consensus
sequence, oligonucleotides were synthesized: 1) to identify by PCR a eDNA
library that contained the sequence of
interest, and 2) for use as probes to isolate a clone of the full-length
coding sequence for PR0792.
A pair of PCR primers (forward and reverse) were synthesized:
forward PCR rp imer 5'-GCGAGAACTGTGTCATGATGCTGC-3' (SEQ ID N0:232)
reverse PCR primer 5'-GTTTCTGAGACTCAGCAGCGGTGG-3' (SEQ ID N0:233)
Additionally, a synthetic oligonucleotide hybridization probe was constructed
from the consensus DNA38106
sequence which had the following nucleotide sequence
Itybri~,iza~,gn probe
5'-CACCGTGTGACAGCGAGAAGGACGGCTGGATCTGTGAGAAAAGGCACAAC-3' (SEQ ID N0:234)
In order to screen several libraries for a source of a full-length clone, DNA
from the libraries was screened
by PCR amplification with the PCR primer pair identified above. A positive
library was then used to isolate clones
encoding the PR0792 gene using the probe oligonucleotide and one of the PCR
primers. RNA for construction of
the cDNA libraries was isolated from human bone marrow tissue (LIB255).
DNA sequencing of the clones isolated as described above gave the full-length
DNA sequence for PR0792
(herein designated as UNQ431 (DNA56352-1358)] (SEQ ID N0:230) and the derived
protein sequence for PR0792.
The entire nucleotide sequence of UNQ431 (DNA56352-1358) is shown in Figure 84
(SEQ ID N0:230).
Clone UNQ431 (DNA56352-1358) contains a single open reading frame with an
apparent uanslational initiation site
at nucleotide positions 67-69 and ending at the stop codon at nucleotide
positions 946-948 (Figure 84). The predicted
polypeptide precursor is 293 amino acids long (Figure 8S). The full-length
PR0792 protein shown in Figure 85 has
an estimated molecular weight of about 32,562 daltons and a pI of about G.53.
Analysis of the full-length PR0792
sequence shown in Figure 85 (SEQ ID N0:231) evidences the presence of the
following: a type I1 transmembrane
domain from about amino acid 31 to about amino acid 54; potential N-
glycosylation sites from about amino acid 73
to about amino acid 76 ar>d from about amino acid 159 to about amino acid 162,
a leueine zipper amino acid sequence
pattern from about amino acid 102 to about amino acid 123, potential N-
myristolation sites from about amino acid
18 to about amino acid 23, from about amino acid 133 to about amino acid 138
and from about amino acid 242 to
about amino acid 247 and a C-type lectin domain signature block from about
amino acid 2b4 to about amino acid 287.
Clone UNQ431 (DNA56352-1358) has been deposited with ATCC on May 6, 1998 and
is assigned ATCC deposit
no. 209846.
Analysis of the amino acid sequence of the full-length PR0792 polypeptide
suggests that it possesses
significant sequence similarity to the CD23 protein, thereby indicating that
PR0792 may be a novel CD23 homolog.
More specifically, an analysis of the Dayhoff database (version 35.45
SwissProt 35) evidenced significant homology
between the PR0792 amino acid sequence and the following Dayhoff sequences,
S34198, A07100 1, A05303_l,
P 841689, P P82839, A10871_l, P Ri2796, P 847199, A46274 and P 832188.
EXAMP1_.F_ 39: Isolation of cDNA Clones Encoding Human PR08G6
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A consensus sequence was obtained relative to a variety of EST sequences as
described in Example 1 above,
wherein the consensus sequence obtained is herein designated DNA44708. Based
on the DNA44708 consensus
sequence, oligonucleotides were synthesized: 1).to idemify by PCR a cDNA
library that contained the sequence of
interest, and 2) for use as probes to isolate a clone of the full-length
coding sequence for PR0866.
PCR primers (forward and reverse) were synthesized:
forward PCR Rrimgr 1 5'-CAGCACTGCCAGGGGAAGAGGG-3' (SEQ ID N0:237)
forward PCR primer 2 5'-CAGGACTCGCTACGTCCG-3' (SEQ ID N0:238)
forward PCR primer 3 5'-CAGCCCCTTCTCCTCCTTTCTCCC-3' (SEQ ID N0:239)
reverse PCR primer 1 5'-GCAGTTATCAGGGACGCACTCAGCC-3' (SEQ ID N0:240)
reverse PCR primer 2 5'-CCAGCGAGAGGCAGATAG-3' (SEQ ID N0:241)
reverse PCR primer 3 5'-CGGTCACCGTGTCCTGCGGGATG-3' (SEQ ID N0:242}
Additionally, a synthetic oligonucleotide hybridization probe was constructed
from the consensus DNA44708
sequence which had the following nucleotide sequence
hybridization probe
5'-CAGCCCCTTCTCCTCCTTTCTCCCACGTCCTATCTGCCTCTC-3' (SEQ ID NO:243)
In order to screen several libraries for a source of a full-length clone, DNA
from the libraries was screened
by PCR amplification with one of the PCR primer pairs identified above. A
positive library was then used to isolate
clones encoding the PR0866 gene using the probe oligonucleotide and one of the
PCR primers. RNA for
construction of the cDNA libraries was isolated from human fetal kidney tissue
{LIB228).
DNA sequencing of the clones isolated as described above gave the full-length
DNA sequence for PR0866
[herein designated as UNQ435 (DNA53971-1359)) (SEQ ID N0:235} and the derived
protein sequence for PR0866.
The entire nucleotide sequence of UNQ435 (DNA53971-1359) is shown in Figure 86
(SEQ ID NO:235).
Clone UNQ435 {DNA53971-1359) contains a single open reading frame with an
apparent translationa3 initiation site
at nucleotide positions 275-277 and ending at the stop codon at nucleotide
positions 1268-1270 (Figure 86). The
predicted polypeptide precursor is 331 amino acids long (Figure 87). The full-
length PR0866 protein shown in
Figure 8? has an estimated molecular weight of about 35,844 daltons and a pI
of about 5.45. Analysis of the full-
length PR0866 sequence shown in Figure 87 (SEQ ID NO:236) evidences the
presence of the following: a signal
peptide from about amino acid 1 to about amino acid 26. Clone UNQ435 (DNA5397I-
1359) has been deposited with
ATCC on April 7, 1998 and is assigned ATCC deposit no. 209750.
Analysis of the amino acid sequence of the full-length PR0866 polypeptide
suggests that it possesses
significant sequence similarity to the mindinlspondin family of proteins,
thereby indicating that PR0866 may be a
novel mindin homolog. More specifically, an analysis of the. Dayhoff database
(version 35.45 SwissProt 35)
evidenced significant homology between the PR0866 amino acid sequence and the
following Dayhoff
sequences,AB006085_l, AB006084_l, AB006086 I, AF017267~1, CWU422I3~1,
AC004160_l, CPM1CRP_l,
S49108, A48569 and 146687.
EXAM,~'LE 40: j~~~~n of cDNA Clones Encoding Human PR0871
A consensus sequence was obtained relative to a variety of EST sequences as
described in Example 1 above,
wherein the consensus sequence obtained is herein designated DNA40324. Based
on the DNA40324 consensus
sequence, oligonucleotides were synthesized: I) to identify by PCR a cDNA
library that contained the sequence of
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interest, and 2) for use as probes to isolate a clone of the full-length
coding sequence for PR0871.
PCR primers (forward and reverse) were synthesized:
forward PCR grimed S'-TGCGGAGATCCTACTGGCACAGGG-3' (SEQ ID N0:246)
forward FCR p~im~ 2 S'-CGAGTTAGTCAGAGCATG-3' (SEQ ID N0:247)
forward PCR primer 3 S'-CAGATGGTGCTGTTGCCG-3' (SEQ ID N0:248)
S reverse PCR urimer i S'-CAACTGGAACAGGAACTGAGATGTGGATC-3' (SEQ ID N0:249)
reverse PAR primer 2 S'-CTGGTTCAGCAGTGCAAGGGTCTG-3' (SEQ ID N0:2S0)
reverse PCR primer 3 S'-CCTCTCCGATTAAAACGC-3' (SEQ ID N0:2S1)
Additionally, a synthetic oligonucleotide 'hybridization probe was constmcted
from the consensus DNA40324
sequence which had the following nucleotide sequence
hybridization g~robe
5'-GAGAGGACTGGTTGCCATGGCAAATGCTGGTTCTCATGATAATGG-3' (SEQ ID N0:2S2)
In order to screen several libraries for a source of a full-length clone, DNA
from the libraries was screed
by PCR amplification with one of the PCR primer pairs idernified above. A
positive library was then used to isolate
clones encoding the PR0871 gene using the probe oligonucleotide and one of the
PCR primers. RNA .for
construction of the cDNA libraries was isolated from human fetal kidney tissue
{LIB227).
DNA sequencing of the clones isolated as described above gave the full-length
DNA sequence for PR0871
[herein designated as UNQ438 (DNA50919-136I)] {SEQ ID N0:244) and the derived
protein sequence for PR0871.
The entire nucleotide sequence of UNQ438 {DNA50919-1361) is shown in Figure 88
(SEQ ID N0:244).
Clone UNQ438 {DNA50919-1361) contains a single open reading frame with an
apparent translational initiation site
at nucleotide positions 191-193 and ending at the stop codon at nucleotide
positions 1607-1609 (Figure 88). The
predicted polypeptide precursor is 472 amino acids long (Figure 89). The full-
length PR0871 protein shown in
Figure 89 has an estimated molecular weight of about 53,847 daltons and a pI
of about 5.75. Analysis of the full
length PR0871 sequence shown in Figure 89 {SEQ ID N0:245) evidences the
presence of the following: a signal
peptide from about amino acid 1 to about amino acid 2l, potential N-
glycosylation sites from about amino acid 109
to about amino acid 112 and from about amino acid 201 to about amino acid 204,
a cyclophilin-type peptidy-prolyl
cis-cans isomerase signature sequence from about amino acid 49 to about amino
acid 66 and regions that are
homologous to cyclophilin-type peptidy prolyl cis-traps isomerases from about
amino acid 96 to about amino acid 140,
from about amino acid 49 to about amino acid 89 and from about amino acid 22
to about amino acid 51. Clone
UNQ438 (DNA50919-1361) has been deposited with ATCC on May 6, 1998 and is
assigned ATCC deposit no.
209848.
Analysis of the amino acid sequence of the full-length PR0871 polypeptide
suggests that it possesses
significant sequence similarity to the cyclophilin family of proteins, thereby
indicating that PR0871 may be a novel
cyclophiiin protein family member. More specifically; an analysis of the
Da;yhoff database (version 3S.4S SwissProt
35) evidenced significant homology between the PR0871 amino acid sequence and
the following Dayhoff sequences,
SPBC16H5 S, S64705, YALS_SCHPO, CYP4 CAEEL, CELC34D4- 7, CYPA CAEEL,
HUMORF006_1,
CYPI MYCTU, AF043642 1 and HSSRCYP 1.
IXAMP1 E 41: Isolat~n of cDNA (',ones Encoding I~uman PR0873
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A consensus sequence was obtained relative to a variety of EST sequences as
described in Example I above.
wherein the consensus sequence obtained is herein designated DNA39621. Based
on the DNA3962I consensus .
sequence, oligonucleotides were synthesized: I ) to identify by PCR a cDNA
library ti~at contained the sequence of
interest, and 2) for use as probes to isolate a clone of the full-length
coding sequence for PR0873.
A pair of PCR primers (forward and reverse) were synthesized:
]'orward PCR primeF 5'-AGGTGCCTGCAGGAGTCCTGGGG-3' (SEQ ID N0:255)
reverse PCR primer 5'- CCACCTCAGGAAGCCGAAGATGCC-3' (SEQ ID N0:256)
Additionally, a synthetic oligonucleotide hybridization probe was constructed
from the consensus DNA39621
sequence which had the following nucleotide sequence:
hybridization probe
.. 5'-GA.ACGGTACAAGTGGCTGCGCTTCAGCGAGGACTGTCTGTACCTG-3' (SEQ ID N0:257)
1n order to screen several libraries for a source of a full-length clone, DNA
from the libraries was screened
by PCR amplification with the PCR primer pair identified above. A positive
library was then used to isolate clones
encoding the PR0873 gene using the probe oligonucleotide and one of the PCR
primers. RNA for construction of
the cDNA libraries was isolated from human fetal liver tissue (LIB229).
IS DNA sequencing of the clones isolated as described above gave the foil-
length DNA sequence for PR0873
[herein designated as UNQ440 (DNA44179-1362)] (SEQ ID N0:253) and the derived
protein sequence for PR0873.
The entire nucleotide sequence of UNQ440 (DNA44179-1362) is shown in Figure 90
(SEQ ID N0:253).
Clone UNQ440 (DNA44179-1362) contains a single open reading frame with an
apparent translational initiation site
at nucleotide positions 139-141 and ending at the stop codon at nucleotide
positions 1774-1776 (Figure 90). The
predicted polypeptide precursor is 545 amino acids long (Figure 91). The full-
length PR0873 protein shown in
Figure 91 has an estimated molecular weight of about 58,934 daltons and a pI
of about 9.45. Analysis of the full-
length PR0873 sequence shown in Figure 9I (SEQ ID N0:254) evidences the
presence of the following features:
a signal peptide from about amino acid 1 to about amino acid 29; a
carboxylesterase type-B serine active site at about
amino acid 312 to about amino acid 327; a carboxylesterase type-B signature 2
motif at about amino acid 218 to about
amino acid 228; and three potential N-glycosylation sites at about amino acid
318 to about amino acid 321, about
amino acid 380 to about amino acid 383, and about amino acid 465 to about
amino acid 468. Clone UNQ440
(DNA44I79-1362) has been deposited with ATCC on May 6, 1998 and is assigned
ATCC deposit no. 209851.
Analysis of the amino acid sequence of the full-length PR0873 polypeptide
suggests that it possesses
significant sequence similarity to a human liver carboxylesterase, thereby
indicating that PR0873 may be a novel
carboxylesterase. More specifically, an analysis of the Dayhoff database
(version 35.45 SwissProt 35) evidenced
significant homology between the PR0873 _amino acid sequence and the following
Dayhoff sequences: ES 10 RAT,
GEN12405, ABOlOb33_l, EST4 RAT, A48809, SASB ANAPL, RNU41662_l, RNU22952 1,
BAL RAT,
GEN13522.
EXAMPLE 42: Isolation of cDNA Clones Encodint=_ Human PRO940
A consensus sequence was obtained relative to a variety of EST sequences as
described in Example 1 above,
wherein the consensus sequence obtained is herein designated DNA47442. Based
on the DNA47442 consensus
sequence, oligonucleotides were synthesized: 1 ) to identify by PCR a cDNA
library that contained the sequence of
interest, and 2) for use as probes to isolate a clone of the full-length
coding sequence for PR0940.
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A pair of PCR primers (forward and reverse) were syntltcsizred:
forward PCR primer 5'-CAAAGCCTGCGCCTGGTCTGTG-3' (SEQ 1D N0:260) ,
reverse PCR primer S' TTCTGGAGCCCAGAGGGTGCTGAG-3' (SEQ ID N0:261)
Additionally, a synthuic oligonucleotide hybridization probe was constructed
from the consensus DNA47442
sequence which had the following nucleotide sequence
hybridization p~obe
5'-GGAGCTGCCACCCATTCAAATGGAGCACGAAGGAGAGTTCACCTG-3' (SEQ ID N0:262)
in order to screen several Lbraries for a source of a full-length clone, DNA
from the libraries was screened
by PCR amplification with the PCR primer pair identified above. A positive
library was then used to isolate clones
encoding the PR0940 gene using the probe oligonucJeotide and one of the PCR
primers. RNA for construction of
IO the cDNA libraries was isolated from human fetal liver tissue (LIB229).
DNA sequencing of the clones isolated as described above gave the full-length
DNA sequence for PR0940
[liereim designated as UNQ477 (DNA54002-1367)j (SEQ ID N0:258) and the derived
protein sequence=for PRO940.
