Note: Descriptions are shown in the official language in which they were submitted.
WO93/11783 2 1 2 S 4 9 ~ PCT/US92/10427
COMPOSITION AND TREATMENT WIT~
BIOLOGICALLY ACTIVE ~ll~ES AND C~ELATING AGENTS
This invention relates to biologically active peptides and
proteins, and more particularly to compositions and uses
involving biologically active peptides and chelating agents.
In accordance with an aspect of the present invention, there
is provided a composition comprising at least one biologically
active amphiphilic peptide or biologically active protein, and a
chelating agent. The peptide or protein is preferably an ion
channel-forming peptide or protein.
In accordance with another aspect of the present invention,
there is provided a process of inhibiting growth of a target cell
in a host which comprises administering to a host at least one
biologically active amphiphilic peptide or biologically active
protein and a chelating agent. The peptide or protein is
preferably an ion channel-forming peptide or protein. The
biologically active amphiphilic peptide or biologically active
protein and the chelating agent are administered in amounts
effective to inhibit growth of a target cell in a host.
Although the scope of the present invention is not intended
to be limited to any theoretical reasoning, Applicant believes
that certain ions such as, for example, calcium ions and
magnesium ions, decrease the activity of certain biologically
active amphiphilic peptides as antibacterial agents. It is
believed that such decrease in activity may be due to competition
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between the ions, which are positively charged, and the peptides
for negative charges on the bacteria. Applicant also has found
that when a chelating agent, which binds ions such as calcium
and/or magnesium ions, is added to the biologically active
amphiphilic peptide, the activity of the biologically active
peptide is enhanced, or potentiated, whereas when known calcium
channel-blocking agents, such as verapamil and diltiazem, are
added to such peptides, no improvement in the biological activity
of the peptides was obtained.
The term "potentiate", as used herein, means either that the
biologically active amphiphilic peptide or protein is effective
in increasing the biological activity of the chelating agents
against a target cell so thereby the chelating agent may be
employed in an amount lower than which would be reguired for
preventing, destroying or inhibiting growth of a target cell
and/or that the peptide or protein may be employed in an amount
lower than which would be required for preventing, destroying, or
inhibiting growth of a target cell.
Chelating agents which may be employed in accordance with
the present invention are agents which are capable of chelating
calcium. Such chelating agents may also increase the
permeability of the peptide or protein through the membrane of a
target cell, such as a bacterium, as described in Marvin, et al.,
"Enhanced Penetration of Externally Added Macromolecules Through
the Outer Membrane of Gram-Negative Bacteria," found in Actor, et
al. (eds.), Antibiotic Inhibition of Bacterial Cell Surface
Assembly and Function, (1988), pgs. 431-435. Such chelating
agents include ethylene dinitrilo tetraacetic acid (EDTA), and
ethylene glycol bis-~-aminoethyl ether N,N,N',N'-tetraacetic acid
(EGTA).
The biologically active amphiphilic peptides employed in the
present invention are generally water soluble to a concentration
of at least 20 mg/ml at neutral pH in water. In addition, the
structure of such peptide provides for flexibility of the peptide
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molecule. When the peptide is placed in water, it does not
assume an amphiphilic stracture. When the peptide encounters an
oily surface or membrane, the peptide chain folds upon itself
into a rod-like structure, sometimes referred to as an -helical
structure.
In general, such peptides have at least lO amino acids, and
preferably at least 20 amino acids. In most cases, such peptides
do not have in excess of 50 amino acids.
In general, the biologically active peptides or proteins
employed in the present invention are ion channel-forming
peptides or proteins. An ion channel-forming peptide or protein
or ionophore is a peptide or protein which increases the
permeability for ions across a natural or synthetic lipid
membrane. B. Christensen et al. PNAS Vol. 85 P. 5072-76 (July,
1988) describes methodology which indicates whether or not a
peptide or protein has ion channel-forming properties and is
therefore an ionophore. As used herein an ion channel-forming
peptide or protein is a peptide or protein which has ion
channel-forming properties as determined by the method of
Christensen et al.
An amphiphilic peptide or protein is a peptide which
includes both hydrophobic and hydrophilic peptide regions.
The administration of the biologically active amphiphilic
peptides or proteins and chelating agent to a target cell may be
by direct administration to the target cell or systemic or
topical administration to a host which includes the target cell,
in order to prevent, destroy, or inhibit the growth of a target
cell.
The compositions of the present invention may be
administered to a host; for example a human or non-human animal,
in an amount effective to inhibit growth of a target cell. Thus,
for example, the compositions may be used as antimicrobial
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agents, or in particular as anti-bacterial agents, or as
antibiotics.
The term "antimicrobial" as used herein means that the
compositions of the present invention inhibit, prevent, or
destroy the growth or proliferation of microbes such as, for
example, bacteria.
The term "antibiotic" as used herein means that the
compositions employed in the present invention produce effects
adverse to the normal biological functions of the target non-host
cell, including death or destruction and prevention of the growth
or proliferation of the non-host target cell when contacted with
the compositions.
The compositions of the present invention have a broad range
of potent antibiotic activity against a plurality of
microorganisms including Gram-positive and Gram-negative
bacteria. The compositions of the present invention allow a
method for treating or controlling microbial infection caused by
organisms which are sensitive to the peptides herein described.
Such treatment may comprise administering to a host organism or
tissue susceptible to or affiliated with a microbial infection an
antimicrobial amount of at least one of the peptides and a
chelating agent.
Because of the antibiotic and antimicrobial properties of
the compositions, they may also be used as preservatives or
sterilants of materials susceptible to microbial contamination.
The compositions may be administered in combination with a
non-toxic pharmaceutical carrier or vehicle such as a filler,
non-toxic buffer, or physiological saline solution. Such
pharmaceutical compositions may be used topically or systemically
and may be in any suitable form such as a liquid, solid,
semi-solid, injectable solution, tablet, ointment, lotion, paste,
capsule, or the like. The compositions may also be used in
combination with adjuvants, protease inhibitors, or compatible
drugs where such a combination is seen to be desirable or
SUBSTITUTE SHEE~
WO93/11783 2 1 2 ~ 4 9 4 PCT/US92/10427
advantageous in controlling infection caused by harmful
microorganisms including bacteria.