The entire nucleotide sequence of UNQ477 (DNA54002-1367) is shown in Figure 92
(SEQ ID N0:258)_
Clone UNQ477 (DNA54002136'~ contains a single open reading frame with an
apparent translational initiation site
IS at nucleotide positions 46-08 and ending at the stop codon at nucleotide
positions 1678-I680 (Figure 92). The
predicted polypeptide preausor is 544 amino acids long (Figure 93). The full-
length PR0940 protein shown in
Figure 93 has an estimated molecxtlar weight of about 60,268 daltons and a pI
of about 9.53. Analysis of the full-
length PR0940 sequence shown in Figure 93 (SEQ ID N0:259) cvidetxa the
presence of the following: a signal
peptide fc~om about amino acid 1 to about amino acid 15, potential
N~lyeosylation sites from about amino acid 100
20 to about amino acid 103, fra~t about amino acid 297 to about aW i~ acid 300
and from about amino acid 306 to about
amino acid 309 and an immunoglobulin and major histocompatibiliry complez
signature sequence block from about
amino acid 365 to about amino acid 37I . Clone UNQ477 (DNA54002-1367) has been
deposited with ATCC on April
7, 1998 and is assigned ATCC deposit no. 209754.
Analysis of the amino acid sequence of the full-length PR0940 polypeptide
suggtsu that it possesses
2S significaru seqt~C similarity w CD33 and the OB binding protein-2. More
specifically, an analysis of the Dayhoff
dataltase (version 35.45 SwissProt 35) evidenced significant homology between
the PR0940 amino acid sequence and
the following Dayhoff sequences, CD33 HUMAN, HSU71382_l, HSU71383_l, D86359 1,
PGBM HUMAN,
MAGS MOUSE, D86983 I, C22B HUMAN, P W01002 and HW2411G 1.
30 SAMPLE 43: Isolation of eDNA Clones ,j_ttcodin>i Human PRO941
A consensus scqtiettoc was oMai>ted relative to a variety of EST sequences as
described in Exarttple I above,
wherein the conscruus seqti~ee obtained is heroin designated DNA359~4I. An EST
sequence proprietary to
Genentectt was employed in the assembly and is herein designated DNA641S
(Figure 96; SEQ ID N0:265). Based
on the DNA35941 conscrrcus sequence, oligonucleotides were synthesized: 1) to
identify by PCR a cDNA library
35 that contained the sequence of interest, and 2). for use as probes to
isolate a clone of the full-length coding sequence
for PR0941.
A pair of PCR primers (forward and reverse) were synthesized:
forward P('.R R~ 5'-C71'GAG'TGTCT'CTGAATCTGCACCC-3' (SEQ iD N0:266)
reverse E~primsr 5'-AAG1'GGTGGAAGCCTCCAGTGTGG-3' (SEQ ID N0:267)
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Additionally, a synthetic oligonucleotide hybridization probe was constructed
from the consensus DNA35941
sequence which had the following nucleotide sequence
hybridization probe
5'-CCACTACGGTATTAGAGCAAAAGTTAAAAACCATCATGGTTCCTGGAGCAGC-3' (SEQ ID N0:268)
In order to screen several libraries for a source of a full-length clone, DNA
from the libraries was screened
by PCR amplification with the PCR primer pair identified above. A positive
library was then used to isolate clones
encoding the PR094I gene using the probe oligonucleotide and one of the PCR
primers. RNA for construction of
the cDNA libraries was isolated from human fetal kidney tissue (LIB227).
DNA sequencing of the clones isolated as described above gave the full-length
DNA sequence for PR0941
[herein designated as UNQ478 (DNA53906-1368)] (SEQ ID N0:263) and the derived
protein sequence for PR0941.
The entire nucleotide sequence of UNQ478 {DNA53906-1368) is shown in Figure 94
(SEQ ID N0:263).
Clone UNQ478 (DNA53906-1368) contains a single open reading frame with an
apparent translational initiation site
at nucleotide positions 37-39 and ending at the stop codon of nucleotide
positions 2353-2355 (Figure 94). The
predicted polypepiide precursor is 772 amino acids long (Figure 95). The full-
length PR0941 protein shown in
Figure 95 has an estimated molecular weight of about 87,002 daltons and a pI
of about 4.64. Analysis of the full-
length PR0941 sequence shown in Figure 95 (SEQ ID N0:264) evidences the
presence of the following: a signal
peptide from about amino acid 1 to about amino acid 21, potential N-
glycosylation sites from about amino acid 57
to about amino acid 60, from about amino acid 74 to about amino add 77, from
about amino acid 419 to about amino
acid 422, from about amino acid 437 to about amino acid 440, from about amino
acid 508 to about amino acid 511,
from about amino acid 515 to about amino acid 518, from about amino acid 516
to about amino acid 519 and from
about amino aad 534 to about amino acid 537, and cadherin extracellular
repeated domain signature sequences from
about amino acid 136 to about amino acid 146 and from about amino acid 244 to
about amino acid 254. Clone
UNQ478 (DNA53906-1368) has been deposited with ATCC on April 7, 1998 and is
assigned ATCC deposit no.
209747.
Analysis of the amino acid sequence of the full-length PR0941 polypeptide
suggests that it possesses
significant sequence similarity to a cadhetin protein, thereby indicating that
PR0941 may be a novel cadberin protein
family member. More specifically, an analysis of the Dayhoff database (version
35.45 SwissProt 35) evidenced
significant homology between the PR0941 amino acid sequence and the following
Dayhoff sequences, I50180,
CADA CHICK, I50178, GEN12782, CADC HUMAN, P W25637, A38992, P 849731, D38992
and 602678.
~,XAMPLE 44: Isolation of cDNA Clones Encoding Human PR0944
A consensus sequence was obtained relative to a variety of FST sequences as
described in Example 1 above,
wherein the cot>sensus seque»ce obtained is herein designated DNA47374. A
variety of proprietary Genentech EST
sequences were employed in the assembly and are shown in Figures 99-107. Based
on the DNA47374 consensus
sequence, oligonucleotictes were synthesized: I) to identify by PCR a cDNA
library that contained the sequence of
interest, and 2) for use as probes to isolate a clone of the full-length
coding sequence for PR0944.
A pair of PCR pritners (forward and reverse) were synthesized:
~gtward PCR~rimer 5'-CGAGCGAGTCATGGCCAACGC-3' (SEQ ID N0:280)
reverse PCR ~m_er_ 5'-GTGTCACACGTAGTCTTTCGCGCTGG-3' (SEQ ID N0:281)
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Additionally, a synthetic oligonucleotide hybridization probe was constructed
from the consensus DNA47374
sequence which had the following nucleotide sequence
hybridig_ation probe
5'-CTGCAGCTGTTGGGCTTCATTCTCGCCTTCCTGGGATGGATCG-?~' (SEQ ID N0:282)
In order to screen several libraries for a source of a full-length clone, DNA
from the libraries was screened
by PCR amplification with the PCR primer pair identified above. A positive
library was then used to isolate clones
encoding the PR0944 gene using the probe oligonucleotide and one of the PCR
primers. RNA for construction of
the cDNA libraries was isolated from human fetal kidney tissue (LIB22?).
DNA sequencing of the clones isolated as described above gave the full-length
DNA sequence for PR0944
[herein designated as UNQ481 (DNA52I85-1370)] (SEQ ID N0:269) and the derived
protein sequence for PR0944.
The entire nucleotide sequence of UNQ481 (DNA52185-1370) is shown in Figure 9?
(SEQ ID N0:269).
Clone UNQ481 (DNA52185-1370) contains a single open reading frame with an
apparent translatioital initiation site
at nucleotide positions 2I9-221 and ending at the stop codon at nucleotide
positions 852-854 (Figure 97). The
predicted polypeptide precursor is 211 amino acids long (Figure 98). The full-
length PR0944 protein shown in
Figure 98 has an estimated moiecuiar weight of about 22,744 dalfons and a pI
of about 8.51. Analysis of the full-
length PR0944 sequence shown in Figure 98 (SEQ ID N0:270) evidences the
presence of the following: a signal
peptide from about amino acid I to about amino acid 21, transmembrane domains
from about amino acid 82 to about
amino acid 102, from about amino acid 1 i 8 to about amino acid I42 and from
about amino acid 161 to about amino
acid 187, a potential N-glycosylation site from about amino acid 72 to about
amino acid 75, a sequence block having
homology to PMP-22IEMP/MP20 family of proteins from about amino acid 70 to
about amino acid 1 l l and a
sequence block having homology to ABC-2 type transport system integral
membrane protein from about amino acid
119 to about amino acid 133. Clone UNQ481 (DNA52185-1370) has been deposited
with ATCC on May 14, 1998
and is assigned ATCC deposit no. 209861.
Analysis of the amino acid sequence of the full-length PR0944 polypeptide
suggests that it possesses
significant sequence similarity to the CPE-R protein, thereby indicating that
PR0944 may be a novel CPE-R
hotnolog. More specifically, an analysis of the Dayhoff database (version
35.45 SwissProt 35) evidenced significant
homology between the PR0944 amino acid sequence and the following Dayhoff
sequences, AB000713_I,
AB000714~l, AF035814'l, AF000959 1, HSU89916 1, EMP2_HUMAN, JC5732, CELF53B3
6, PM22 MOUSE
and CGU49797 1.
EXAMPLE 45: Isolation of cDNA Clones Encoding Hummn PRO983
A consensus sequence was obtained relative to a variety of EST sequences as
described in Example 1 above,
wherein the consensus sequence obtained is herein designated DNA47473. Various
proprietary Genentech EST
sequences were employed in the assembly, wherein those EST sequences are shown
in Figures 110-116. Based on
the DNA4?473 consensus sequence, oligonucieotides were synthesized: 1) to
identify by PCR a cDNA library that
contained the sequence of interest, and 2) for use as probes to isolate a
clone of the full-length coding sequence for
PR0983.
A pair of PCR primers (forward and reverse) were synthesized:
forward PCR primer 5'-GCACCACCGTAGGTACTTGTGTGAGGC-3' (SEQ ID N0:292)
reverse 5'-AACCACCAGAGCCAAGAGCCGGG -3' (SEQ ID N0:293)
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Additionally, a synthetic oligonucleotide hybridization probe was constructed
from the consensus DNA47473
sequence which had the following nucleotide sequence
hybridization probe
5'-CAGCGGAATCATCGATGCAGGGGCCTCAATTAATGTATCTGTGATGTTAC-3' (SEQ 1D N0:294)
In order to screen several libraries for a source of a full-length clone, DNA
from the libraries was screened
by PCR amplification with the PCR primer pair identified above. A positive
library was then used to isolate clones
encoding the PR0983 gene using the probe oligonucleotide and one of the PCR
primers. RNA for construction of
the cDNA libraries was isolated from human bone marrow (LIB256).
DNA sequencing of the clones isolated as described above gave the full-length
DNA sequence for PR0983
[herein designated as UNQ484 (DNA53977-1371)] (SEQ ID N0:283) and the derived
protein sequence for PR0983.
The entire nucleotide sequence of UNQ484 (DNA53977-1371) is shown in Figure
108 (SEQ ID N0:283).
Clone UNQ484 (DNA53977-1371) contains a single open reading frame with an
apparent translational initiation site
at nucleotide positions 234-236 and ending at the stop codon at nucleotide
positions 963-965 (Figure 108). The
predicted polypeptide precursor is 243 amino acids long (Figure 109). The full-
length PR0983 protein shown in
Figure 109 has an estimated molecular weight of about 27,228 daltons and a p1
of about 7.43. Analysis of the full-
length PR0983 sequence shown in Figure 109 (SEQ ID N0:284) evidences the
presence of the following features:
a putative transmembrane domain from about amino acid 224 to about amino acid
239; a potential N-glycosylation
site from about amino acid 68 to about amino acid 71; and three potential N-
myristoylation sites from about amino
acid 59 to about amino acid 64, from about amino acid 64 to about amino acid
69, and froth about amino acid 235
to about amino acid 240. Clog UNQ484 (DNA53977-1371) has been deposited with
ATCC on May 14, 1998 and
is assigned ATCC deposit no. 209862.
Analysis of the amino acid sequence of the foil-length PR0983 polypeptide
suggests that it possesses
significant sequence similarity to the vesicle-associated protein, VAP-33,
thereby indicating that PR0983 may be a
novel vesicle associated membrane protein. More specifically, an analysis of
the Dayhoff database (version 35.45
SwissProt 35) evidenced significant homology between the PR0983 amino acid
sequence and the following Dayhoff
sequences: VP33 APLCA, CELF33D11 12, CELF42G2 2, S50623, YDFC SCHPO, CELF54H5
2, CELZC196 8,
CEF57A10 3, MSP3~GLORO, CEC15H11_l.
EXAMPLE 46: Isolation of eDNA Clones Enco~ng Human PR01057
A consensus sequence was obtained relative to a variety of EST sequences as
described in Example 1 above,
wherein the consensus sequence obtained is herein designated DNA49808. Based
on the DNA49808 consensus
sequence, oligonucleotides were synthesized: 1) to identify by PCR a cDNA
library that contained the sequence of
interest, and 2) for use as probes to isolate a clone of the full-length
coding sequence for PR01057.
PCR primers (forward and reverse) were synthesized:
forward PCR primer 5'-GCATCTGCAGGAGAGAGCGAAGGG-3' (SEQ ID N0:297)
~e_vetse PCR primer 5'-CATCGTTCCCGTGAATCCAGAGGC-3' (SEQ ID N0:298)
Additionally, a synthetic oligonucleotide hybridization probe was constructed
from the comcensus DNA49808
sequence which had the following nucleotide sequence
hybridization vrobe
5'-GAAGGGAGGCCTTCCTTTCAGTGGACCCGGGTCAAGAATACCCAC-3' (SEQ ID N0:299)
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In order to screen several libraries for a source of a full-length clone, DNA
from the libraries was screened
by PCR amplification with the PCR primer pair identified above. A positive
library was then used to isolate clones
encoding the PR01057 gene using the probe oligonucleotide and one of the PCR
primers. RNA for construction of
the cDNA libraries was isolated from human fetal kidney tissue (LIB227).
DNA sequencing of the clones isolated as described above gave the full-length
DNA sequence for PROlOS7
(t>erein designated as UNQS22 (DNAS?2S3-1382)) (SEQ ID N0:29S) and the derived
protein sequence for PR010S7.
The entire nucleotide sequence of UNQS22 (DNAS7253-1382) is shown in Figure
117 (SEQ ID N0:29S).
Clone UNQS22 (DNAS7253-1382) contains a single open reading frame with an
apparent translational initiation site
at nucleotide positions 275-27? and ending at the stop codon at nucleotide
positions 1514-1516 (Figure 117). The
predicted polypeptide precursor is 413 amino acids long (Figure 118). The full-
length PR01057 protein shown in
Figure 118 has an estimated molecular weight of about 47,070 daltons and a p1
of about 9.92. Analysis of the full-
length PROlOS7 sequence shown in Figure 118 (SEQ ID N0:296) evidences the
presence of the following: a signal
peptide from about amino acid 1 to about amino acid 16, potential N-
glycosylation sites from about amino acid 90
to about amino acid 93, from about amino acid 110 to about amino acid 113 and
from about amino acid 193 to about
amino acid 196, a glycosaminoglycan attachment site from about amino acid 236
to about amino acid 239 and a serine
protease histidine-containing active site from about amino acid 165 to about
amino acid 170. Clone UNQ522
(DNA57253-1382) has been deposited with ATCG on May 14, 1998 and is assigned
ATCC deposit no. 209867.
Analysis of the amino acid sequence of the full-length PROIOS7 polypeptide
suggests that it possesses
significant sequence similarity to various protease proteins, thereby
indicating that PROlOS7 may be a novel protease.
More specifically, an analysis of the Dayhoff database (version 3S.4S
SwissProt 3S) evidenced significant homology
between the PROlOS7 amino acid sequence and the following Dayhoff
sequences,TRYE DROER, P 814159,
A69660, EBN1 EBV, S65494, GEN12688, A51084 1, P R99S71, AS7S14 and AF003200 1.
EXI~VIPLE 47: Isolation o~ cDNA Cones Encodi~; Human PR01071
A consensus sequence was obtained relative to a variety of EST sequences as
described in Example 1 above,
2S wherein the consensus sequence obtained is herein designated DNA5303S.
Based on the DNA5303S consensus
sequence, it was determined that that consensus sequence shared significant
sequence identity with Incyte EST clone
no. 2872569, a clone that upon review appeared to encode a full Iengtlt
protein. As such, Incyte EST clone no.
28?2569 was purchased and its insert was obtained and sequenced so as to
confirm the proper sequence. This
sequence is herein designated UNQS28 or DNAS8847-1383.
DNA sequencing of the clone isolated as described above gave the foil-length
DNA sequence for PR01071
[herein designated as UNQS28 (DNA58847-1383)] (SEQ m N0:300) and the derived
protein sequence for PR01071.
The entire rtucieotide sequence of UNQ528 (DNA58847-1383) is shown in Figure
119 (SEQ ID N0:300).
Clone UNQS28 (DNAS8848-1383) contains a single open reading frame with an
apparent translational initiation site
at nucleotide positions 133-13S and ending at the stop codon at nucleotide
positions 1708-1710 (Figure 119). The
3S predicted polypeptide precursor is 525 amino acids long (Figure 120). The
foil-length PR01071 protein shown in
Figure 120 has an estimated molecular weight of about 58,416 daltons and a pI
of about 6.62. Analysis of the fuil-
length PR01071 sequence shown in Figure 120 (SEQ ID N0:301) evidences the
presence of the following: a signal
peptide from about amino acid 1 to about amino acid 2S, a potential N-
glycnsylation site from about amino acid 2S1
to about amino acid 254, a thrombospondin-I homology block from about amino
acid 385 to about amino acid 399
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and von Willibtands factor type C homology blocks from about amino acid 385 to
about amino acid 399, from about
amino acid 445 to about amino acid 459 and from about amino acid 42 to about
amino acid 56. Clone UNQ528
(DNA58847-1383) has been deposited with ATCC on May 20, 1998 and is assigned
ATCC deposit no. 2098?9.
Analysis of the amino acid sequence of the full-length PR01071 polypeptide
suggests that it possesses
significant sequence similarity to the thrombospondin protein, thereby
indicating that PR01071 may be a novel
thrombospondin homolog. More specifically, an analysis of the Dayhoff database
(version 3.45 SwissProt 35)
evidenced significant homology between the PR01071 amino acid sequence and the
following Dayhoff sequences,
AB002364 1, D67076 I, BTPC1NPGN_1, CET13H10 1, CEF25H8,5, CEF53B6 2, CEC26C6
6, HSSEMG_l;
CET21B6 4 and BTY08561 1.
EXAMPLE 48: Isolation of cDNA Clones Encodine Human PR01072
A consensus sequence was obtained relative to a variety of EST sequences as
described in Example 1 above,
wherein the consensus sequence obtained is herein designated DNA53125. Based
on the DNA53125 consensus
sequence, oligonucleotides were synthesized: 1) to identify by PCR a cDNA
library that contained the sequence of
interest, and 2) for use as probes to isolate a clone of the full-length
coding sequence for PR01072.
PCR primers (forward and reverse) were synthesized:
forward PCR primer 5'-CCAGGAAATGCTCCAGGAAGAGCC-3' (SEQ ID N0:305)
reverse PCR primer 5'-GCCCATGACACCAAATTGAAGAGTGG-3' (SEQ ID N0:306)
Additionally, a synthetic oligonucleotide hybridization probe was constructed
from the consensus DNA53125
sequence which had the following nucleotide sequence
hX ridization_probe
5'-AACGCAGGGATCTTCCAGTGCCCTTACATGAAGACTGAAGATGGG-3' (SEQ ID NO:307)
In order to screen several libraries for a source of a full-length clone, DNA
from the libraries was screened
by PCR amplification with the PCR primer pair identified above. A positive
library was then used to isolate clones
encoding the PR01072 gene using the probe oligonucleotide and one of the PCR
primers. RNA for construction of
the cDNA libraries was isolated from human fetal lung tissue (LIB26}.
DNA sequencing of the clones isolated as described above gave the full-length
DNA sequence for PR01072
[herein designated as UNQ529 (DNA58747-1384)) (SEQ ID N0:302) and the derived
protein sequence for PR01072.
The entire nucleotide sequence of UNQ529 (DNA58747-1384) is shown in Figure
121 (SEQ ID N0:302).
Clone UNQ529 (DNA58747-1384) contains a single open reading frame with an
apparent translational.initiation site
at nucleotide positions 65-67 and ending. at the stop codon at nucleotide
positions 1073-1075 (Figure I21). The
predicted polypeptide precursor is 336 amino acids long (Figure 122). The full-
length PR01072 protein shown in
Figure 122 has an estimated molecular weight of about 36,865 daltons and a pI
of about 9.15. Analysis of the full-
length PR01072 sequence shown in Figure 122 (SEQ ID N0:303) evidences the
presence of the following: a signal
peptide from about amino acid 1 to about amino acid 21, short-chain alcohol
dehydrogenase protein homology blocks
from about amino acid 134 to about amino acid 144, from about amino acid 44 to
about amino acid 56 and from about
amino acid 239 to about atnitio acid 248 and potential N-glycosylation sites
from about amino acid 212 to about amino
acid 215 and from about amino acid 239 to about amino acid 242. Clone UNQ529
(DNA58747-1384) has been
deposited with ATCC on May 14, 1998 and is assigned ATCC deposit no. 209868.
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Analysis of the amino acid sequence of the full-length PR01072 polypeptide
suggests that it possesses
significant sequence sinlilariry to the reductase family of proteins, thereby
indicating that PROI072 tray be a novel
reductase. More specifically, an analysis of the Dayhoff database (version
35.45 SwissProt 35) evidenced significant
homology between the PR01072 amino acid sequence and the following Dayhoff
sequences, P W03198, P W15759>
P_R60800, MTV037'3, CEC15H11 6, ATAC00234314, MTV022~13, SCU43704_1, OXIR
STRAT AND
CELCOI G8 3.
EXAMPLE 49: Isolation of cDNA Clones Encoding Human PR01075
A consensus sequence was obtained relative to a variety of EST sequences as
described in Example 1 above,
wherein the consensus sequence obtained is herein designated DNA34363. Based
on the DNA34363 sequence,
oligonucleotides were synthesized: 1) to identify by PCR a cDNA library that
contained the sequence of interest, and
2} for use as probes to isolate a clone of the full-length coding sequence for
PRO10'75.
PCR primers (forward and reverse) were synthesized:
forward,~CR ~rimel 5'-TGAGAGGCCTCTCTGGAAGTTG-3' (SEQ ID N0:312)
forward PCR primer_ 5'-GTCAGCGATCAGTGAAAGC-3' (SEQ ID N0:3I3)
forward PCR ,primer 5'-CCAGAATGAAGTAGCTCGGC-3' (SEQ ID N0:314)
forward PCR timer 5'-CCGACTCAAAATGCATTGTC-3' (SEQ ID N0:315)
forward PCR rn imer 5'-CATTTGGCAGGAATTGTCC-3' (SEQ ID N0:31b}
forward PCR Timer 5'-GGTGCTATAGGCCAAGGG-3' (SEQ ID N0:317)
reverse PCR grimer 5'-CTGTATCTCTGGGCTATGTCAGAG-3' (SEQ ID N0:318)
reverse PCR primer 5'-CTACATATAATGGCACATGTCAGCC-3' (SEQ ID N0:319)
Additionally, a synthetic oligonucleotide hybridization probe was constructed
from the consensus DNA34363
sequence which had the following nucleotide sequence
ridi~ation probe
5'-CGTCTTCCTATCCTTACCCGACCTCAGATGCTCCCTTCTGCTC:CTG-3' (SEQ ID N0:320}
In order to screezt several libraries for a source of a full-length clone; DNA
from the libraries was screened
by PCR atnpii5cation with the PCR primer pair identified above. A positive
library was then used to isolate clones
encoding the PRO1075 gene using the probe oligonucieotide and one of the PCR
primers. RNA for construction of
the cDNA libraries was isolated from human skin tumor tissue (LIB324).
DNA sequencing of the clones isolated as described above gave the full-length
DNA sequence for PR01075
(herein designated as UNQ532 (DNA57689-I385)] (SEQ ID N0:308) and the derived
protein sequence for PROI075:
TI~ entire rntcleotide sequence of UNQ532 (DNA57689-1385) is shown in Figure
124 (SEQ ID N0:308).
Clone UNQ532 (DNA57689-1385) contains a single open reading frame with an
apparent uanslational initiation site
at nucleotide positions 137-139 and ending at the stop codon at nucleotide
positions 1355-1357 (Figure 124). The
predicted polypeptide precursor is 406 amino acids long (Figure 125). The full-
length PR01075 protein shown in
Figure 125 has an estimated molecular weight of about 46,927 daltons and a p1
of about 5.21. Analysis of the full-
length PR01075 sequence shown in Figure 125 (SEQ ID N0:309) evidences the
presence of the following: a signal
peptide from about amino acid 1 to about amino acid 29, an endoplasraic
reticulum targeting sequence from about
amino acid 403 to about amino acid 406, a tyrosine kinase phosphorylation site
from about amino acid 203 to about
amino acid 211 and a sequence block having homology to the thioredoxin family
of proteins from about amino acid
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50 to about amino acid 66. Clone UNQ532 (DNA57689-1385) has been deposited
with ATCC on May 14, 1998 and
is assigned ATCC deposit no. 209869.
Analysis of the amino acid sequence of the full-length PR01075 polypeptide
suggests that it possesses
significant sequence similarity to protein disulfide isomerase, thereby
indicating that PR01075 may be a novel protein
disulfide isotnerase. More specifically, an analysis of the Dayhoff database
(version 35.45 SwissProt 35) evidenced
significant homology between the PR01075 amino acid sequence and the following
Dayhoff sequences,
CELC30H7 2, CELC06A6 3, CELF42G8 3, S57942, ER72 CAEEL, CELC07A12 3, CEH06001
4 and
P 851696.
EXAMPLE 50: lsolatio~t of cDNA Cion~ Encoding Human PR0181
A cDNA sequence isolated in the amylase screen described in Example 2 above
was found; by BLAST and
FastA sequence alignment, to have sequence homology to a nucleotide sequence
encoding the cornichon protein. This
cDNA sequence is herein designated DNA13242 (Figure 130; SEQ ID N0:323). Based
on the sequence homology,
oligonucleotide probes were generated from the sequence of the DNA13242
molecule and used to screen a human
placenta (LIB89) library prepared as described in paragraph 1 of Example 2
above. The cloning vector was pRKSB
(pRKSB is a precursor of pRKSD that does not contain the SfiI site; see,
Holmes et al., Science, 253:1278-1280
(1991)), and the cDNA size cut was less than 2800 bp.
The oligonucleotide probes employed included:
forward PCR primer 5'-GTGCAGCAGAGTGGCTTACA-3' (SEQ ID N0:326)
reverse PCR primer 5'-ACTGGACCAATTCTTCTGTG-3' (SEQ ID N0:32?)
lybridization~robe
S'-GATATTCTAGCATATTGTCAGAAGGAAGGATGGTGCAAATTAGCT-3' (SEQ ID N0:328)
A fitll length clone was identified that contained a single open reading frame
with an apparent translational
initiation site at nucleotide positions 14-16 and ending at the stop codon
found at nucleotide positions 446-448 (Figure
128; SEQ 1D N0:321). The predicted polypeptide precursor is 144 amino acids
long, has a calculated molecular
weight of approximately 16,699 dahons and an estimated pI of approximately
5.6. Analysis of the full-length PR0181
sequence shown in Figure 129 (SEQ 1D N0:322) evidences the presence of the
following: a signal peptide from about
amino acid 1 to about amino acid 20, a putative type II transmembrane domain
from about amino acid 11 to about
amino acid 31 and other transmembrane domains from about amino acid 57 to
about amino acid 77 and from about
amino acid 123 to about amino acid 143. Clone UNQ155 (DNA23330-1390) has been
deposited with ATCC on April
14, 1998 and is assigned ATCC deposit no: 209775.
Analysis of the amino acid sequence of the full-length PR0181 polypeptide
suggests that it possesses
significant sequence similarity to the corttichon protein, thereby indicating
that PR0181 may be a novel cornichon
homolog. More specifically, an analysis of the Dayhoff database (version 35.45
SwissProt 35) evidenced significant
homology between the PR0181 amino acid sequence and the following Dayhoff
sequences, AF022811,1,
CET09E8,3, S64058, YGF4~YEAST, YB60_YEAST, EBU89455_1, SIU36383~3 and PH1371.
EXAN1~LE 51: js~~ion of cDNA Clones EncodinE Human PR0195
A cDNA sequence was isolated in the amylase screen described in Example 2
above and is herein
designated DNA13199 (Figure 134; SEQ ID N0:332). The DNA13199 sequence was
then compared to a variety
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of expressed sequence tag (EST) databases which included public EST databases
(e.g., GenBank) to identify existing
homologies. The homology search was performed using the computer program BLAST
or BLAST2 (Altshul et al.,
l~ethc,~ds jp~mologyr 2t~6:4b0-080 (1996)). Those comparisons resulting in a
BLAST score of 70 (or in some
cases 90) or greater that did not encode knOWn proteins were clustered and
assembled into consensus DNA sequences
with the program "phrap" (Phil Green, University of 'Washington, Seattle,
Washington;
hap:l/bozeman.mbt.washington.edulphrap.docs/phrap.html). The consensus
sequence obtained therefrom is herein
designated as DNA22778.
Based on the DNA22778 sequence; oligonucleotide probes were generated and used
to screen a human
placenta library (i.IB89) prepared as described in paragraph I of Example 2
above. The cloning vector was pRRSB
(pRKSB is a precursor of pRI~SD that does not contain the SfiI site; see,
Holmes et al., 'enc , 2_x:1278-1280
IO (1991)), and the cDNA size cut was less than 2800 bp.
PCR primers (forward and reverse) were synthesized:
forward PCR Timer 5'-ACAAGCTGAGCTGCTGTGACAG-3' (SEQ ID NO:333)
reverse PCR primer 5'-TGATTCTGGCAACCAAGATGGC-3' (SEQ ID N0:334)
Additionally, a synthetic oligonucleatide hybridization probe was constructed
from the DNA22778 sequence which
had the following nucleotide sequence
hybridization robe
5'-ATGGCCTTGGCCGGAGGTTCGGGGACCGCTTCGGCTGAAG-3' (SEQ ID N0:335)
1n order to screen several libraries for a source of a full-length clone, DNA
from the libraries was screened
by PCR amplification with the PCR primer pair identified above. A positive
library was then used to isolate clones
encoding the PR0195 gertc using the probe oligonucleotide and one of the PCR
primers.
A full length clone was identified that contained a single open reading frame
with an apparent translational
initiation site at nucleotide positions 70-72 and ending at the stop codon
found at nucleotide positions 1039-1041
(Figure 132; SEQ 1D N0:330). The predicted polypeptide precursor is 323 amino
acids long, has a calculated
molecular weight of approximately 36,223 daltons and an estimated pI of
approximately 5.06. Analysis of the full-
length PROI95 sequence shown in Figure 132 (SEQ ID N0:330) evidences the
presence of the following: a signal
peptide from about amino acid I to about amino acid 3I, a transmembrane domain
from about amino acid 241 to
about amino acid 260 and a potential N-glycosylation site from about amino
acid 90 to about amino acid 93. Clone
UNQ169 (DNA26847-1395) has been deposited with ATCC on April 14, 1998 and is
assigned ATCC deposit no.
209772.
Analysis of the amino acid sequence of the full-length PR0195 polypeptide
suggests that it possesses no
significant sequence similarity to any known protein. However, an analysis of
the Dayhoff database (version 35.45
SwissProt 35) evidenced some degree of homology between the PR019~5 amino acid
sequence and the following
Dayhoff sequences, P P91380, AF035118_l, HUMTROPCS 1, NUOD_SALTY and E70002.
EXAMPLE 52: Isolation of cDN~A Clones Encoding Human PR0865
A cDNA sequence isolated in the amylase screen described in Example 2 above
was herein designated
DNA37642 (Figure 137, SEQ I?? N0:338). The DNA37642 sequence was then compared
to a variety of expressed
sequence tag (EST) databases which included public EST databases (e.g.;
GenBank) and a proprietary EST DNA
database (LIFESEQ"', htcyte Pharmaceuticals, Palo Alto, CA) to identify
homologies therebetween. The homology
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search was performed using the computer program BLAST or BLAST2 (Altshul et
al., Methods~n Enzymology
266:46080 (I996)). Those comparisons resulting in a BLAST score of 70 (or in
some cases 90) or greater that did
not encode Irnown proteins were clustered and assembled into consensus DNA
sequences with the program "phrap"
( P h t 1 Green, University of Washington, Seattle, Washington;
http:/Ibozeman.mbt.Washington.edulphrap.docslphrap.html). The consensus
sequence obtained is herein designated
DNA48615.
Based on the DNA486I5 consensus sequence, probes were generated and used to
screen a human fetal
kidney {LIB227) library prepared as described in paragraph 1 of Example 2
above. The cloning vector was pRKSB
(pRKSB is a precursor of pRKSD that does not contain the SfiI site; see,
Holmes et al., S fence, 53:1278-1280
(199i)), and the cDNA size cut was less than 2800 bp.