The compositions of the present invention may be
administered to a host; in particular an animal, in an effective
antibiotic and/or anti-microbial amount.
~ Depending on the use, a composition in accordance with the
invention will contain an effective anti-microbial amount of one
or more of the hereinabove described peptides which have such
activity.
The compositions of the present invention may be used in the
treatment of external burns and to treat and/or prevent skin and
burn infections. In particular, the compositions may be used to
treat skin and burn infections caused by organisms such as, but
not limited to, P. aeruginosa, S. aureus, and other Streptococcus
species.
The compositions are also useful in the prevention or
treatment of eye infections. Such infections may be caused by
bacteria such as, but not limited to, P. aeruginosa, S. aureus,
and N. qonorrhoeae.
The compositions may also be administered to plants in an
effective antimicrobial amount to prevent or treat microbial
contamination thereof.
In general, the peptide or protein is employed to provide
peptide dosages of from l mg to 500 mg per kilogram of host
weight, when administered systemically. When administered
topically, the peptide or protein is used in a concentration of
from .05% to 10%.
The chelating agent, such as those hereinabove described, or
derivatives or analogues thereof, when used topically, is
generally employed in a concentration of about .001% to about
25%. When used systemically, the chelating agent is generally
employed in an amount of from l mg/kg to about 2 grams/kg of host
weight per day.
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; -6-
In accordance with a preferred embodiment, the peptide used
in conjunction with a chelating agent such as those hereinabove
described, or derivatives or analogues thereof is a basic
(positively charged) polypeptide having at least ten amino acids
wherein the polypeptide includes both hydrophobic and hydrophilic
amino acids. In one embodiment, the polypeptide may have at
least sixteen amino acids wherein the polypeptide includes at
least eight hydrophobic amino acids and at least eight
hydroplilic amino acids. Still more particularly, the
hydrophobic amino acids are in groups of two adjacent amino
acids, and each group of two hydrophobic amino acids is spaced
from another group of two hydrophobic amino acids by at least one
amino acid other than a hydrophobic amino acid (preferably at
least two amino acids) and generally by no greater than four
amino acids, and the amino acids between pairs of hydrophobic
amino acids may or may not be hydrophilic.
The hydrophilic amino acids are generally also in groups of
two adjacent amino acids in which at least one of the two amino
acids is a basic hydrophilic amino acid, with such groups of two
hydrophilic amino acids being spaced from each other by at least
one amino acid other than a hydrophilic amino acid (preferably at
least two amino acids) and generally no greater than four amino
acids, and the amino acids between pairs of hydrophilic amino
acids may or may not be hydrophobic.
In accordance with a particularly preferred embodiment, the
polypeptide comprises a chain of at least four groups of amino
acids, with each group consisting of four amino acids. Two of
the four amino acids in each group are hydrophobic amino acids,
and two of the four amino acids in each group are hydrophilic,
with at least one of the hydrophilic amino acids in each group
being a basic hydrophilic amino acid and the other being a basic
or neutral hydrophilic amino acid.
The hydrophobic amino acids may be selected from the class
consisting of Ala, Cys, Phe, Gly, Ile, Leu, Met, Val, Trp, Tyr,
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norleucine (Nle), norvaline (Nva), and cyclohexylalanine (Cha).
The neutral hydrophilic amino acids may be selected from the
class consisting of Asn, Gln, Ser, and Thr. The basic
hydrophilic amino acids may be selected frc~ the class consisting
of Lys, Arg, His, Orn, homoarginine (Har), 2,4-diaminobutyric
acid (Dbu), and p-aminophenylalanine.
Each of the groups of four amino acids may be of the
sequence ABCD, BCDA, CDAB, or DABC, wherein A and B are each
hydrophobic amino acids and may be the same or different, one of
C or D is a basic hydrophilic amino acid, and the other of C or D
is a basic or neutral hydrophilic amino acid and may be the same
or different. In a preferred embodiment, the polypeptide chain
may comprise 5 or 6 groups of this sequence. In each group, each
of A, B, C and D may be the same in some or all of the groups or
may be different in some or all of the groups.
The polypeptide chain preferably has at least 20 amino
acids, and no greater than 50 amino acids. It is to be
understood, however, that the polypeptide does not have to
consist entirely of the groups described above. The polypeptide
may have amino acids extending from either or both ends of the
noted groups forming the polypeptide chain and/or there may be
amino acids between one or more of the at least four groups and
still remain within the scope of the invention.
The groups of amino acids may be repeating groups of amino
acids, or the amino acids in the various groups may vary provided
that in each group of the at least four groups of amino acids
there are two hydrophobic and two hydrophilic amino acids as
hereinabove noted.
Thus, in a preferred embodiment, the biologically active
polypeptide comprises a chain including at least four groups of
amino acids, each containing four amino acids. Two of the four
amino acids in each group are hydrophobic, at least one amino
acid is basic hydrophilic, and the remaining one is basic or
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neutral hydrophilic, with the polypeptide chain preferably having
at least 20 amino acids but no greater than 50 amino acids.
In one embodiment, each of the at least four groups of amino
acids which are in the peptide chain is of the sequence A-B-C-D,
B-C-D-A, C-D-A-B or D-A-B-C wherein A and B are hydrophobic
amino acids, one of C or D is basic hydrophilic amino acid, and
the other of C or D is basic or neutral hydrophilic amino acid.
The resulting polypeptide chain, therefore, may have one of the
following sequences:
(Xl)a(A-B-C-D)n(yl)b
(X2)a(B-C-D-A)n(Y2)b
(X3)a(c-D-A-B)n(y3)b
(x4)a(D-A-B-c)n(y4)b
wherein Xl is D; C-D- or B-C-D-, Yl is -A or
-A-B or -A-B-C
X2 is A-, D-A- or C-D-A-
Y2 is -B, -B-C or B-C-D
X3is B-, A-B-, D-A-B-
Y3 is -C, -C-D, -C-D-A
X4is C-, B-C-, A-B-C-
Y4 is -D, -D-A, -D-A-B
a is 0 or l; b is 0 or l
and n is at least 4.