10. PCR primers (forward and reverse) were synthesized:
forward PCR primer 1 5'-AAGCTGCCGGAGCTGCAATG-3' (SEQ ID N0:339) '
forward PCR primer 2 5'-TTGCTTCTTAATCCTGAGCGC-3' (SEQ ID N0:340)
forward PCR primer 3 5'-AAAGGAGGACTTTCGACTGC-3' (SEQ ID N0:341)
rgverse PCR primer 1 5'-AGAGATTCATCCACTGCTCCAAGTCG-3' (SEQ ID N0:342)
reverse PCR primer 2 5'-TGTCCAGAAACAGGCACATATCAGC-3' (SEQ ID N0:343)
Additionally, a synthetic oligonucleotide hybridization probe was constructed
from the consensus DNA48615
sequence which had the following nucleotide sequence
hybridization~robe
5'-AGACAGCGGCACAGAGGTGCTTCTGCCAGGTTAGTGGTTACTTGGATGAT-3' (SEQ ID N0:344)
In order to screen several libraries for a source of a full-length clone, DNA
from the libraries was screened
by PCR amplification with the PCR primer pairs identified above. A positive
library was then used to isolate clones
encoding the PR0865 gene using the probe oligonucleotide and one of the PCR
primers.
A full length clone was identified that contained a single open reading frame
with an apparent translational
initiation site at nucleotide positions 173-175 and ending at the stop colon
found at nucleotide positions 1577-1579
(Figure 135; SEQ ID N0:336). The predicted polypeptide precursor is 468 amino
acids long, has a calculated
molecular weight of approximately 54,393 daltons and an estimated pI of
approximately 5.63. Analysis of the full-
Iength PR0865 sequence shown in Figure 136 (SEQ ID N0:337) evidences the
presence of the following: a signal
peptide from about amino acid 1 to about amino acid 23, potential N-
glycosylation sites from about amino acid 280
to about amir~ acid 283 and from about amino acid 384 to about amino acid 387,
a potential amidation site from
about amino acid 94 to about amino acid 97, glycosaminoglycan attachment sites
from about amino acid 20 to about
amino acid 23 and from about amino acid 223 to about amino acid 226, an
aminotransferase class-V pyridoxyl-
phosphate amino acid sequence block from about amino acid 216 to about amino
acid 222 and an amino acid sequence
block similar to that found in the interleukin-7 protein from about amino acid
338 to about amino acid 343. Clone
UNQ434 (DNA539?4-1401) has been deposited with ATCC on April 14, 1998 and is
assigned ATCC deposit no.
209774.
Analysis of the amino acid sequence of the full-length PR0865 polypeptide
suggests that it possesses no
significant sequence similarity to any known protein. However, an analysis of
the Dayhoff database (version 35.45
SwissProt 35) evidenced some degree of homology between the PR0865 amino acid
sequence and the following
Dayhoff sequences, YMNO YEAST, ATFCA4 43, S44168, P W14549 and RABTCRG4 1.
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EXAMPLI~ 53: Isolation o~cDNA Clones Encodin~Human PR0827
A cDNA sequence isolated in the amylase screen described in Example 2 above
was found, by BLAST and.
FastA sequence alignment, to have sequence homology to nucleotide sequences
encoding various integrin proteins.
This cDNA sequence is herein designated DNA47751 (see Figure 140; SEQ ID
N0:347). Based on the sequence
homology, probes were generated from the sequence of the DNA47751 molecule and
used to screen a human fetal
pigment epithelium library (LIB113) prepared as described in paragraph 1 of
Example 2 above. The cloning vector
was pRKSB (pRKSB is a precursor of pRKSD that does not contain the Sfil site;
see, Holmes et al., Science,
25:1278-1280 {I991)), and the cDNA size cut was less than 2800 bp.
PCR primers (forward and reverse) were synthesized:
forward PCR rimer 5'-AGGGACAGAGGCCAGAGGACTTC-3' (SEQ ID N0:348)
reyerse PCR primp 5'-CAGGTGCATATTCACAGCAGGATG-3' {SEQ ID N0:349)
Additionally, a synthetic oligonucleotide hybridization probe was constructed
from the consensus DNA47751
sequence which had the following nucleotide sequence
hvbrid_~zation probe
5'-GGAACTCCCCTTCGTCACTCACCTGTTCTTGCCCCTGGTGTTCI:T-3' (SEQ ID
N0:350)
In order to screen several libraries for a source of a full-length clone, DNA
from the Iibraties was screened
by PCR amplification with the PCR primer pair identified above. A positive
library was then used to isolate clones
encoding the PR0827 gene using the probe oligonucleotide and one of the PCR
primers.
A full length clone was identified that contained a single open reading frame
with an apparent translational
initiation site at nucleotide positions 134-136 and ending at the stop codon
found at nucleotide positions 506-508
(Figure 138; SEQ ID N0:345). The predicted polypeptide precursor is 124 amino
acids long, has a calculated
molecular weight of approximately 13,352 daltons and an estimated pI of
approximately 5.99. Analysis of the full-
length PR0827 sequence shown in Figure 139 (SEQ ID N0:346) evidences the
presence of the following: a signal
peptide from about amino acid 1 to about amino acid 22, a cell attachment
sequence from about amino acid 70 to
about amino acid 72, a potential N-glycosylation site 'from about amino acid
98 to about amino acid 101 and an
integrin alpha chain protein homology sequence from about amino acid 67 to
about amino acid 81. Clone UNQ468
(DNA57039-1402) has been deposited with ATCC on April 14, 1998 and is assigned
ATCC deposit no. 209777.
Analysis of the amino acid sequence of the full-length PR0827 polypeptide
suggests that it possesses
significant sequence similarity to the VLA-2 integrin protein and various
other integrin proteins, thereby indicating
that PR0827 may be a novel integrin or splice variant thereof. More
specifically, an analysis of the Dayhoff database
(version 35.45 SwissProt 35) evidenced significant homology between the PR0240
amino acid sequence and the
following Dayhoff sequences, S44142, ITA2 HUMAN, ITAl RAT, ITAl HUMAN, ITA4
HUMAN,
fTA9 HUMAN, AF032108 l, iTAM MOUSE, ITA8 CRICK and ITA6 CHICK.
EXAMPLE 54: Isolation of cDNt~ Cjones Encoding_Human P,~tOI I 14
A cDNA sequence isolated in the amylase screen described in Example 2 was
found, by the WU-BLAST2
sequence alignment computer program, to have certain sequence identity to
other lo~own interferon receptors. This
cDNA sequence is herein designated DNA48466 (Figure 143; SEQ ID N'0:352).
Based on the sequence identity,
probes were generated from the sequence of the DNA48466 molecule anti used to
screen a human breast carconoma
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library (LIBI35) prepared as described in paragraph 1 of Example 2 above. The
cloning vector was pRKSB (pRKSB
is a precursor of pRKSD that does not contain the Sfil site; see, Holmes et
al., Science, 253:1278-1280 (1991)), and
the cDNA size cut was less than 2800 bp.
The oligonucleotide probes employed were as follows:
forward PCR primer S'-AGGCTTCGCTGCGACTAGACCTC-3' (SEQ ID N0:354)
reverse PCR primer S'-CCAGGTCGGGTAAGGATGGTTGAG-3' (SEQ ID N0:355)
)~bridization probe
S'-TTTCTACGCATTGATTCCATGTTTGCTCACAGATGAAGTGGCCATTCTGC-3' (SEQ ID N0:356)
A full length clone was identified that contained a single open reading frame
with an apparent translational
initiation site at nucleotide positions 250-252, and a stop signal at
nucleotide positions 1183-1185 (Figure 141, SEQ
ID N0:351). The predicted polypeptide precursor is 311 amino acids long, has a
calculated molecular weight of
approximately 35,076 daltons and an estimated p1 of approximately 5.04.
Analysis of the full-length PR01114
interferon receptor sequence shown in Figure 142 (SEQ 1D N0:352) evidences the
presence of the following: a signal
peptide from about amino acid 1 to about amino acid 29, a transmembrane domain
from about amino acid 230 to
about amino acid 255, potential N-glycosylation sites from about amino acid 40
to about amino acid 43 and from
about amino acid 134 to about amino acid 137, an amino acid sequence block
having homology to tissue factor
proteins from about amino acid 92 to about amino acid 119 and an amino acid
sequence block having homology to
integrin alpha chain proteins from about amino acid 232 to about amino acid
262. Clone UNQ557 (DNA57033-1403)
has been deposited 'with ATCC on May 27, 1998 and is assigned ATCC deposit no.
209905.
An analysis of the Dayhoff database (version 35.45 SwissProt 35), using a WU-
BLAST2 sequence alignment
analysis of the full-length sequence shown in Figure 142 (SEQ ID N0:352),
evidenced significant homology between
the PR01114 interferon receptor amino acid sequence and the following Dayhoff
sequences: 601418,
INRl MOUSE, P 871035, INGS HUMAN, A26595_i, A26593_l, I56215 and TF HUMAN.
EXAMPLE 55: TsolatiQn of cDNA Clones Encoding Human PR0237
A consensus sequence was obtained relative to a variety of EST sequences as
described in Example 1 above,
wherein the consensus sequence obtained is herein designated DNA30905. Based
on the DNA30905 consensus
sequence, oligonucieotides were synthesized: I) to identify by PCR a cDNA
library that contained the sequence of
interest, and 2) for use as probes to isolate a clone of the full-length
coding sequence for PR0237.
PCR primers (forward and reverse) were synthesized:
forward PCR primer 5'-TCTGCTGAGGTGCAGCTCATTCAC-3' (SEQ ID N0:359)
reverse PCR vrimer 5'-GAGGCTCTGGAAGATCTGAGATGG-3' (SEQ ID N0:360)
Additionally, a synthetic oligonucleotide hybridization probe was constructed
from the consensus DNA30905
sequence which had the following rrucleotide sequence
hybridization Bxobe
5'-GCCTCTTTGTCAACGTTGCCAGTACCTCTAACCCATTCCTCAGTCGCCTC-3' (SEQ ID N0:361)
In order to screen several libraries for a source of a full-length clone, DNA
from the libraries was screened
by PCR amplification with the PCR primer pair identified above. A positive
library was then used to isolate clones
encoding the PR0237 gene using the probe oligonucleotide and one of the PCR
primers. RNA for construction of
the cDNA libraries was isolated from human fetal brain tissue {LIB153).
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DNA sequencing of the clones isolated as described above gave the full-length
DNA sequence for PR0237
[herein designated as UNQ211 (DNA34353-1428)] (SEQ ID N0:357) and the derived
protein sequence for PR0237.
The entire nucleotide sequence of UNQ211 (DNA34353-1428) is shown in Figure
144 (SEQ ID N0:357).
Clone UNQ211 (DNA34353-1428) contains a single open reading frame with an
apparent translational initiation site
at nucleotide positions 586-588 and ending at the stop codon at nucleotide
positions 1570-1572 (Figure 144). The
predicted polypeptide precursor is 328 amino acids long (Figure 145): The full-
length PR0237 protein shown in
Figure 145 has an estimated molecular weight of about 36,238 daltons and a pI
of about 9.90. Analysis of the full-
length PR0237 sequence shown in Figure 145 (SEQ ID N0:358) evidences the
presence of the following: a signal
peptide from about amino acid 1 to about amino acid 23, a transmembrane domain
from about amino acid 177 to
about amino acid 199, potential N-glycosylation sites from about amino acid
118 to about amino acid i21, from about
amino acid 170 to about amino acid 173 and from about amino acid 26(1 to about
amino acid 263 and eukaryotic-type
carbonic anhydrase sequence homology blocks from about amino acid 222 to about
amino acid 270, from about amino
acid 128 to about amino acid 164 and from about amino acid 45 to about amino
acid 92. Clone UNQZ11
(DNA34353-1428) has been deposited with ATCC on May 12, 1998 and is assigned
ATCC deposit no. 209855.
Analysis of the amino acid sequence of the full-length PR0237 polypeptide
suggests that it possesses
significant sequence similarity to the carbonic anhydrase protein. More
specifically, an analysis of the Dayhoff
database (version 35.45 SwissProt 35) evidenced significant homology between
the PR0237 amino acid sequence and
the following Dayhoff sequences, AF050106_l, OACALP_1, CELD1022 8, CAH2~HUMAN,
1CAC,
CAHS~HUMAN, CARP HUMAN, CAH3 HUMAN, CAH1 HUMAN and 2CAB.
~XA~~uIPLE 56: ~solatiort of cDNA Clones Encoding Human PR0541
A consensus sequence was obtained relative to a variety of EST sequences as
described in Example 1 above,
wherein the consensus sequence obtained is herein designated DNA42259. Based
on the DNA42259 consensus
sequence, oligonucleotides were synthesized: 1) to identify by PCR a cDNA
library that contained the sequence of
interest, and 2) for use as probes to isolate a clone of the full-length
coding sequence for PR0541.
PCR primers (forward and reverse) were synthesized:
forward PCR primer 5'-GGACAGAATTTGGGAGCACACTGG-3' (SEQ ID N0:364)
forward PAR primer 5'-CCAAGAGTATACTGTCCTCG-3' (SEQ ID N0:365)
reverse PCR primer 5'-AGCACAGATTTTCTCTACAGCCCCC-3' (SEQ ID NO:366)
reverse PCR primer 5'-AACCACTCCAGCATGTACTGCTGC-3' (SEQ ID N0:367)
Additionally, a synthetic oligonucieotide hybridization probe was constructed
from the consensus DNA42259
sequence which had the following nucleotide sequence
hvbridizattq~n probe
5'-CCATTCAGGTGTTCTGGCCCTGTATGTACACATTATACACAGGTCGTGTG-3' (SEQ ID N0:368)
In order to screen several libraries for a sowce of a full-length clone, DNA
from the libraries was screened
by PCR amplification with one of the PCR primer pairs identified above. A
positive library was then used to isolate
clones encoding the PR0541 gene using the probe oIigonucleotide and one of the
PCR primers. RNA for
construction of the cDNA libraries was isolated from human fetal kidney ~ussue
(LIB227).
DNA sequencing of the clones isolated as described about gave the full-length
DNA sequence for PR0541
(herein designated as UNQ342 (DNA45417-1432)] (SEQ ID N0:362) and the derived
protein sequence for PR0541.
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The entire nucleotide sequence of UNQ342 (DNA45417-1432) is shown in Figure
146 (SEQ ID N0:362).
Clone UNQ342 (DNA45417-1432) contains a single open reading frame with an
apparent translationaI initiation site
at nucleotide positions 469-47I and ending at the stop codon at nucleotide
positions 1969-1971 (Figure 146). The
predicted polypeptide precursor is 500 amino acids long (Figure 147), The full-
length PR0541 protein shown in
Figure 147 has an estimated molecular weight of about 56,888 daltons and a p1
of about 8.53. Analysis of the full-
length PR0541 sequence shown in Figure 147 (SEQ ID N0:363) evidences the
presence of the following: a signal
peptide from about amino acid 1 to about amino acid 20, amino acid sequence
blocks having homology to
extracelIular proteins SCPlTpx-1/AgSIPR-l/Sc7 from about amino acid 165 to
about amino acid I86, from about
amino acid 196 to about amino acid 218, from about amino acid 134 to about
amino acid 146, from about amino acid
96 to about amitw acid 108 and from about amino acid 58 to about amino acid 77
and a potential N-glycosylation site
from about amino acid 28 to about amino acid 31. Clone UNQ342 (DNA45417-1432)
has been deposited with ATCC
on May 27, 1998 and is assigned ATCC deposit no. 209910.
Analysis of the amino acid sequence of the full-length PR0541 polypeptide
suggests that it possesses
significant sequence similarity to a trypsin inhibitor protein, thereby
indicating that PR054i may be a novel irypsin
inhibitor. More specifically, an analysis of the Dayhoff database (version
35.45 SwissProt 35) evidenced significant
homology between the PR0541 amino acid sequence and the following Dayhoff
sequences, D45027-1, AB009609 1,
JC5308, CRS3 HORSE, TPXl HUMAN, HELD HELHO, GEN14351, A28112_1, CETOSA10 4 and
P W11485.
EXAMPLE 57: IsolatiQn~f cDNA Clones Encoding Human~R0273
A consensus sequence was obtained relative to a variety of EST sequences as
described in Example 1 above,
wherein the consensus sequence obtained is herein designated DNA36465. Based
on the DNA36465 consensus
sequence, oligonucleotides were synthesized: 1) to identify by PCR a cDNA
library that contained the sequence of
interest, and 2) for use as probes to isolate a cione of the full-length
coding sequence for PR0273.
A pair of PCR primers (forward and reverse) were synthesized:
forward PCR~ritr~e_r 5'-CAGCGCCCTCCCCATGTCCCTG-3' (SEQ ID N0:371)
reverse PCR primer 5'-TCCCAACTGGTTTGGAGTTTTCCC-3' (SEQ ID N0:372)
Additionally, a synthetic oligotntcleotide hybridization probe was constructed
from the consensus DNA36465
sequence which had the following nucleotide sequence
hvbridiz~t,, ion.prob~
5'-CTCCGGTCAGCATGAGGCTCCTGGCGGCCGCTGCTCCTGCTGCTG-3' (SEQ ID NO:373)
In order to screen several libraries for a source of a full-length clone, DNA
from the libraries was screened
by PCR amplification with the PCR primer pair identified above. A positive
library was than used to isolate clones
encoding the PR0273 gene using the probe oligonucleotide and one of the PCR
primers. RNA for construction of
the cDNA libraries was isolated from human fetal kidney tissue.
DNA sequencing of the clones isolated as described above gave the full-length
DNA sequence for PR0273
jherein designated as UNQ240 (DNA39523-1192)] (SEQ ID N0:369) and the derived
protein sequence for PR0273.
The entire nucleotide sequence of UNQ240 {DNA39523-1192) is shown in Figure
148 (SEQ ID N0:369).
Clone UNQ240 (DNA39523-1192) contains a single open reading frame with an
apparent translational initiation site
at nucleotide positions 167-169 and ending at the stop codon at nucleotide
positions 500-502 {Figure 148). The
predicted polypeptide precursor is 111 amino acids long {Figure 149). Clone
UNQ240 (DNA39523-1192) has been
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deposited with the ATCC. It is understood that the deposited clone contains
the actual sequence and that the
sequences provided herein are merely representative based on current
sequencing techniques. Moreover, given the
sequences provided herein and knowledge of the universal genetic code, the
corresponding nucleotides for any given
amino acid can be routinely identified and vice versa.
Analysis of the amino acid sequence of the full-length PR0273 polypeptide
suggests that portions of it
possess sequence identity with human macrophage inflammatory protein-2,
cytokine-induced neutrophil
chemoamactant 2, and neutrophiI chemotactic factor 2-beta, thereby indicating
that PR0273 is a novel chemokine.
As discussed further below, the cDNA was subcloned into a baculovirus vector
and expressed in insect cells
as a C-terminally tagged IgG fusion protein. N-terminal sequencing of the
resultant protein identified the signal
sequence cleavage site, yielding a mature poIypeptide of 77 amino acids. The
mature sequence, showing 3I-40%
identity to other human CXC chemokines, includes the four canonical cysteine
residues but lacks the ELR motif.
Northern analysis demonstrates expression at least in the small intestine,
colon, spleen, lymph node and kidney. By
in situ hybridization, also described in detail below, mRNA is localized to
the lamina propria of intestinal villi and
to renal tubules.
EXAMPLE 58: Isolation of cDNA Clones Encoding Human PR070I
A consensus sequence was obtairred relative to a variety of EST sequences as
described in Example l above,
wherein the consensus sequence obtained is herein designated DNA39848. Based
on the DNA39848 consensus
sequence, oligonucleotides were synthesized: I) to identify by PCR a cDNA
library that contained the sequence of
interest; and 2) for use as probes to isolate a clone of the full-length
coding sequence for PR0701.
A pair of PCR primers (forward and reverse) were synthesized:
forward PCR rimer 5'-GGCAAGCTACGGAAACGTCATCGTG-3' (SEQ ID N0:376)
reverse PCR rimer 5'-AACCCCCGAGCCAAAAGATGGTCAC-3' (SEQ ID N0:377)
Additionally, a synthetic oligonucleotide hybridization probe was constructed
from the consensus DNA39848
sequence which had the following nucleotide sequence:
hybridization probe
5'-GTACCGGTGACCAGGCAGCAAAAGGCAACTATGGGCTCCTGGATCAG-3' (SEQ ID N0:378).
In order to screen several libraries for a source of a full-length clone, DNA
from the libraries was screened
by PCR amplification with the PCR primer pair identified above. A positive
library was then used to isolate clones
encoding the PR0701 gene using the probe oligonucleotide and one of the PCR
primers. RNA for construction of
the cDNA libraries was isolated from human fetal kidney tissue (LIB227).
DNA sequencing of the cioties isolated as described above gave the full-length
DNA sequence for PR0701
[herein designated as UNQ365 (DNA44205-1285)j (SEQ ID N0:374) and the derived
protein sequence for PR0701.
The entire nucleotide sequence of UNQ365 (DNA44205-1285) is shown in Figure
150 (SEQ ID N0:374).
Clog UNQ365 (DNA44205-1285) contains a single open reading frame with an
apparent transiational initiation site
at nucleotide positions 50-52 and ending at the stop codon at nucleotide
positions 2498-3000 (Figure 150). The
predicted polypeptide precursor is 816 amino acids long (Figure 151). The full-
length PR0701 protein shown in
Figure 151 has an estimated molecular weight of almut 91,794 daltons, a p1 of
about 5.88 and NX(S!T) being 4.
Clone UNQ365 (DNA44205-1285) has been deposited with the ATCC on March 31,
1998. It is understood that the
clone was the correct and actual sequence, wherein the sequences provided
herein are representative based on
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sequencing techniques.
Still regarding the amino acid sequence shown in Figure 151, there is a
potential signal peptide cleavage site
at about amino acid 25. There are potential N-glycosylation sites at about
amino acid positions 83, 511, 716 and 803.
The carboxyIesterases type-B signature 2 sequence is at about residues 125 to
135. Regions homologous with
carboxylesterase type-B are also at about residues 54-74, 197-212 and 221-261.
A potential transmembrane region
corresponds approximately to amino acids 671 through about 700. The
corresponding nucleic acids can be routinely
determined from the sequences provided herein.
Analysis of the amino acid sequence of the full-Length PR0701 polypeptide
suggests that it possess
significant homology to the neuroligins from rattus norvegicus indicating that
PR0701 may be a novel human
neuioligin.
EXAMPLE 59: Isolation of cDNA Clones Encoding Human PR0704
A consensus sequence was obtained relative to a variety of EST sequences as
described in Example 1 above,
wherein the consensus sequence obtained is herein designated DNA43033. Based
on the DNA43033 consensus
sequence, oligonucleotides were synthesized: 1) to identify by PCR a cDNA
library that contained the sequence of
interest, and 2) for use as probes to isolate a clone of the full-length
coding sequence for PR0704.
A pair of PCR primers (forward and reverse) were synthesized:
forward PCR Q_rimer 5'-CCTTGGGTCGTGGCAGCAGTGG-3' (SEQ ID N0:381);
reverse PCR primer 5'-CACTCTCCAGGCTGCATGCTCAGG-3' (SEQ ID N0:382).
Additionally, a synthetic oligonucleotide hybridization probe was constructed
from the DNA43033 consensus
sequence which had the following nucleotide sequence:
hybridization lobe
5'-GTCAAACGTTCGAGTACTTGAAACGGGAGCACTCGCTGTCGAAGC-3' (SEQ ID N0:383).
In order to screen several libraries for a source of a full-length clone, DNA
from the libraries was screened
by PCR art>plification with the PCR primer pair identified above. A positive
library was then used to isolate clones
encoding the PR0704 gene using the probe oligonucleotide and one of the PCR
primers. RNA for construction of
the cDNA libraries was isolated from human fetal kidney tissue (LIB227).
DNA sequencing of the clones isolated as described above gave the full-length
DNA sequence for PR0704
[herein designated as UNQ368 (DNA50911-1288)] (SEQ ID N0:3?9) and the derived
protein sequence for PR0704.
The entire ttttcleotide sequence of UNQ368 (DNA50911-1288) is shown in Figure
152 (SEQ ID N0:379).
Clone UNQ368 (DNA50911-1288) contains a single open reading frame with an
apparent translatianal initiation site
at nucleotide positions 8-10 and ending at the stop codon at nucleotide
positions 1052-1054 (Figure 152). The
predicted polypeptide precursor is 348 amino acids long (Figure 153). The full-
length PR0704 protein shown in
Figure 153 has an estimated molecular weight of about 39,711 and a pI of about
8.7. Clone UNQ368 (DNA50911-
1288) has been deposited with the ATCC on March 31, 1998. Regarding the
sequence, it is understood that the
deposited clone contains the correct sequence, and the sequences provided
herein are based on known sequencing
techniques.
Analysis of the amino acid sequence of the full-length PR0704 polypeptide
suggests that portions of it
possess significant homology to the vesicular integral membrane protein 36,
thereby indicating that PR0704 tttay be
a novel vesicular integral membrane protein.
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Still analyzing the amino acid sequence of SEQ ID N0:380, the putative signal
peptide is at about amino
acids 1-39 of SEQ ID N0:380. The transmembrane domain is at amino acids 310-
335 of SEQ ID N0:380. A
potential N-glycosylation site is at about amino acids 180-183 of SEQ ID
N0:380. The corresponding nucleotides
can be routinely determined given the sequences provided herein.
EXAMPLE 60: Iso-,lalion o~eDNA Clones Encgding Human PR07Q6
A consensus sequence was obtained relative to a variety of EST sequences as
described in Example 1 above,
wherein the consensus sequence obtained is herein designated DNA40669. Based
on the DNA40669 consensus
sequence, oligonucleotides were synthesized: 1) to identify by PCR a cDNA
library that contained the sequence of
interest, and 2) for use as probes to isolate a clone of the full-length
coding sequence for PR0706.
A pair of PCR primers (forward and reverse) were synthesized:
forward PCR 'met 5'-CCAAGCAGCTTAGAGCTCCAGACC-3' (SEQ ID N0:386)
reverse PCR primer 5'-TTCCCTATGCTCTGTATTGGCATGG-3' (SEQ ID N0:387)
Additionally, a synthetic oligotntcleotide hybridization probe was constructed
from the consensus DNA40669
sequence which had the following nucleotide sequence
h bridi anon probe
5'-GCCACTTCTGCCACAATGTCAGCTTTCCCTGTACCAGAAATGGCTGTGTT-3' (SEQ ID N0:388)
In order to screen several libraries for a source of a full-length clone, DNA
from the -libraries was screened
by PCR amplification with the PCR primer pair identified above. A positive
library was then used to isolate clones
encoding the PR0706 gene using the probe oligonucleotide and one of the PCR
primers. RNA for construction of
the cDNA libraries was isolated from human fetal brain tissue (LIB153).
DNA sequencing of the clones isolated as described above gave the full-length
DNA sequence for PR0706
[herein designated as UNQ370 (DNA48329-1290)] (SEQ ID N0:384) and the derived
protein sequence for PR0706.
It is understood that the deposited clone contains the actual sequence, and
that the sequences provided herein are
representative based on current sequencing techniques.
The entire nucleotide sequence of UNQ370 (DNA48329-1290) is shown in Figure
154 (SEQ ID N0:384).
Clone UNQ370 (DNA48329-1290) contains a single open reading frame with an
apparent translational initiation site
at nucleotide positions 279-281 and ending at the stop codon at nucleotide
positions 1719-1721 (Figure 154). The
predicted polypeptide precursor is 480 amino acids long (Figure 155). Tlte
full-length PR0706 protein shown in
Figure 155 has an estimated molecular weight of about 55,239 daltons and a pI
of about 9.30. Clotic UNQ370
(DNA48329-1290) has been deposited with the ATCC on April 21, 1998.
Still regarding the amino acid sequence shown in Figure 155, there is a
potential signal peptide cleavage site
at about amino acid 19. There are potential N-glycosylation sites at about
amino acid positions 305 and 354. There
is a potential tyrosine ldnase phosphorylation site at about amino acid
position 333. A region homologous with
histidine acid phosphatase is at about residues 87-102. The corresponding
nucleic acid regions can be routinely
determined given the provided sequences, i.e., the eadons can be determined
from the specifically named amino acids
given.
Analysis of the amino acid sequence of the full-length PR0706 polypeptide
suggests that portions of it
possess significam homology to the human prostatic acid phosphatase precursor
thereby indicating that PR0706 may
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be a novel human prostatic acid phosphatase.
EXAMPLE 6i: Isolation of cDNrA Clones )incoding Human PR0707
A consensus sequence was obtained relative to a variety of EST sequences as
described in Example 1 above,
wherein the consensus sequence obtained is herein designated DNA42775. Based
on DNA42775, olieonucleotides
were synthesized: l) to identify by PCR a cDNA library that contained the
sequence of interest, and 2) for use as
probes to isolate a clone of the full-length coding sequence for PR0707.
A pair of PCR primers (forward and reverse) were synthesized:
forward PCR p~ir~er 5'-TCCGTCTCTGTGAACCGCCCCAC-3' (SEQ ID N0:391);
reverse PCR primer 5'-CTCGGGCGCATTGTCGTTCTGGTC-3' (SEQ ID N0:392}.
Additionally, a synthetic oligonucleotide hybridization probe was constructed
from the DNA42775 sequence which
had the following nucleotide sequence:
hybridization probe
5'-CCGACTGTGAAAGAGAACGCCCCAGATCCACTTATTCCCC-3' (SEQ ID N0:393).
In order to screen several libraries for a source of a foil-length clone, DNA
from the libraries was screened
by PCR amplification with the PCR primer pair identified above. A positive
library was then used to isolate clones
encoding the PR0707 gene using the probe oligonucleotide and one of the PCR
primers. RNA for construction of
the cDNA libraries was isolated from human fetal kidney tissue {LIB22?).
DNA sequencing of the clones isolated as described above gave the full-length
DNA sequence for PR0707
[herein designated as UNQ371 (DNA4830tr1291)] (SEQ ID N0:389) and the derived
protein sequence for PR0707.
The entire nucleotide sequence of UNQ371 (DNA48306-1291) is shown in Figure
156 (SEQ ID N0:389).
Clone UNQ371 (DNA48306-I291) contains a single open reading frame with an
apparent translational initiation site
at nucleotide positions 371-373 and ending at the stop codon at nucleotide
positions 3119-3121 of SEQ ID N0:389.
The predicted polypeptide precursor is 91b amino acids long (Figure 157). The
full-length PR0707 protein shown
in Figure 157 has an estimated molecular weight of about 100,204 daltons and a
pI of about 4.92. Clone UNQ371
(DNA48306-1291) has been deposited with ATCC on May 27, 1998. It is understood
that the clone UNQ371 which
is deposited is that which encodes PR0707, and that the sequences herein are
merely representations based on known
sequencing techniques which may be subject to minor errors.
Regarding analysis of the amino acid sequence, the signal sequence appears to
be at about 1 through 30 of
SEQ )D N0:390. Cadhetins extracellular repeated domain signature sequence is
at about amino acids 121-131, 230-
240, 333-345, 4401150, and 550-X60 of SEQ ID N0:390. Tyrosine kinase
phosphorylation sites are at about amino
acids 124-132 and 580-5$6 of SEQ ID NO:390. A potential transmembrane domain
is at about amino acids 682-715
t 5. The nucleic acid positions can be derived by referring to the
corresponding codon for the named amino acid.
Analysis of the amino acid sequence of the full-length PR0707 polypeptide
suggests that portions of it
possess significant homology to the cadherin FIBS protein, expressed in human
fibroblasts, thereby indicating that
PR0707 may be a novel cadherin.
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EXAMPLE 62: Isolation of eDNA Clones Encoding Human PR0322
A consensus sequence was obtained relative to a variety of EST sequences as
described in Example 1 above,
wherein the consensus sequence obtained is herein designated DNA48336. Based
on the DNA48336 consensus
sequence, oligonucleotides were synthesized: I) to identify by PCR a cDNA
library that contained the sequence of
interest, and 2) for use as probes to isolate a clone of the full-length
coding sequence for PR0322.
A pair of PCR primers (forward and reverse) were synthesized:
forward PCR primer 5'-CAGCCTACAGAATAAAGATGGCCC-3' (SEQ ID N0:396)
reverse PCR primer 5'-GGTGCAATGATCTGCCAGGCTGAT-3' (SEQ. ID N0:397)
Additionally, a synthetic oligonucleotide hybridization probe was constructed
from the DNA48336 consensus
sequence which had the following nucleotide sequence:
~bridization probe
5'-AGAAATACCTGTGGTTCAGTCCATCCCAAACCCCTGCTACAACAGCAG-3' (SEQ ?D N0:398).
In order to screen several libraries for a source of a full-length clone, DNA
from the libraries was screened
by PCR amplification with the PCR primer pair identified above. A positive
library was then used to isolate clones
encoding the PR0322 gene using the probe oligonucleotide and one of the PCR
primers. RNA for construction of
the cDNA libraries was isolated from human fetal kidney tissue (LIB227).