It is to be understood that the peptide chain may include
amino acids between the hereinabove noted groups of four amino
acids provided that the spacing between such groups and the
charge on the amino acids does not change the characteristics of
the peptide chain which provide amphiphilicity and a positive
charge and do not adversely affect the folding characteristics of
the chain to that which is significantly different from one in
which the hereinabove noted group of four amino acids are not
spaced from each other.
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WO93/11783 PCT/US92/10427
_g
As representative examples of peptides in accordance with
the present invention, there may be mentioned.
I Ala-Phe-Ser-Lys-Ala-Phe-Ser-Lys-Ala-Phe-Ser-
Lys-Ala-Phe-Ser-Lys-Ala-Phe-Ser-Lys (SEQ ID
NO:l)
II Ala-Phe-Ser-Lys-Ala-Phe-Ser-Lys-Ala-Phe-Ser-
Lys-Ala-Phe-Ser-Lys-Ala-Phe-Ser-Lys-Ala-Phe-
Ser-Lys-(SEQ lD NO:2)
III Phe-Ser-Lys-Ala-Phe-Ser-
Lys-Ala-Phe-Ser-Lys-Ala-
Phe-Ser-Lys-Ala-(SEQ ID NO:3)
IV Ser-Lys-Ala-Phe-Ser-Lys-Ala-
Phe-Ser-Lys-Ala-Phe-Ser-Lys-Ala-
Phe-Ser-Lys-Ala-Phe-(SEQ ID NO:4)
V Lys-Ala-Phe-Ser-Lys-Ala-Phe-Ser-Lys-Ala-Phe-Ser-
Lys-Ala-Phe-Ser-(SEQ ID NO:5)
The peptide may have amino acids extending from either end
of the chain. For example, the chains may have a Ser-Lys
sequence before the "Ala" end, and/or an Ala-Phe sequence after
the "Lys" end. Other amino acid sequences may also be attached
to the "Ala" and/or the "Lys" end.
Similarly, in any polypeptide chain having at least four
groups of amino acids of the sequence as described above, the
chain may have, for example, a C-D sequence before the first
A-B-C-D group. Also other amino acid sequences may be attached
to the "A" and/or the "D" end of one of these polypeptide chains.
Also there may be amino acids in the chain which space one or
more groups of the hereinabove noted four amino acids from each
other.
The peptides may be produced by known techniques and
obtained in substantially pure form. For example, the peptides
may be synthesized on an automatic synthesizer. Journal of the
American Chemical Society, Vol. 85 Pages 2149-54(1963). It is
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10-
also possible to produce such peptides by genetic engineering
techniques.
In accordance with another preferred embodiment, the peptide
employed in conjunction with a chelating agent such as those
hereinabove described, or derivatives or analogues thereof may be
a magainin peptide.
A magainin peptide is either a magainin such as Magainin J,
II or III or an analogue or derivative thereof. The magainin
peptides may include the following basic peptide structure X12
-- Rll-Rll-R12-R13-Rll-R14-R12-Rll-R14-
12 R11 R11 R11 R14a (R15)n R14a R14
wherein R11 is a hydrophobic amino acid, R12 is a basic
hydrophilic amino acid; R13 is a hydrophobic, neutral
hydrophilic, or basic hydrophilic amino acid; R14 and R14a are
hydrophobic or basic hydrophilic amino acids, R15 is glutamic
acid or aspartic acid, or a hydrophobic or basic hydrophilic
amino acid, and n is O or 1. In a preferred embodiment, R13 is a
hydrophobic or neutral hydrophilic amino acid, R14a is a
hydrophobic amino acid, and R15 is glutamic acid or aspartic
acid.
Thus, for example, a magainin peptide may include the
following structure:
Y12 - X12,
where X12 is the hereinabove described basic peptide
structure and Y12 is
( i ) R12
(ii) R14 - R12;
(iii) Rll~R14a~R12;
( iv ) R14-Rl l-Rl4a-Rl2
R11, R12, R14, and R14a are as previously defined.
A magainin peptide may also have the following structure:
-X12-Z12-
wherein X12 is as previously defined and Z12 is:
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(i) Rl6 where Rl6 is a basic hydrophilic amino acid or
asparagine or glutamine; or
(ii) Rl6-Rl7 where Rl7 is a neutral hydrophilic amino
acid, a hydrophobic amino acid, or a basic hydrophilic amino
acid. Preferably, Rl7 is a neutral hydrophilic amino acid.
A magainin peptide may also have the following structure:
(Y12)a X12 (Z12)b
where Xl2, Yl2, and Z12 are as previously defined, and a is
O or l and b is O or l.
The magainin peptides may also include the following basic
peptide structure Xl3:
R14-R11-R14 -R12-Rll-Rl1-R12-R13-Rl1-R14-
R12 -Rl 1 -Rl 1 -R12 - ~
ll~ Rl2~ Rl3~ Rl4~ and Rl4a are amino acids as
hereinabove described.
The magainin peptide may also include the following
structure Xl3-Zl~; wherein Xl3 is the hereinabove described basic
peptide structure and Z13 is
ll)n ( ll)n (Rll)n-(Rl4a)n-(Rl5)n-(Rl4a)n-
(Rl4) ~(Rl6)n~(Rl7)n~~ wherein Rll, Rl4, Rl4a, lS' 16
Rl7 are amino acids as hereinabove described, and n is O or l,
and each n may be the same or different.
The magainin peptides generally include at least fvurteen
amino acids and may include up to forty amino acids. A magainin
peptide preferably has 22 or 23 amino acids. Accordingly, the
hereinabove described basic peptide structures of a magainin
peptide may include additional amino acids at the amino end or at
the carboxyl end, or at both ends.
As representative examples of such magainin peptides, there
may be mentioned the following peptides, the structures of which
are given in the accompanying sequence listing as well as
appropriate analogues and derivatives thereof:
(a) (NH2) (SEQ ID NO:6) (OH) or (NH2)
(Magainin I)
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(b) (NH2) (SEQ ID NO:7) (OH) or (NH2)
(Magainin II)
(c) (NH2) (SEQ ID NO:8) (OH) or (NH2)
(Magainin III)
The following are examples of peptide derivatives or analogs
of the basic structure:
(d) (NH2) (SEQ ID NO:9) (OH) or (NH2)
(e) (NH2) (SEQ ID NO:10) (OH) or (NH2)
(f) (NH2) (SEQ ID NO:ll) (OH) or (NH2)
Magainin peptides are described in Proc. Natl. Acad Sci.