DNA sequencing of the clones isolated as described above gave the full-length
DNA sequence for PR0322
[herein designated as UNQ283 {DNA48336-1309)) (SEQ ID N0:394) and the derived
protein sequence for PR0322.
It is understood that UNQ283 (DNA48336-1309) in fact encodes PRO322, and that
SEQ ID N0:394 is a
representation of the sequence based on sequencing techniques known in the
art.
The entire nucleotide sequence of UNQ283 (DNA48336-1309) is shown in Figure
158 (SEQ ID N0:394).
Clone UNQ283 (DNA48336-1309) contains a single open reading frame with an
apparent translational initiation site
at nucleotide positions 166-168 and ending at the stop codon at nucleotide
positions 946-948 (Figure 158). The
predicted polypeptide precursor is 260 amino acids long (Figure 159). The full-
length PR0322 protein shown in
Figure 159 has an estimated molecular weight of about 28,028 daltons and a p1
of about 7.87. Clone UNQ283
(DNA48336-1309) has been deposited with ATCC and is assigned ATCC deposit no.
209669.
Regarding the amino acid sequence of Figure 159, a potential N-glycosylation
site is at amino acid 110 of
SEQ ID N0:395. The serine proteases, trypsin family and histidine active site
is identified at amino acids 69 through
74 of SEQ ID N0:395 and the consensus sequence is identified at amino acids
207 through 217 of SEQ ID N0:395.
The kringle domain proteins motif is identified at amino acids 205 tluoug;h
217 of SEQ ID N0:395. The putative
signal peptide is encoded at about amino acids 1-23.
Analysis of the amino acid sequence of the full-length PR0322 polypeptide
suggests that portions of it
possess significant homology to neuropsin and other serine proteases, thereby
indicating that PR0322 is a novel
serine protease related to'neuropsin.
EXAMPLE 63: Isolation of cDNA Clones Encoding Numan_PR0526
A consensus sequence was obtained relative to a variety of EST sequences as
described in Example 1 above,
wherein the consensus sequence obtained is herein designated DNA39626. Based
on the DNA39626 consensus
sequence, oligonucleotides were synthesized: I) to identify by PCR a cDNA
library that contained the sequence of
interest, and 2) for use as probes to isolate a clone of the full-length
coding sequence for PR0526.
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A pair of PCR primers (forward and reverse) were synthesized:
forward PCR ri~mer 5'-TGGCTGCCCTGCAGTACCTCTACC-3' (SEQ ID N0:401);
_rgverse,~~R rimer 5'-CCCTGCAGGTCATTGGCAGCTAGG-3' (SEQ 1D N0:402).
Additionally, a synthetic oligonucleotide hybridization probe was constructed
from the DNA39626 consensus
sequence which had the following nucleotide sequence:
~bridization probe
5'-AGGCACTGCCTGATGACACCTTCCGCGACCTGGGCAACCTCACAC-3' {SEQ ID N0:403).
In order to screen several libraries for a source of a full-length clone, DNA
from the libraries was screened
by PCR amplification with the PCR primer pair identified above. A positive
library was then used to isolate clones
encoding the PR0526 gene using the probe oligonucleotide and one of the PCR
primers. RNA for construction of
the cDNA libraries was isolated from human fetal liver tissue (LIB228).
DNA sequencing of the clones isolated as described above gave the full-length
DNA sequence for PR0526
[herein designated as UNQ330 (DNA44184-1319)] (SEQ ID N0:399) and the derived
protein sequence for PR0526.
The entire nucleotide sequence of UNQ330 (DNA44184-1319) is shown in Figure
160 (SEQ ID N0:399).
Clone UNQ330 (DNA44184-1319) contains a single open reading frame with an
apparent translational initiation site
at nucleotide positions 514-516 and ending at the stop codon at nucleotide
positions 1933-1935 (Figure 160). The
predicted polypeptide precursor is 473 amino acids long (Figure 161). The full-
length PR0526 protein shown in
Figure 161 has an estimated molecular weight of about 50,708 daltons and a p1
of about 9.28. Clone UNQ330
(DNA44184-1319) has been deposited with the ATCC on March 26, 1998. It is
understood that the clone contains
the actual sequence, whereas the sequences presented herein are representative
based on current sequencing
techniques.
Analysis of the amino acid sequence of the full-length PR0526 polypeptide
suggests that portions of it
possess significant homology to the ieucine repeat rich proteins including
ALS, SLIT, carboxypeptidase and platelet
glycoprotein V thereby indicating that PR0526 is a novel protein which is
involved in protein-protein interactions.
Still analyzing SEQ 1D N0:400, the signal peptide sequence is at about amino
acids 1-26. A leucine zipper
pattern is at about amino acids 135-156. A glycosaminoglycan attachment is at
about amino acids 436139. N-
glycosylation sites are at about amino acids 82-85, 179-182, 237-240 and 423-
426. A von Willebrand factor (VWF)
type C domains) is found at about amino acids 411-42$. The skilled artisan can
understand which nucleotides
correspond to these amino acids based on the sequences provided herein.
EXAMPLE 64: Isolation _o~' cDNA Clones EncodZng Human PR0531
An ECD database was searched and an expressed sequence tag (EST) from
LIFESEQT"', Incyte
Pharmaceuticals; Palo Alto, CA was identified which showed homology to
protocadherin 3. Based on this sequence,
a search was performed using the computer program BLAST or BLAST2 (Altshul et
al., Methods in E_n~ymology
xø:460-480 (1996)) as a comparison of the ECD protein sequences to a 6 frame
translation of the EST sequence.
Those comparisons resulting in a BLAST score of 70 (or in sori~e cases 90) or
greater that did not encode known
proteins were clustered and assembled into consensus DNA sequences with the
program "phrap" (Phil Green,
University of Washington, Seattle, Washington;
http:l/bozeman.mbt.washington.edulphrap.docslphrap.html).
A consensus DNA sequence was assembled relative to other EST sequences using
phrap. Based on the
consensus sequence obtained, oligonucleotides were synthesized: 1) to identify
by PCR a cDNA library that contained
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the sequence of interest, and 2) for use as probes to isolate a clone of the
full-length coding sequence for PR0531.
A pair of PCR primers (forward and reverse) were synthesized:
forward PCR prim~e_r 5'-CTGAGAACGCGCCTGAAACTGTG-3' (SEQ 1D N0:406);
reverse PCR primer 5'-AGCGTTGTCATTGACATCGGCG-3' (SEQ ID N0:407).
Additionally, a synthetic oiigonucleotide hybridization probe was constructed
from the consensus DNA sequence
which had the following nucleotide sequence:
hybridisation probe
5'-TTAGTTGCTCCATTCAGGAGGATCTACCCTTCCTCCTGAAATCCGCGGAA-3' (SEQ 1D N0:408).
In order to screen several libraries for a source of a full-length clone, DNA
from the libraries was screened
by PCR amplification with the PCR primer pair identified above. A positive
library was then used to isolate clones
encoding the PR0531 gene using the probe oligonucleotide and one of the PCR
primers. RNA for construction of
the cDNA libraries was isolated from human fetal brain tissue (LIB153). The
cDNA libraries used to isolate the
eDNA clones were constructed by standard methods using commercially available
reagents such as those from
Invitrogen, San Diego, CA. The cDNA was primed with oligo dT containing a NotI
site, linked with blunt to SaII
hemikinased adaptors, cleaved with Notl, sized appropriately by gel
electrophoresis, and cloned in a defined
orientation into a suitable cloning vector (such as pRKB or pRICD; pRKSB is a
precursor of pRKSD that does not
contain the SfI site; see, Holmes et al., c' nee, X53:1278-1280 (1991)) in the
unique XhoI and NotI sites.
DNA sequencing of the clones isolated as described above gave thE: full-length
DNA sequence for PR0531
(herein designated as UNQ332 (DNA48314-1320)] (SEQ ID N0:404) and the derived
protein sequence for PR0531:
The entire represenmtive nucleotide sequence of UNQ332 (DNA48314-1320) is
shown in Figure 162 (SEQ
ID N0:404}. 1t is understood that the actual sequence is that within the clone
deposited with the ATCC as
DNA48314-1320. Clone UNQ332 tDNA48314-1320) contains a single open reading
frame with an apparent
translational initiation site at nucleotide positions I71-173 and ending at
the stop colon at nucleotide positions 2565
2567 (Figure 162). The predicted polypeptide precursor is 789 amino acids long
(Figure 163). The full-length
PR0531 protein shown in Figure 163 has an estimated molecular weight of about
87,552 daltons ate a p! of about
4.84. Clone UNQ332 (DNA48314-1320) has been deposited with the ATCC on March
26, 1998:
Analysis of the amino acid sequence of the full-length PR0531 polypeptide
suggests that portions of it
possess significant homology to protocadherin 3. Moreover, PR0531 is found in
the brain, Like other protocadherins,
thereby indicating that PR0531 is a novel member of the cadherin superfamily.
Still analyzing the amino acid sequence of SEQ ID N0:405, the cadherin
exaacellular repeated domain
signature is found at about amino acids 122-132, 231-241, 336-34b, 439-449 and
549-559 of SEQ ID N0:405. An
ATP/GTP-binding site motif A (P-loop) is found at about amino acids 285-292 of
SEQ 1D N0:405. N-glycosyladon
sites are found at least at about amino acids 56?-570, 786-790, 418-421 and
336-339 of SEQ ID NO:405. The signal
peptide is at about amino acids 1-26, and the transmembrane domain is at about
amino acids 685-712 of SEQ ID
N0:405.
EXAMPLE 65: Isolation of~~j~A Clones En~od~g Human PRO 34
A consensus sequence was obtained relative to a variety of EST sequences as
described in Example 1 above,
wherein the cons sequence obtained is herein designated DNA43038. Based on the
43048 consensus sequence,
oiigonucleotides were synthesized; 1} to identify by PCR a cDNA library that
contained the sequence of interest, and
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2) for use as probes to isolate a clone of the full-length coding sequence for
PR0534.
A pair of PCR primers (forward and reverse) were synthesized:
f~,rward PCR primer S'-CACAGAGCCAGAAGTGGCGGAATC-3' (SEQ ID N0:411);
reverse PCR primer S'-CCACATGTTCCTGCTCTTGTCCTGG-3' (SEQ ID N0:412).
Additionally, a synthetic oligonucleotide hybridization probe was constructed
from the consensus DNA43038
sequence which had the following nucleotide sequence:
hybridization probe
S'-CGGTAGTGACTGTACTCTAGTCCTGTTTTACACCCCGTGGTGCCG-3' {SEQ ID NO:413).
In order to screen several libraries for a source of a full-length clone, DNA
from the libraries was screened
by PCR amplification with the PCR primer pair identified above. A positive
library was then used to isolate clones
~ encoding the PR0534 gene using the probe oligonucleotide and one of the PCR
primers. RNA for conswction of
the cDNA libraries was isolated from human fetal lung tissue (LIB26).
DNA sequencing of the clones isolated as described above gave the full-length
DNA sequence far PR0534
[herein designated as UNQ335 (DNA48333-1321)) (SEQ ID N0:409) and the derived
protein sequence for PR0534.
The entire nucleotide sequence of UNQ33S (DNA48333-1321) is shown in Figure
164 (SEQ ID N0:409).
Clone UNQ335 (DNA48333-1321) contains a single open reading frame with an
apparent translational initiation site
at nucleotide positions 87-89 and ending at the stop codon at nucleotide
positions 1167-1169 {Figure 164). The
predicted polypeptide precursor is 360 amino acids long (Figure 165). The full-
length PR0534 protein shown in
Figure 165 has an estimated molecular weight of about 39,885 daltons and a pI
of about 4.?9. Clone UNQ33S
(DNA48333-1321) has been deposited with ATCC on March 26, 1998. It is
understood that the deposited clone
contains the actual sequence, and that the sequences provided herein are
representative based on current sequencing
techniques.
Analysis of the amino acid sequence of the full-length PR0534 polypeptide
suggests that portions of it
possess significant sequence identity with the protein disulfide isomerase,
thereby indicating that PR0534 may be a
novel disulfide isomerase.
Still analyzing the amino acid sequence of PR0534, the signal peptides is at
about amino acids 1-25 of SEQ
ID N0:410. The transmembrane domain is at about amino acids 321-340 of SEQ ID
N0:410. The disulfide
isomerase corresponding region is at amino acids 212-302 of SEQ ID N0:410. The
thioredoxin domain is at amino
acids 211-227 of SEQ ID N0:410. N-glycosylation sites are at: 165-168, 18I-
184, 187-190, 194-197, 206-209, 278-
281, and 293-296 of SEQ ID N0:410. The corresponding nucleotides can routinely
be determined from the
sequences provided herein. PR0534 has'a transmembrane domain rather than an ER
retention peptide like other
protein disulfide isomerases. Additionally, PR0534 may have an intron at the 5
prime end.
EXAMPLE 66: lcntation of cDNA Clones Enco~tQ r~uman PR0697
A consensus sequence was obtained relative to a variety of EST sequences as
described in Example 1 above,
wherein the consensus sequence obtained is here'sn designated DNA43052. Based
on this consensus sequence,
oligorntcleotides were synthesized: 1) to identify by PCR a cDNA library that
contained the sequence of interest, and
2) for use as probes to isolate a clone of the full-length coding sequence for
PR0697.
A pair of PCR primers (forward and reverse) were synthesized:
forward PCR Timer S'-CCTGGCTCGCTGCTGCTGCTC-3' (SEQ ID N0:416);
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~everse PCR primer 5'-CCTCACAGGTGCACTGCAAGCTGTC-3' (SEQ ID N0:4I7).
Additionally, a synthetic oligonucleotide hybridization probe was constn~cted
from the DNA43052 consensus
sequence which had the following nucleotide sequence:
3lybriøization Brobe
5'-CTCTTCCTCTTTGGCCAGCCCGACTTCTCCTACAAGCGCAGAATTGC-3' {SEQ ID N0:418).
In order to screen several libraries for a source of a full-length clone, DNA
from the libraries was screened
by PCR amplification with the PCR primer pair identifted above. A positive
library was then used to isolate clones
encoding the PR0697 gene using the probe oligonueleotide and one of the PCR
primers. RNA for construction of
the cDNA libraries was isolated from human fetal kidney tissue (LIB227).
DNA sequencing of the clotres isolated as described above gave the fall-length
DNA sequence for PR0697
[herein designated as UNQ361 (DNA50920-1325)) (SEQ 1D N0:414) and the derived
protein sequence for PR0697.
The entire nucleotide sequence of UNQ361 (DNA50920-1325) is shown in Figure
166 {SEQ ID N0:414).
Clone UNQ361 (DNA50920-1325) contains a single open reading frame with an
apparent translational initiation site
at nucleotide positions 446 and ending at the stop codon at nucleotide
positions 929-931 (Figure 166). The
predicted polypeptide precursor is 295 amino acids long (Figure 167). The full-
length PR0697 protein shown in
Figure 167 has an estimated molecular weight of about 33,518 daltons and a pi
of about 7.74. Clone UNQ361
(DNA50920-1325) was deposited with the ATCC on March 26, 1998. It is
understood that the deposited clone
contains the actual sequence, and that the sequences provided herein are
representative based on current sequencing
techniques.
Analysis of the amino acid sequence of the full-length PR0697 polypeptide
suggests that portions of it
possess significant sequence identity with sl;RPs, thereby indicating that
PR0697 may be a novel sFRP family
member.
Still analyzing tta: amino acid sequence of PR0697, the signal peptides is at
about amino acids 1-20 of SEQ
ID N0:415. The cystein rich domain, having identity with the frizzled N-
terminus, is at about amino acids 6-153
of SEQ ID N0:415. The corresponding nucleotides can routinely be determined
from the sequences provided herein.
EXAMPLE 67: Isolation of cDNA Clones Encoding Human PR0717
A consensus sequence was obtained relative to a variety of EST sequences as
described in Example 1 above,
wherein the consensus sequence obtained is herein designated DNA42829. Based
on the DNA42829 consensus
sequence, oligonucleotides were synthesized: 1) to identify by PCR a cDNA
library that contained the sequence of
interest, and 2) for use as probes to isolate a clone of the full-length
coding sequence for PR0717.
A pair of PCR primers (forward and reverse) were synthesized:
forward PCR~rimer 5'-AGCTTCTCAGCCCTCCTGGAGCAG-3' (SEQ ID NO:421);
reverse PCR primer 5'-CGGGTCAATAAACCTGGACGCTTGG-3' (SEQ ID N0:422).
Additionally, a synthetic oligonucleotide hybridization probe was constructed
from the DNA42829 consensus
sequence which had the following nucleotide sequence:
$ybridization probe
5'-TATGTGGACCGGACCAAGCACTTCACTGAGGCCACCAAGATTG-3' (SEQ ID N0:423).
In order to screen several libraries for a source of a full-length clone; DNA
from the libraries was screened
by PCR amplification with the PCR primer pair identified above. A positive
library was then used to isolate clones
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encoding the PR0737 gene using the probe oligonucleotide and one of the PCR
primers. RNA far construction of
the cDNA libraries was isolated from human fetal liver tissue (LIB229).
DNA sequencing of.the clones isolated as described above gave the full-length
DNA sequence for PR0717
(herein designated as UNQ385 (DNA50988-1326)] (SEQ ID N0:419) and the derived
protein sequence for PR0717.
The entire nucleotide sequence of UNQ385 (DNA50988-1326) is shown in Figure
168 (SEQ iD N0:419).
Clone UNQ385 (DNA50988-1326) contains a single open reading frame with an
apparent uanslational initiation site
at nucleotide positions 17-19 and ending at the stop codon at nucleotide
positions 1697-1699 (Figure 168). The
predicted polypeptide precursor is 560 amino acids long (Figure 169). The full-
length PR0717 protein shown in
Figure 169 has an estimated molecular weight of about 58,427 daltons and a pI
of about 6.86. Clone UNQ385
(DNA50988-1326) has been deposited with the ATCC on April 28, 1998. Regarding
the sequence, it is understood
that the deposited clone contains the correct sequence, and the sequences
provided herein are based on known
sequencing techniques.
Analysis of the amino acid sequence of the full-length PR0717 polypeptide
suggests that PR0717 may be
a novel 12 transmembrane receptor. The reverse complement strand of DNA50988
has a stretch that matches
identically with human regulatory myosin light strand.
Still analyzing the amino acid sequence of SEQ ID N0:420, transmembrane
domains are at about amino
acids 30-50, 61-79, 98-112, 126-146, 169-182, 201-215, 248-268, 280-300, 318-
337, 341-357, 375-387, and 420-441
of SEQ ID N0:420. N-glycosylation sites are at about amino acids 40-43 and 43-
46 of SEQ ID N0:420. A
glycosaminoglycan attachment site is at about amino acids 468-471 of SEQ ID
N0:420. The corresponding
nucleotides can be routinely determined given the sequences provided herein.
EXAMPLE 68: Isolati n of cDNA Clones Encoding~Human PR0731
A database was used to search expressed sequence tag (EST) databases. The EST
database used herein was
the proprietary EST DNA database LIFESEQTM, of Incyte Pharmaceuticals, Palo
Alto, CA. Incyte clone 2581326
was herein identified and termed DNA42801. Based on the DNA42801 sequence,
oligonucleotides were synthesized:
1 ) to identify by PCR a cDNA library that contained the sequence of interest,
and 2) for use as probes to isolate a
clone of the full-length coding sequence for PR0731.
A pair of PCR primers (forward and reverse) were synthesized:
forward ~('~R~rimer 5'-GTAAGCACATGCCTCCAGAGGTGC-3' (SEQ ID N0:426);
rever_~e PClt~rimer 5'-GTGACGTGGATGCTTGGGATGTTG-3' (SEQ ID N0:427).
Additionally, a synthetic oligonucleotide hybridization probe was constructed
from the DNA42801 sequence which
had the following nucleotide sequence:
hybridization r
5'-TGGACACCTTCAGTATTGATGCCAAGACAGGCCAGGTCATTCTGCGTCGA-3' (SEQ ID N0:428).
In order to screen several libraries for a source of a full-length clone, DNA
from the Libraries was screened
by PCR amplification with the PCR primer pair identified above. A positive
library was then used to isolate clones
encoding the PR0731 gene using the probe oligonucleotide and one of the PCR
primers. RNA for construction of
the cDNA libraries was isolated from human bone marrow tissue (LIB255). The
cDNA libraries used to isolate the
cDNA clones were constructed by standard methods using commercially available
reagents such as those from
Invitrogen, San Diego, CA. The cDNA was primed with oligo dT containing a NotI
site, linked with blunt to SaII
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hemikinased adaptors, cleaved with NotI, sized appropriately by gel
electrophoresis, and cloned in a defined
orientation into a suitable cloning vector (such as pRKB or pRK.D; pRKSB is a
precursor of pRKSD that does not
contain the SfiI site; see, Hohnes et al., Science, x_3:1278-1280 (1991)) in
the unique XhoI and Notl sites.
DNA sequencing of the clones isolated as described above gave the full-length
DNA sequence for PR0731
[herein designated as UNQ395 (DNA48331-1329)] (SEQ ID N0:424) and the derived
protein sequence for PR0731.
The entire nucleotide sequence of UNQ395 (DNA48331-1329) is shown in Figures
170A-B (SEQ ID
N0:424). Clone UNQ395 (DNA48331-1329) contains a single open reading frame
with an apparent translational
initiation site at nucleotide positions 329-331 and ending at the stop codon
at nucleotide positions 3881-3883 (Figures
170A-B). The predicted polypeptide precursor is 1184 amino acids long (Figure
17l ). The full=length PR0731
protein shown in Figure I71 has an estimated molecular weight of about 129,022
daltons and a p! of about 5.2. Clone
IO UNQ395 (DNA48331-1329) was deposited with the ATCC on March 31, 1998.
Regarding the sequence, it is
tmderstood that the deposited clone contains the correct sequence, and the
sequences provided herein are based on
lrnown sequencing techniques.
Analysis of the amino acid sequence of the full-length PR0731 polypeptide
suggests that portions of it
possess significant identity and similarity to members of the protocadherin
family, thereby indicating that PR0731
may be a novel protocadherin.
Still analyzing the amino acid sequence of SEQ ID N0:425, the putative signal
peptide is at about amino
acids 1-13 of SEQ ID N0:425. The transmembrane domain is at amino acids 719-
739 of SEQ ID N0:425. The N-
glycosylation of SEQ ID N0:425 are as follows: 415-418, 582-586, 659-662, 662-
665, and 857-860. The cadherin
extracellular repeated domain signatures are at about amino acids (of SEQ ID
N0:425): I23-133, 232-242, 340-350,
448-458, and 553-563. The corresponding nucleotides can be routinely
determined given the sequences provided
herein.
EXAMPLE 69: Isolation of cDNA Clones Encoding Human PRO218
A consensus sequence was obtained relative to a variety of EST sequences as
described in Example 1 above,
wherein the consensus sequence obtained is herein designated DNA174I1. Two
proprietary Genentech EST
sequences were employed in the consensus assembly and are shown in Figure I74
and 175. Based on the DNA17411
consensus sequence, oligonucleotides were synthesized: I) to identify by PCR a
cDNA library that contained the
sequence of interest, and 2) for use as probes to isolate a clone of the full-
length coding sequence for PR0218.
A pair of PCR primers (forward.and reverse) were synthesized:
forwar~PCR primer 5'-AAGTGGAGCCGGAGCCTTCC-3' (SEQ ID I'd0:433);
reverse PCR primer 5'-TCGTTGTTTATGCAGTAGTCGG-3' (SEQ ID N0:434).
Additionally, a synthetic oligonucleotide hybridization probe was constructed
from the consensus DNA17411
sequence which had the following nucleotide sequence:
hvbridization robe
5'-ATTGTTTAAAGACTATGAGATACGTCAGTATGTTGTACAGG-3' (SEQ ID N0:435).
In order to screen several libraries for a source of a full-length clone, DNA
from the libraries was screened
by PCR amplification with the PCR primer pair identified above. A positive
library was then used to isolate clones
encoding the PR0218 gene using the probe oligonucleotide and one of the PCR
primers. RNA for construction of
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the cDNA libraries was isolated from human fetal kidney tissue (i.IB28).
DNA sequencing of the clones isolated as described above gave the full-length
DNA sequence for PR0218
[herein designated as UNQ192 (DNA30867-1335)] (SEQ ID N0:429) and the derived
protein sequence for PR0218.
The entire nucleotide sequence of UNQ192 (DNA30867-1335) is shown in Figure
172 (SEQ ID N0:429).
Clone UNQ192 (DNA30867-1335) contains a single open reading frame with an
apparent translational initiation site
at nucleotide positions 150-152 and ending at the stop colon at nucleotide
positions 1515-151? (Figure 172). The
predicted polypeptide precursor is 455 amino acids long (Figure 173). The full-
length PR0218 protein shown in
Figure 173 has an estimated molecular weight of about 52,917 daltons and a pI
of about 9.5. Clone UNQ192
(DNA30867-1335) has been deposited with the ATCC on April 28, 1998. Regarding
the sequence, it is understood
that the deposited clone contains the correct sequence, and the sequences
provided herein are based on known
sequencing techniques.
Analysis of the amino acid sequence of the full-length PR0218 polypeptide
suggests that PR0218 may be
a novel transmembrane protein.
Still analyzing the amino acid sequence of SEQ ID N0:430, the putative signal
peptide is at about amino
acids 1 through 23 of SEQ ID N0:430. Transmembrane domains are potentially at
about amino acids 37-55, 81-102,
150-168, 288-311, 338-356, 375-398, and 425-444 of SEQ ID N0:430. N-
glycosylation sites are at about amino
acids 67, 180, and 243 of SEQ ID N0:430. Eukaryotic cobalamin-binding protein
is at about amino acids 151-160
of SEQ ID N0:430. Tl~ cotiesponding nucleotides can be routinely determined
given the sequences provided herein.
EJ~AMPLE 70: ~Z~~n of cDNA Clones Encoding Human PR0768
A consensus sequence was obtained relative to a variety of EST sequences as
described in Example 1 above,
wherein the consensus sequence obtained is herein designated DNA43448. Based
on the DNA43448 consensus
sequence, oligonucleotides were synthesized: 1) to identify by PCR a cDNA
library that contained the sequence of
interest, and 2) for use as probes to isolate a clone of the full-length
coding sequence for PR0768.
A pair of PCR primers (forward and reverse) were synthesized:
forward PCR primer 5'-GGCTGACACCGCAGTGCTCTTCAG-3' (SEQ ID N0:438);
reverse PCR primer 5'-GCTGCTGGGGACTGCAATGTAGCTG-3' (SEQ 1D N0:439).
Additionally, a synthetic oligonucleotide hybridization probe was constructed
from the DNA43448 consensus
sequence which had the following nucleotide sequence:
h_Ybridization probe
5'-CATCCTCCATGTCTCCCATGAGGTCTCTATTGCTCCACGAAGCATC-3' (SEQ ID N0:440).
In order to screen several libraries for a source of a full-length clone, DNA
from the libraries was screened
by PCR amplification with the PCR primer pair identified above. A positive
library was then used to isolate clones
encoding the PR0768 gene using the probe oligonucleotide and one of the PCR
primers. RNA for construction of
the cDNA libraries was isolated from human bone marrow tissue tl,IB255).
DNA sequencing of the clones isolated as described above gave the full-length
DNA sequence for PR0768
[herein designated as UNQ4U6 {DNAS573?-1345)] (SEQ ID N0:436) and the derived
protein sequence for PRO768.
The entire nucleotide sequence of UNQ406 (DNA55737-1345) is shown in Figures
176A-B (SEQ ID
N0:436). Clone UNQ406 (DNA55737-1345) contains a single open reading frame
with an apparent translational
initiation site at nucleotide positions 20-22 and ending at the stop colon at
nucleotide positions 3443-3445 (Figures
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WO 99/46281 ' PCT/US99/05028
176A-B). The predicted polypeptide precursor is 1141 amino acids long (Figure
177). The full-length PR0768
protein shown in Figure 177 has an estimated molecular weight of about 124;671
daltons and a pLof about 5.82.
Clone UNQ406 (DNA55737-1345) has been deposited with the ATCC on Apri16, 1998.
Regarding the sequence,
it is urxlerstood that the deposited clone contains the correct sequence, and
the sequences provided herein are based
on Irnown sequencing techniques.
Analysis of the amino acid sequence of the ful3-length PR0768 polypeptide
suggests that portions of it
possess significant sequence identity and similarity with integrin 7.
Still analyzing the amino acid sequence of SEQ ID N0:437, the putative signal
peptide is at about amino
acids 1-33 of SEQ ID N0:437. The transmembrane domain is at amino acids 1039-
1064 of SEQ ID N0:437. N-
glycosylation sites are at amino acids: 86-89, 746-749, 949-952, 985-988 and
1005-1008 of SEQ ID N0:437.
Integrin alpha chain protein domains are identified at about amino acids: 1064-
1071, 3841109, 1041-.1071, 3I7-346,
443-465, 385-407, 2I5-224, 634-647, 85-99, 322-346, 470-479, 442-46fi, 379-408
and I03I-1047 of SEQ ID
N0:437. The corresponding nucleotides can be routinely determined given the
sequences provided herein.
EXAI~IPLE 71: Isolation of cDNA ~on~s Encod~i g Human PR0773
A cor>sensus sequence was obtained relative to a variety of EST sequences as
described in Example 1 above,
whereia the consensus sequence obtained is herein designated DNA43330. Based
on the DNA43330 sequence,
oligorn»cleotides were synthesized: 1) w identify by PCR a cDNA library that
contained the sequence of interest, and
2) for use as probes to isolate a clone of the full-length coding sequence for
PR0771.
A pair of PCR primers (forward and reverse) were synthesized:
fo~vard PC Timer 5'-CAGCAATATTCAGAAGCGGCAAGGG-3' (SEQ ID N0:443);
reverse PCR primer 5'-CATCATGGTCATCACCACCATCATCATC-3' (SEQ ID N0:444).
Additionally, a synthetic oligonucleotide hybridization probe was constructed
from the DNA43330 consensus
sequence which had the following nucleotide sequence:
hybridization probe
5'-GGTTACTACAAGCCAACACAATGTCATGGCAGTGTTGGACAGTGCTGG-3' (SEQ ID N0:445).
In order to screen several libraries for a source of a full-length clone; DNA
from the libraries was screened
by PCR amplification with the PCR primer pair identified above. A positive
library was then used to isolate clones
eroding the PR0771 gene using the probe oligotrucleotide and one of the PCR
primers. RNA for construction of
the cDNA libraries was isolated from human fetal kidney tissue (LIB28).
DNA sequencing of the clones isolated as described above gave the full-length
DNA sequence for PR0771
[herein designated as UNQ409 (DNA49829-1346)) (SEQ ID N0:441) and the derived
protein sequence for PR0771.
The entire ~tcleotide sequere of UNQ409 (DNA49829-1346) is shown in Figure 178
(SEQ ID N0:441).
Clotre UNQ409 (DNA49829-1346) contains a single open reading frame with an
apparent translationai initiation site
at nucleotide positions 134-136 and ending at the stop codon at nucleotide
positions 1442-1444 (Figure 178). The
predicted polypeptide precursor is 436 amino acids long (Figure 179). 7"he
full-length PR0771 protein shown in
Figure 179 has an estimated molecular weight of about 49,429 daltons and a p1
of about 4.8. Clone UNQ409
(DNA49829-1346) has been deposited with the ATCC on April 7, 1998. Regarding
the sequence, it is understood
that the deposited clone contains the correct sequence, and the sequences
provided herein are based on lrnown
sequencing techniques.
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Analysis of the amino acid sequence of the full-length PR0771 polypeptide
suggests that portions of ii
possess significant homology to the testican protein, thereby indicating that
PR0771 may be a novel testican
homologue.
Still analyzing the amino acid sequence of SEQ ID N0:442, the putative signal
peptide, leucine zipper
pattern, N-myristoylation sites, and thyroglobulin type-1 repeats are also
shown in Figure 179. The corresponding
nucleotides can be routinely determined given the sequences provided herein.
EXAMPLE 72: Isolation of cDNA~lones Encoding Numan PR0733
A consensus sequence was obtained relative to a variety of EST sequences as
described in Example 1 above,
wherein the consensus sequence obtained is herein designated DNA45600. Based
on the DNA45600 consensus
sequence, oligonucleotides were synthesized: 1) to identify by PCR a cDNA
library that contained the sequence of
interest, and 2) for use as probes to isolate a clone of the full-length
coding sequence for PR0733.
A pair of PCR primers (forward and reverse) were synthesized:
forward PCR primer 5'-CCCAGCAGGGATGGGCGACAAGA-3' (SEQ ID N0:448);
reverse PCR primer 5'-GTCTTCCAGTTTCATATCCAATA-3' (SEQ ID N0:449).
Additionally, a synthetic oligonucleotide hybridization probe was constructed
from the DNA45600 consensus
sequence which had the following nucleotide sequence:
~ybridization_p~obe
5'-CCAGAAGGAGCACGGGGAAGGGCAGCCAGATCTTGTCGCCCAT-3' (SEQ ID N0:450).