Vol. 84 pp. 5449-53 (Aug. 1987). The term "magainin peptides" as
used herein refers to the basic magainin structure as well as
derivatives and analogs thereof, including but not limited to the
representative derivatives or analogs.
In accordance with a further embodiment, the peptide
employed in conjunction with a chelating agent may be a PGLa
peptide or an XPF peptide.
A PGLa peptide is either PGLa or an analogue or derivative
thereof. The PGLa peptides preferably include the following
basic peptide structure X14:
11 17 R12 Rll R14 Rll Rll
11 R14 R12 Rll Rll R12 Rll
11 11 12
11' 12' 14' 17 P Y
The PGLa peptides generally include at least seventeen amino
acids and may include as many as forty amino acids. Accordingly,
the hereinabove described basic peptide structure for a PGLa
peptide may include additional amino acids at the amino end or at
the carboxyl end or at both the amino and carboxyl end.
Thus, for example, a PGLa peptide may have the following
structure:
-Yl 4-X14-
where X14 is as previously defined and
Y14 i S
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-13-
(i) Rll;
( ii ) R14-Rll
where Rll is as previously defined.
~ For example, a PGLa like peptide may also have the following
structure:
14 Z14
where X14 is as previously defined; and Z14 is:
(i) Rll; or
( ii ) Rll-Rll
where Rll is as previously defined.
A PGLa peptide may also have the following structure:
( 14)a 14 ( 14)b
where X14; Y14 and Z14 are as previously defined, a is 0 or
1 and b is O or 1.
An XPF peptide is either XPF or an analogue or derivative
thereof. The XPF peptides preferably include the following basic
peptide structure X16:
--Rll-R17-R12-Rll-R14-R18-R17-
Rl l-R14-R12 -Rl 1 -Rll-R12 -
Rll-Rll- Rll-R12-R15-Rl~
11' R12, R14, R15 and R17 are as previously defined
and R18 is glutamine or asparagine.
The XPF peptides generally include at least nineteen amino
acids and may include up to forty amino acids. Accordingly, the
hereinabove described basic peptide structure of XPF may include
additional amino acids at the amino end, or at the carboxyl end
or at both the amino and carboxyl endfi.
Thus, for example, an XPF peptide may include the following
structure:
-Yl~-X16-
where X16 is as previously defined and Y16 is
(i) R11 or
( ii ) R14-R
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where Rll and R14 are is as previously defined.
An XPF peptide may include the following structure:
-X16-Z16-
where X16 is as previously defined and Z16 is
(i) 11; or
(ii) 11 18; or
(iii) Rll-R18-Proline; or
( iv) Rll-R18-Prline R12
An XPF peptide may also have the following structure:
( 16)a 16( 16)b
where X16, Y16 and Z16 are as previously defined: a is 0 or
1 and b is 0 or 1.
Preferred are XPF or PGLa peptides, which are characterized
by the following as listed in the accompanying sequence listing:
PGLa : (SEQ ID N0:12) (NH2)
XPF : (SEQ ID N0:13)
A review of XPF and PGLa can be found in Hoffman et al, EMB0
J. 2:711-714, 1983; Andreu et al, J. Biochem. 149:531-535, 1985;
Gibson et al J. Biol. Chem. 261:5341-5349, 1986; and Giovannini
et al, Biochem J. 243:113-120, 1987.
In accordance with yet another embodiment, the peptide
employed in conjunction with a chelating agent such as those
hereinabove described, may be a CPF peptide or appropriate
analogue or derviative thereof.
A basic CPF peptide structure as well as analogues and
derivatives thereof are herein sometimes referred to collectively
as CPF peptides.
The CPF peptide is preferably one which includes the
following peptide structure X20:
-R21-R21-R22-R22-R21-R21-R23-R21-
R2 1-R2 1-R23 -R2 1-R2 1 -R24-R25 RZ 1-
wherein R21 is a hydrophobic amino acid;
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WO93~11783 2 1 2 5 4 9 4 PCT/US92/10427
R22 is a hydrophobic amino acid or a basic hydrophilic amino
acid;
R23 is a basic hydrophilic amino acid; and
R24 is a hydrophobic or neutral hydrophilic amino acid; and
R25 is a basic or neutral hydrophilic amino acid.
~ The hereinabove basic structure is hereinafter symbolically
indicated as X20.
The hydrophobic amino acids may be Ala, Cys, Phe, Gly, Ile,
Leu, Met, Val, Trp, Tyr, norleucine (Nle), norvaline (Nva), and
cyclohexylalanine (Cha).
The neutral hydrophilic amino acids may be Asn, Gln, Ser,
and Thr.
The basic hydrophilic amino acids may be Lys, Arg, His, Orn,
homoarginine (Har), 2,4-diaminobutyric acid ~Dbu), and
-aminophenylalanine.
The CPF peptide may include only the hereinabove noted amino
acids or may include additional amino acids at the amino end or
carboxyl end or both the amino and carboxyl end. In general, the
peptide does not include more than 40 amino acids.
The CPF peptides including the above basic peptide structure
may have from l to 4 additional amino acids at the amino end.
Accordingly, such preferred peptides may be represented by the
structural formula:
Y20-X20-
wherein X20 is the hereinabove described basic peptide
structure and Y20 is
(i) R25-' or
(ii) R22-R25; or
(iii) R2l-R22-R25; or
(iv) 22 R21-~22-R25; preferably
Glycine -R21-R22 R2s
wherein R2l, R22, and R25 are as previously defined.
The carboxyl end of the basic peptide structure may also
have additional amino acids which may range from l to 13
additional amino acids.
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-16-
In a preferred embodiment, the basic structure may have from
1 to 6 additional amino acids at the carboxyl end, which may be
represented as follows:
X20 Z20 wherein
X20 is the hereinabove defined basic peptide structure and
Z20 is
(i) R21 '
(ii) R21-R21-;
( iii ) R21 R21 R24;
( iv) R21-R21-R24-R24;
(v) R21 R21 R24 24 26;
(vi) 21 R21~R24~R24-R26-Gln; or
(vii) 21 R2l-R24-R24-R26-Gln-Gln~
wherein R21 and R24 are as previously defined, and R26 is
proline or a hydrophobic amino acid.
Preferred peptides may be represented by the following
structural formula:
(Y2o)a 20 ( 20)b
wherein X20, Y20 and Z20 are as previously defined and a is
0 or 1 and b is 0 or 1.
Representative examples of CPF peptides which are useful in
the present invention have been described in the literature and
comprise the following sequences the structures of which are
given in the accompanying sequence listing:
(SEQ ID N0:14)
(SEQ ID N0:15)
(SEQ ID N0:16)
(SEQ ID N0:17)
(SEQ ID N0:18)
(SEQ ID N0:19)
(SEQ ID N0:20)
(SEQ ID N0:21)
(SEQ ID N0:22)
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(SEQ ID N0:23)
(SEQ ID N0:24)
(SEQ ID N0:25)
(SEQ ID N0:26)
A review of the CPF peptides can be found in Richter, K.,
Egger, R., and Kreil (1986) J. Biol. Chem. 261, 3676-3680;
Wakabayashi, T. Kato, H., and Tachibaba, ~. (1985) Nucleic Acids
Research 13, 1817-1828; Gibson, B.W., Pc ter, L., Williams,
D.H., and Maggio, J.E. (1986) J. Biol. Chem. 261, 5341-5349.
CPF peptides which may be employed in the present invention
are represented by the following:
Glyl2Ser3LeuGly4AlaLeuLysAla5LeuLysIleGly678LeuGlyGly9(10)
GlnGln
Where:
1 = Phe, Leu
2 = Gly, Ala
3 = Phe, Leu
4 = Lys, Leu
= Ala, Gly, Thr
6 = Ala, Thr
7 = His, Asn
8 = Ala, Met, Phe, Leu
9 = Ala, Ser, Thr
= Pro, Leu
The numbered amino acids may be employed as described in any
combination to provide either a basic CPF peptide structure or an
analogue or derivative. The term CPF peptide includes the basic
peptide structure as well as analogues or deriva-ives thereof.
In accordance with yet another embodiment, the peptide may
include one of the following basic structures X31 through X37
wherein:
X31 is -[R31-R32-R32-R33-R31-R32-R32i~n;
X32 is -[R32-R32-R33-R31-R32-R32-R31]- ;
SUBSTITU I E SHEET
w093/1l783 2 1 2 5 ~1 ~ 4 -18- PCT/US92/10427
X33 is -IR32-R33-R3l-R32-R32-R3l-R32l~n;
X34 is -[R33-R31-R32-R32-R31-R32-R32]- ;
X35 is -[R31-R32-R32-R31 R32 32 33 n
36 l 32 32 31 32 32 33 31] n; and
X37 is -lR32-R31-R32-R32-R33-R31-R32]~n;
wherein R31 is a basic hydrophilic amino acid, R32 is a
hydrophobic amino acid, R33 is a neutral hydrophilic or
hydrophobic amino acid, and n is from 2 to 5.
The basic hydrophilic amino acids may be selected from the
class consisting of Lys, Arg, His, Orn, homoarginine (Har),
2,4-diaminobutyric acid (Dbu), and p-aminophenylalanine.
The hydrophobic amino acids may be selected from the class
consisting of Ala, Cys, Phe, Gly, Ile, Leu, Met, Val, Trp and
Tyr,norleucine (Nle), norvaline (Nva), and cyclohexylalanine
(Cha).
The neutral hydrophilic amino acids may be selected from the
class consisting of Asn, Gln, Ser and Thr.
In accordance with one embodiment, when the peptide includes
the structure X31, the peptide may include the following
structure:
Y31-X31, wherein X31 is as hereinabove described, and Y
i s :
(i) R32;
(ii) R32 R32;
(iii) R3l-R32 R32;
(iV) R33-R31-R32 R32;
(v) R32-R33-R3l-R32 R32;
( ) 32 R32 R33 R3l-R32-R32~ wherein R31, R32, and R33 are
as hereinabove described
In accordance with another embodiment, when the peptide
includes the structure X31, the peptide may include the following
structure: -
X31-Z31, wherein X31 is as hereinabove described, and Z31
i s :
SUBSTITUT~ SH~ET
212~49 l
WO93/11783 PCT/US92/10427
-19-
(i) R31;
( ) 31 R32;
(iii) R31-R32 R32;
- (iv) R31-R32 R32 33
(v) 31 R32~R32-R33-R31; or
(vi) R31~R32~R32~R33 R31 32
In accordance with yet another embodiment, the peptide may
include the following structure:
(Y31)a X31 (Z31)b' wherein Y31 and Z31 are as previously
defined, a is 0 or 1, and b is 0 or 1.
When the peptide includes the structure X32, the peptide may
include the following structure:
Y32 ~ X32, wherein X32 is as hereinabove described, and Y32
i s :
(i) R31;
(ii~ R32 R31;
(iii) R32-R32 R31;
(iv~ R31-R32-R32 R31;
(v) 33 R31 R32 R32 R31; or
(vi) R32-R33-R3l-R32 R32 31
In another embodiment, when the peptide includes the
structure X32, the peptide may include the following structure:
X32 ~ Z32~ wherein X32 is as hereinabove described, and Z32
i s :
(i) R32;
) R32 R32;
(iii) R32-R32 R33;
(iV) R32-R32-R33 R31;
(v) 32 R32 R33~R31~R32; or
(vi) R32-R32-R33-R31 R32 32
In accordance with yet another embodiment, the peptide may
include the following structure:
(Y32)a X32 (Z32)b~ wherein Y32 and Z32 are as previously
defined, a is 0 or 1, and b is 0 or 1.