In order to screen several libraries for a source of a full-length clone, DNA
from the libraries was screened
by PCR amplification with the PCR primer pair identified above. A positive
library was then used to isolate clones
encoding the PR0733 gene using the probe oligonucleotide and one of the PCR
primers. RNA for construction of
the cDNA libraries was isolated from human bone marrow tissue {LIB255).
DNA sequencing of the clones isolated as described above gave the full-length
DNA sequence for PR0733
[herein designated as UNQ411 f,DNA5219b-1348)] (SEQ ID N0:446) and the derived
protein sequence for PR0733.
The entire nucleotide sequence of UNQ411 (DNA52196-1348) is shown in Figures
180A-B (SEQ ID
N0:446). Clone UNQ411 (DNA52196-1348) contains a single open reading frame
with an apparent translational
initiation site at nucleotide positions 106-108 and ending at the stop codon
at nucleotide positions 793-795 (Figures
180A-B). The predicted polypeptide precursor is 229 amino acids long (Figure
181). The full-length PR0733 protein
shown in Figure 181 has an estimated molecular weight of about 26,017 daltons
and a pI of about 4.73. Clone
UNQ411 (DNA52196-1348) has been deposited with the ATCC on April 7, 1998.
Regarding the sequence, it is
understood that the deposited clone contains the correct sequence, and the
sequences provided herein are based on
lawwn sequencing techniques.
Analysis of the amino acid sequence of the full-length PR0733 polypeptide
suggests that portions of it
possess significant sequence identity and similarity to the TllST2 receptor
binding protein precursor and therefore
may have a similar function in cell signaling. If it is a cytoltine, it may be
useful in the treatment of inflammation
and cancer.
Still analyzing the amino acid sequence of SEQ ID N0:447, the putative signal
peptide, transmembratx
domain, N-myristoylation site, and tyrosine !anise site are also shown in
Figure 181. The corresponding nucleotides
can be routinely determined given the sequences provided herein.
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EXAMPLE 73: I~ol tion of eDNA Clones Encoding Human PR016~
An expressed sequence tag (EST) DNA database (Merck/Washington University) was
searched and an EST
AA397543 was identified which showed homology to human pancreatitis-associated
protein. The EST AA397543 .
Cole was purchased anrl its insert obtained and sequenced and the sequence
obtained is shown in Figuze 182 (SEQ
ID N0:451).
The entire nucleotide sequence of PR0162 is shown in Figure 182 (SEQ ID
N0:451). DNA sequencing
of the clone gave the full-length DNA sequence for FR0162 [herein designated
as UNQ429 (DNA56965-1356)) (SEQ
ID N0:451) and the derived protein sequence for PR0162. Clone UNQ429 (DNA56965-
1356) contains a single open
reading frame with an apparent translational inntiation site at nucleotide
positions 86-88 and ending at the stop colon
at nucleotide positions 611-613 (Figure 182). The predicted polypeptide
precursor is 175 amino acids long (Figure
183). The full-length PROI62 protein shown in Figure 183 has an estimated
molecular weight of about 19,330
daltons and a pI of about 7.25. Clone UNQ429 (DNA56965-1356) has been
deposited with the ATCC. Regarding
the sequence, it is understood that the deposited clone contains the correct
sequence, and the sequences provided
herein are based an lrnown sequencing techniques.
Analysis of the amino acid sequence of the full-length PR0162 polypeptide
suggests that portions of it
possess significant homology to the human pancreatitis-associated protein,
thereby indicating that PR0162. may be
a novel pancreatitis-associated protein.
Still analyzing the amino acid sequence of SEQ ID N0:452, the putative signal
peptide is at about amino
acids 1-26 of SEQ ID N0:452. A C-type leetin domain signature is at about
amino acids 146-171 of SEQ ID
N0:452. The corresponding nucleotides can be routinely determined given the
sequences provided herein.
EXALVIPL]~74: Isolation of cDNA Clones Encoding Human O?88
A consensus DNA sequence (designated herein as DNA49308) was assembled
relative to other EST
sequer~es using phrap as described in Example 1 above. Based upon an observed
homology between the DNA49308
consensus sequence and the Incyte EST clo0ne no. 2777282, the Incyte EST clone
no. 2777282 was purchased and
its insert obtained and sequenced, which gave the full-length DNA sequence for
PR0788 [herein designated as
UNQ430 (DNA56405-1357)] (SEQ ID N0:453) and the derived protein sequence for
PR0788.
Clone UNQ430 (DNA56405-I357) contains a single open reading frame with an
apparent translational
initiation site at nucleotide positions 84-86 and ending at the stop colon at
nucleotide positions 459-461 (Figure 184).
The predicted poiypeptide precursor is 125 amino acids long (Figure 185): The
full-length PR0788 protein shown
in Figure 185 has an estimated molecular weight of about 13,115 daltons and a
p1 of about 5.90, CIone UNQ430
(DNA56405-1357) has been deposited with the ATCC. Regarding the sequence, it
is understood that the deposited
clone contains the correct sequence, and the sequences provided herein are
based on latown sequencing techniques.
Still analyzing Figure 185, a signal peptide is shown at about amino acids 1-
17 of SEQ ID N0:454. An N-
glycosylation site is at about amino acids 46-49 of SEQ ID N0:454.
EXAMPLE 75: Isolation of cDNA Clones EncodingJHurr~n PR01008_
A consensus DNA sequence was assembled relative to other EST sequences using
phrap as described in
Example 1 above. This consensus sequence is herein desig aced as DNA49804. An
EST proprietary to Genentcch
was employed in the consensus assembly and is herein designated as DNA16508
(Figure 188; SEQ ID N0:457).
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WO 99/46281 PCT/US99/05028
Based upon an observed homology between the DNA49804 sequence and Merck EST
clone no. AA143670, the
Merck EST clone no. AA143670 was purchased and its insert obtained and
sequenced. That sequence is shown
herein in Figure 186 (SEQ ID N0:455).
Sequencing gave the foil length sequence for PR01008 [herein designated as
UNQ492 (DNA57530-1375))
(SEQ ID N0:455) and the detyved protein sequence for PR01408 were identified.
The entire nucleotide sequence of UNQ492 (DNA57530-1375) is shown in Figure
186 (SEQ ID N0:455).
Clone UNQ492 (DNA57530-I375) contains a single open reading frame with an
apparent translational initiation site
at nucleotide positions 138-140 and ending at the stop colon at nucleotide
positions 936-938 (Figure 186). The
predicted polypeptide precursor is 266 amino acids long (Figure 187). The full-
length PR01008 protein shown in
Figure 187 has an estimated molecular weight of about 28,672 daltons and a pI
of about 8.85. Clone UNQ492
(DNA57530-1375) has been deposited with the ATCC on May 20, 1998. Regarding
the sequence, it is understood
that the deposited clone contains the correct sequence, and the sequences
provided herein are based on known
sequencing techniques.
Analysis of the amino acid sequence of the full-length PR01008 polypeptide
suggests that portions of it
possess significant sequence identity andlor similarity with mdkk-l, thereby
indicating, that PR01008 may be a novel
IS member of this family and have head inducing activity.
Still analyzing the amino acid sequence of SEQ ID N0;456, the putative signal
peptide is at about amino
acids. l-23 of SEQ ID N0:456. The N-glycosylation site is at about amino acids
256-259 of SEQ ID N0:456, and
the fungal zn-(2)-cys(6) binuclear cluster domain is at about amino acids 110-
126 of SEQ ID N0:456. The
corresponding nucleotides can of all the amino acids can be routinely
determined given the sequences provided herein.
EXAMPLE 76: Isolation of cDNA Clues Encoding Human PR01012
A consensus DNA sequence was assembled relative to other EST sequences using
phrap as described in
Example 1 above, wherein the consensus seguence is herein designated DNA49313.
Based on the DNA49313
consensus sequence, oligonucleotides were synthesized: 1) to identify by PCR a
cDNA library that contained the
sequence of interest, and 2} for use as probes to isolate a clone of the full-
length coding sequence for PR01012.
A pair of PCR primers (forward and reverse) were synthesized:
~~rd PCR primer 5'-ACTCCCCAGGCTGTTCACACTGCC-3' (SEQ ID N0:460);
rever;zg PC$ rg imer 5'-GATCAGCCAGCCAATACCAGCAGC-3' {SEQ ID N0:461}.
Additionally, a synthetic oligonucleotide hybridization probe was constructed
from the DNA49313 consensus
sequence which had the following nucleotide sequence:
~,Y~jdization globe
5'-GTGGTGATGATAGAATGCTTTGCCGAATGAAAGGAGTCAACAGCTATCCC-3' (SEQ ID N0:462).
In order to screen several libraries for a source of a full-length clone, DNA
from the libraries was screened
by PCR amplification with the PCR primer pair identified above. A positive
library was then used to isolate clones
encoding the PR01012 gene using the probe oligonucleotide and one of the PCR
primers. RNA for construction of
the cDNA libraries was isolated from human fetal kidney tissue (LIB227).
DNA sequencing of the clones isolated as described above gave the full-length
DNA sequence for PR01012
[herein designated as UNQ495 (DNA5b439-1376)] (SEQ ID N0:458) and the derived
protein sequence for PR01012.
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WO 99146281 PCT/US99/05028
The entire nucleotide sequence of UNQ495 (DNA56439-1376) is shown in Figures
189A-B (SEQ ID
N0:458). Clone UNQ495 (DNA56439-1376) contains a single open reading frame
with an apparent translational
initiarion site at tntcleotide positions 4U4-406 and ending at the stop colon
at nucleotide positions 2645-2647 (Figures
189A-B). The predicted polypeptide precursor is 747 amino acids long (Figure
190). The full-length PR01012
protein shown in Figure 190 has an estimated molecular weight of about 86,127
daltons and a pI of about 7.46. Clone
UNQ495 (DNA56439-1376) has been deposited with ATCC. on May 14, 1998,
Regarding the sequence, it is
understood that the deposited clone contains the correct sequence, and the
sequences provided herein are based on
known sequencing techniques.
Analysis of the amino acid sequence of the full-length PR01012 poiypeptide
suggests that portions of it
possess sequence identity with disulfide isomerase thereby indicating that
PRO1012 may be a novel disulfide
IO isomerase related protein.
Still analyzing the amino acid sequence of SEQ ID N0:459, the cytochrome C
family heme-binding site
signature is at about amino acids 158-163 of SEQ 1D N0:459. The Nt-DNAJ domain
signature is at about amino
acids 77-96 of SEQ 1D N0:459. An N-glycosylation site is at about amino acids
484-487 of SEQ ID N0:459. The
ER targeting sequence is at about amino acids ?44-747 of SEQ ID N0:459. 1t is
understood that the polypeptide and
nucleic acids disclosed can be routinely fortrted with or without, these
portions as desired, in alternative embodiments.
For example, it may be desirable to produce PROlUi2 without the ER targeting
sequence.. Ttte corresponding
nucleotides can be routinely determined given the sequences provided herein.
E7~AMPLE 77: Isolation of cDNA Clones ~ncodiltg. Human PROi014
A consensus DNA sequence was assembled relative to other EST sequences using
phrap as described in
Example 1 abobe, wherein the consensus sequence obtained is herein designated
DNA49811. Based upon an
observed homology between the DNA49811 sequence and Incyte EST clone no.
2612207, Incyte EST clone no:
2612207 was purchased aril its insert was obtained and sequenced, wherein the
sequence obtained is shown in Figure
191 (SEQ OD N0:463).
DNA sequencing gave the full-length DNA sequence for PR01014 [herein
designated as UNQ497
(DNA56409-1377)] (SEQ ID N0:463) and the derived protein sequence for PR01014.
The entire rtueleotide sequence of UNQ497 (DNA56409-1377) is shown in Figure
191 (SEQ ID N0:463).
Clone UNQ497 (DNA56409-1377) contains a single open reading frame with an
apparent translational initiation site
at nucleotide positions 66-68 and ending at the stop colon at nucleotide
positions 966-968 (Figure 191), The
predicted polypepade precursor is 300 amino acids long (Figure 192). the full-
length PR01014 protein shown in
Figure 192 has an estimated molecular weight of about 33,655 daltons and a pI
of about 9.31. Clone UNQ497
(DNASb409-1377) has been deposited with the ATCC on May 20, 1998. Regarding
the sequence, it is understood
that the deposited clone contains the correct sequence, and the sequences
provided herein aye based on known
sequencing techniques.
Analysis of the amino acid sequence of the full-length PR01014 polypeptide
suggests that portions of it
possess sequence idenriry with reductase, thereby indicating that PR01014 may
be a novel member of the reductase
family.
Still analyzing the amino acid sequence of SEQ ID N0:464, the putative signal
peptide is at about amino
acids 1-19 of SEQ ID N0:464, The cAMP and cGMP dependent protein kinase
phosphorylation sites are at about
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WO 99/46281 PCT/US99/05028
amino acids 30-33 and 58-dl of SEQ ID N0:464. Short chain alcohol
dehydrogenase family proteins are at about
amino acids 165-202, 37-49, 112-122 and 210-219 of SEQ ID N0:464. The
corresponding nucleotides of these
domains and any other amino acids provided herein can be routinely determined
given the sequences provided herein.
EXAMPLE 78: Isolation of cDNA Clones EncodingHurt~an PR0~017
A consensus DNA sequence was assembled relative to other EST sequences using
phrap as described in
Example 1 above, wherein that consensus DNA sequence is herein designated
DNA53235. Based upon an observed
homology between the DNA53235 consensus sequence and the Merck EST clone no.
AA243086, the Merck EST
clone no. AA243086 was purchased and its insert obtained and sequenced,
wherein the sequence obtained is shown
in Figure 193 (SEQ ID N0:465). DNA sequencing gave the full-length DNA
sequence for PR01017 [herein
designated as UNQ500 (DNA56112-1379)) (SEQ ID N0:465) and the derived protein
sequence for PR01017.
The entire nucleotide sequence of UNQ500 (DNA56112-1379) is shown in Figure
193 (SEQ ID N0:465).
Clone UNQ500 (DNA56112-1379) contains a single open reading frame with an
apparent aanslational initiation site
at nucleotide positions 128-130 and ending at the stop codon at nucleotide
positions 1370-1372 (Figure 193). The
predicted polypeptide precursor is 414 amino acids long (Figure 194). The full-
length PR01017 protein shown in
Figure 194 has an estimated molecular weight of about 48,414 daltons and a p1
of about 9.54. Clone UNQ500
{DNA56112-1379) has been deposited with the ATCC. Regarding the sequence, it
is understood that the deposited
clone contains the correct sequence, and the sequences provided herein are
based an known sequencing techniques.
Analysis of the amino acid sequence of the full-length PR01017 polypeptide
suggests that portions of it
possess sequence identity with HNK-1 sulfotransferase, thereby indicating that
PR01017 may be a novel
sulfotransferase.
Still analyzing the amino acid sequence of SEQ ID N0:466, the putative signal
peptide is at about amino
acids 131 of SEQ ID N0:466. N-glycosylation sites are at about amino acids 134-
137, 209-212, 280-283 and 370-
273 of SEQ ID N0:466. The TNFRINGFR family cystein-rich region protein is at
about amino acids 329-332 of
SEQ m NO:466. The corresponding nucleotides can be routinely determined given
the sequences provided herein.
The protein can be secreted.
~, M~ PLE 79: Isolation of cDNA Clones EncodinE Human PR0474
A consensus DNA sequence was assembled relative to other EST sequences using
phrap as desctibed in
Example 1 above, wherein the consensus sequence obtained is herein designated
DNA49818. Based upon an
observed homology between the DNA49818 consensus sequence and the Merck EST
clone no. H77889, the Merck
EST clone no.1377889 was purchased and its insert obtained and sequenced,
wherein the sequence obtained is herein
shown in Figure 195 (SEQ ID N0:467). DNA sequencing gave the full-length DNA
sequence for PR0474 [herein
designated as UNQ502 (DNA56045-1380)] (SEQ ID N0:467) and the derived protein
sequence for PR0474.
Ttie entire nucleotide sequence of UNQ502 (DNA56045-1380) is shown in Figure
193 (SEQ ID N0:467).
Clone UNQ502 (DNA56045-1380) contains a single open reading frame with an
apparent translational initiation site
at nucleotide positions 106-108 and ending at the stop codon at nucleotide
positions 916-918 (Figure 195). The
predicted polypeptide precursor is 270 amino acids long (Figure 196). The full-
length PR0474 protein shown in
Figure 196 has an estimated molecular weight of about 28,317 daltons and a p1
of about 6Ø Clone UNQ502
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