SUBSTITUTE SH~T
wo 93,ll,832 1 2 ~ ~ 9 ~ PCT/US92/10427
-20-
In accordance with another embodiment, when the peptide
includes the structure X33, the peptide may include the following
structure:
Y33 - X33 wherein X33 is as hereinabove described, and Y33
i s :
( ) R32;
( ) 31 R32;
(iii) R32-R31 R32;
(iv) R32-R32-R31 R32;
(v) R31-R32-R32-R31-R32; or
) 33 31 R32 R32 R3l-R32~ wherein R31, R32, and R33 are
as hereinabove described.
In accordance with another embodiment, when the peptide
includes the structure X33, the peptide may include the following
structure:
X33 - Z33 wherein X33 is as hereinabove described, and Z33
i s :
(i) R32;
( ) 32 33;
(iii) R32-R33 R31;
(iv) R32-R33-R3l R32;
(v) 32 R33 R31-R32-R32; or
(vi) R32-R33-R3l-R32 R32 31
In accordance with yet another embodiment, the peptide may
include the following structure:
(Y33)a X33 (Z33)b' wherein Y33 and Z33 are as previously
defined, a is 0 or 1, and b is 0 or 1.
In accordance with yet another embodiment, when the peptide
includes the structure X34, the peptide may include the following
structure:
Y34 - X34, wherein X34 is as hereinabove described, and Y34
i s :
(i) R32;
SUBSTITUTE Sl !~ET
212~4~4
W093/l1783 PCT/US92/10427
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(ii) 32 R32;
(iii) R31-R32 R32;
(iv) R32-R31-R32 R32;
(v) R32-R32-R31-R32-R32; or
( ) 31 R32 R32 R31~R32~R32~ wherein R31, R32 and R33 are
as hereinabove described.
In accordance with another embodiment, when the peptide
includes the structure X34, the peptide may include the following
structure:
X34-Z34, wherein X34 is as hereinabove described, and Z34
i s :
(i) R33;
( ) 33 31;
(iii) R33-R31 R32;
(iv) R33-R31-R32 R32;
( v ) R3 3-R3 1~R32 -R32 R3 1;
(vi) R33-R31-R32-R32 R31 32
In accordance with yet another embodiment, the peptide may
include the following structure:
(Y34)a X34 (Z34)b' wherein X34 and Z34 are as previously
defined, a is 0 or 1, and b is 0 or 1.
In accordance with a further embodiment, when the peptide
includes the structure X35, the peptide may include the following
structure:
Y35-X35, wherein X35 is as hereinabove described, and Y35
i s :
(i) R33;
(ii) R32 R33;
(iii) R32-R32 R33;
(iv) R31-R32 R32 33
(v) R32-R31-R32 R32 33
( ) R32 R32 R31 R32-R32-R33~ wherein R31, R32, and R33 are
as hereinabove described.
. .
SUBSTITUT_ SHEET
212~ 3 '~
W093/11783 PCT/US92/1~27
-22-
In accordance with another embodiment, when the peptide
includes the structure X35, the peptide may include the following
structure:
X35 - Z35 wherein X35 is as hereinabove described, and Z35
i s :
(i) R31;
(ii) 31 32;
(iii) R3l-R32 R32;
(iv) R3l-R32-R32 R3l;
(v) 3l R32~R32-R3l~R32; or
(vi) R3l-R32-R32-R3l R32 32
In accordance with yet another embodiment, the peptide may
include the following structure:
(Y35)a X35 (Z35)b' wherein X35 and Z35 are as previously
defined, a is O or l, and b is O or l.
In accordance with a further embodiment, when the peptide
includes the structure X36, the peptide may include the following
structure:
Y36 ~ X36 wherein X36 is as hereinabove described, and Y36
i s :
(i) R31;
(ii) R33-R31;
(iii) R32~R33 R3l;
( iv) R32-R32-R33 R31 i
(v) 3l R32-R32-R33-R3l; or
( ) 32 R3l R32 R32-R33-R3l~ wherein R3l, R32, and R33 are
as hereinabove described.
In accordance with another embodiment, when the peptide
includes the structure X36, the peptide may include the following
structure:
X36-Z36, wherein X36 is as hereinabove described, and Z36
i s :
(i) R32;
(ii) R32-R32;
SlJ~STITl)TE SHEET
WO93/11783 2 1 2 5 4 9 ~ PCT/US92/10427
-23-
(iii) R32-R32 R31;
(iv) R32-R32-R31 R32;
(v) R32-R32-R31-R32 R32;
- (vi) R32-R32-R31-R32 R32 33
In accordance with yet another embodiment, the peptide may
include the following structure:
(Y36)a X36 (Z36)b' wherein Y36 and Z36 are as previously
defined, a is O or 1, and b is 0 or 1.
In accordance with one embodiment, when the peptide includes
the structure X37, the peptide may includes the structure
Y37-X37, wherein X37 is as hereinabove described, and Y37 is:
( ) 32;
( ) 31 R32;
(iii) R33-R31 R32;
(iv) R32-R33-R31 R32;
(v) R32-R32-R33-R31 R32;
( ) 31 R32 R32 R33-R31-R32~ wherein R31, R32, and R33 are
as hereinabove described.
In accordance with a further embodiment, when the peptide
includes the structure X37, the peptide may include the following
structure:
X37 - Z37 wherein X37 is as hereinabove described, and Z37
i s :
(i) R32;
) R32 R31;
(iii ) R32-R31 R32;
(iv) R32-R31-R32 R32;
(v) 32 R31~R32~R32~R33; or
(vi) R32-R31-R32 R32 R33 31
In accordance with yet another embodiment, the peptide may
include the following structure:
(Y37)a X37 (Z37)b~ wherein Y37 and Z37 are as previously
defined, a is 0 or 1, and b is 0 or 1.
SUBSTITUTE SHEET
212~94
W093/11783 PCT/US92/10427
-24-
In a preferred embodiment, n is 3, and most preferably the
peptide has one of the following structures:
(Lys Ile Ala Gly Lys Ile Ala)3-NH2 (SEQ ID NO:27)
(Lys Ile Ala Lys Ile Ala Gly)3-NH2 (SEQ ID NO:28)
(Lys Ile Ala Gly Lys Ile Gly)3-NH2 (SEQ ID NO:29)
(Lys Leu Ala Gly Lys Leu Ala)3-NH2 (SEQ ID NO:30)
(Lys Phe Arg Gly Lys Phe Ala)3-NH2 (SEQ ID NO:31)
(Lys Ala Leu Ser Lys Ala Leu)3-NH2 (SEQ ID NO:32)
(Lys Leu Leu Lys Ala Leu Gly)3-NH2 (SEQ ID NO:33)
(Lys Ala Ile Gly Lys Ala Ile)3-NH2 (SEQ ID NO:34)
(Gly Ile Ala Lys Ile Ala Lys)3-NH2 (SEQ ID NO:35)
(Lys Ile Ala Lys Ile Phe Gly)3-NH2 (SEQ ID NO:36)
(Gly Ile Ala Arg Ile Ala Lys)3-NH2 (SEQ ID NO:37)
(Lys Phe Ala Arg Ile Ala Gly)3-NH2 (SEQ ID NO:38)
(Gly Phe Ala Lys Ile Ala Lys)3-NH2 (SEQ ID NO:39)
(Lys Ile Ala Gly Orn Ile Ala)3-NH2 (SEQ ID NO:40)
(Lys Ile Ala Arg Ile Ala Gly)3-NH2 (SEQ ID NO:41)
(Orn Ile Ala Gly Lys Ile Ala)3-NH2 (SEQ ID NO:42)
(Gly Ile Ala Arg Ile Phe Lys)3-NH2 (SEQ ID NO:43)
(Lys Nle Ala Gly Lys Nle Ala)3-NH2 (SEQ ID NO:44)
(Lys Nle Ala Gly Lys Ile Ala)3-NH2 (SEQ ID NO:4S)
(Lys Ile Ala Gly Lys Nle Ala)3-NH2 (SEQ ID NO:46)
(Lys Nva Ala Gly Lys Nva Ala)3-NH2 (SEQ ID NO:47)
(Lys Nva Ala Gly Lys Ile Ala)3-NH2 (SEQ ID NO:48)
(Lys Leu Leu Ser Lys Leu Gly)3-NH2 (SEQ ID NO:49)
(Lys Leu Leu Ser Lys Phe Gly)3-NH2 (SEQ ID NO:50)
(Lys Ile Ala Gly Lys Nva Ala)3-NH2 (SEQ ID NO:51)
(His Ile Ala Gly His Ile Ala)3-NH2 (SEQ ID NO:52)
(Ala Gly Lys Ile Ala Lys Ile)3-NH2 (SEQ ID NO:53)
(Ile Ala Lys Ile Ala Gly Lys)3-NH2 (SEQ ID NO:54)
(Lys Ile Ala Gly Arg Ile Ala)3-NH2 (SEQ ID NO:55)
(Arg Ile Ala Gly Arg Ile Ala33-NH2 (SEQ ID NO:56)
(Lys Val Ala Gly Lys Ile Ala)3-NH2 (SEQ ID NO:57)
(Lys Ile Ala Gly Lys Val Ala)3-NH2 (SEQ ID NO:58)
Sl)BSTITUTE SHEET
2125~94
WO93/11783 PCT/US92/1~27
-25-
(Ala Lys Ile Ala Gly Lys lle)3-NH2 (SEQ ID NO:59)
(Orn Ile Ala Gly Orn Ile Ala)3-NH2 (SEQ ID NO:60)
(Lys Phe Ala Gly Lys Ile Ala)3-NH2 (SEQ ID NO:61)
(Lys Ile Ala Gly Lys Phe Ala)3-NH2 (SEQ ID NO:62)
(Lys Cha Ala Gly Lys Ile Ala)3-NH2 (SEQ ID NO:63)
(Lys Nle Ala Lys Ile Ala Gly)3-NH2 (SEQ ID NO:64)
(Arg Ile Ala Gly Lys Ile Ala)3-NH2 (SEQ ID NO:65)
(Har Ile Ala Gly Har Ile Ala)3-NH2 (SEQ ID NO:66)
(Xaa Ile Ala Gly Lys Ile Ala)3-NH2 (SEQ ID NO:67)
(Lys Ile Ala Gly Xaa Ile Ala)3-NH2 (SEQ ID NO:68)
In (SEQ ID NO:67) and (SEQ ID NO:68), Xaa is
p-aminophenylalanine.
In accordance with another embodiment, the biologically
active amphiphilic peptide includes the following basic structure
X40
R31-R32-R32-R33-R34-R32-R32-R31-R32-R32-R32-R34-R32-R32,
wherein R31, R32, and R33 are as hereinabove described, and
R34 is a basic hydrophilic or hydrophobic amino acid.
In accordance with one embodiment, the peptide may include
the following structure:
Y40-X40, wherein X40 is as hereinabove described, and Y40
i s :
(i) R32;
(ii) R32-R32;
(iii) R34-R32 R32;
(iv) R33-R34-R32-R32;
(v) R32 33 34 32 32;
(v) 32 R32 R33-R34-R32-R32, or
( ) 31 R32 R32 R33~R34~R32~R32,Wherein R31, R32, R33 and
R34 are as hereinabove described.
In accordance with another embodiment, the peptide may
include the following structure:
X40-Z40, wherein X40 is as hereinabove described and Z40 is:
SIJBSTITUT~ SH~ET
212543~
W093/11783 PCT/US92/10427
-26-
(i) R31;
( ) 31 R32;
(iii) R31-R32 R32;
(iv) R31-R32 R32 33
(v) R31-R32-R32-R33 R34;
(vi) R31-R32-R32-R33-R34-R32; or
( ) 31 R32 R32-R33-R34-R32-R32~ wherein R31, R32, R33,
and R34 are as hereinabove described.
In accordance with yet another embodiment the peptide may
include the following structure:
( 40)a X40 (Z40)b~ wherein Y40 and Z40 are as previously
defined, a is 0 or 1, and b is 0 or 1. In a preferred
embodiment, the peptide has the following structural formula as
indicated in the sequence listing contained herein:
(SEQ ID NO:69)-NH2.
In another preferred embodiment, the peptide has the
following structural formula as indicated in the sequence listing
contained herein:
(SEQ ID N0:70)-NH2.
In accordance with a further embodiment, the peptide has one
of the following structural formulae as indicated in the sequence
listing contained herein:
(SEQ ID N0:71)-NH2
(SEQ ID N0:72)-NH2
(SEQ ID N0:73)-NH2
(SEQ ID N0:74)-NH2
(SEQ ID N0:75)-NH2
(SEQ ID N0:76)-NH2
(SEQ ID N0:77)-NH2
(SEQ ID N0:78)-NH2
(SEQ ID N0:79)-NH2
(SEQ ID N0:80)-NH2
(SEQ ID N0:81)-NH2
(SEQ ID N0:82)-NH2
SUBSTITUTE SH~ET
2125~94
WO93/11783 PCT/US92/10427
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(SEQ ID N0:83)-NH2
(SEQ ID NO:84)-NH2
(SEQ ID N0:85)-NH2
In accordance with another embodiment the peptide has the
following structural formula:
-(Lys Ile Ala Lys Lys Ile Ala)n-, wherein n is from 2 to 5.
Preferably, n is 3, and the peptide has the following structural
formula:
(Lys Ile Ala Lys Lys Ile Ala)3-NH2.
(SEQ ID N0:86)
In accordance with another embodiment, the peptide is
selected from the qroup consisting of the following structural
formulae as given in the accompanying sequence listing:
(SEQ ID N0:87)-NH2
(SEQ ID NO:88)-NH2
(SEQ ID N0:89)-NH2
(SEQ ID N0:90)-NH2.
In accordance with another embodiment, the peptide includes
the following basic structure X50:
41 42 42 41 42 42 R41 R41 R42 R41 R41'
wherein R41 is a hydrophobic amino acid, and R42 is a basic
hydrophilic or neutral hydrophilic amino acid.
In one embodiment, the peptide includes the basic structure
Y50-X50, wherein X50 iS as hereinabove described and Y50 is:
(i) R41;
(ii) R42-R41; or
(iii) R42-R42-R41, wherein R41 and R42 are as hereinabove
describe.
In one embodiment, R41 is leucine. In another embodiment,
R42 is lysine. Representative, examples of such peptides include
those having the following structures:
(SEQ ID N0:91)
(SEQ ID N0:92)
(SEQ ID N0:93)
S~JBSTITUTE SH~ET
W093/11783 212 5 ~ ~ ~ PCT/US92/10427
-28-
(SEQ ID NO:94).
In yet another embodiment, the peptide includes the
following basic structure X52:
42 41 42 42 41 41 R42 R42 R41 R42 R42'
wherein R41 is a hydrophobic amino acid, and R42 is a basic
hydrophilic or neutral hydrophilic amino aci-d.
In one embodiment, R41 is leucine. In another embodiment,
R42 is lysine-
In one embodiment, the peptide includes the basic structureY52-X52, wherein X52 is as hereinabove described, and Y52 is:
(i) R42;
(ii) R41-R42; or
(iii) R4l-R41 R42;
(iv) R42-R4l-R4l R42;
( v ) R42 -R42 -R41 -R41 R42 -
In one embodiment, the peptide may have the following
structure:
Lys Lys Leu Leu Lys Lys Leu Lys Lys Leu
Leu Lys Lys Leu Arg Arg
(SEQ ID NO:95).
In another embodiment, the peptide includes the basic
structure X52-Z52, wherein X52 is as hereinabove described, and
Z52 is:
(i) R41;
(ii) R41-R41; or
(iii) R41-R41 R42;
(iv) R41-R41-R42 R42;
( v ) R41-R41-R42 -R42 R41 -
In one embodiment, the peptide may have the following
structure:
Lys Leu Lys Lys Leu Leu Lys Lys Leu
Lys Lys Leu Leu Lys Lys Leu
SUBSTITUTE SH~ET
W093/11783 2 1 2 5 ~ ~ 4 PCT/US92/10427
-29-
(SEQ ID N0:96).
In another embodiment, the peptide may include the
structure:
- (Y52) -X52-(Z52)b~ wherein X52, Y52' 52
hereinabove described, and a is 0 or 1, and b is 0 or 1.
The above peptides may be acetylated with a CH3C0-group at
the N-terminal.
In accordance with another embodiment, each of the amino
acid residues may be a D-amino acid residue or glycine.
In still another embodiment, the peptide employed in
conjunction with a chelating agent such as those hereinabove
described, or derivatives or analogues thereof, is a cecropin.
The cecropins and analogues and derivatives thereof are described
in Ann. Rev. Microbiol 1987, Vol. 41 pages 103-26, in particular
p. 108 and Christensen at al PNAS Vol. 85 p. 5072-76, which are
hereby incorporated by reference.
The term cecropin includes the basic structure as well as
analogues and derivatives.
In yet another embodiment, the peptide employed in
conjunction with a chelating agent such as those hereinabove
described, or derivatives or analogues thereof is a sarcotoxin.
The sarcotoxins and analogues and derivatives thereof are
described in Molecular Entomology, pages 369-78, in particular p.
375 Alan R. Liss Inc. (1987), which is hereby incorporated by
reference.
The term sarcotoxin includes the basic materials as well as
analogues and derivatives.
Ion channel-forming proteins or peptides which may be
employed include defensins, also known as human neutrophil
antimicrobial peptides (~ P), major basic protein (MBP) of
eosinophils, bactericidal permeability-increasing protein (BPI),
a pore-forming cytotoxin called variously perforin, cytolysin, or
S~JBSTITUTE SHEET
wo g3/ll78~ 1 2 ~ PCT/US92/1~27
-30-
pore-forming protein, lactoferrin, B12-binding protein,
eosinophil cationic protein (ECP), and eosinophil-derived
neurotoxin. Defensins are described in Selsted, et al., J. Clin.
Invest., Vol. 76, pgs. 1436-1439 (1985). MBP proteins are
described in Wasmoen, et al., J. Biol. Chem., Vol. 263, pgs.
12559-12563 (1988). BPI proteins are described in Ooi, et al.,
J. Biol. Chem., Vol. 262, pgs. 14891-14894 (1981). Perforin is
described in Henkart, et al. J. Exp. Med., 160:75 (1984), and in
Podack, et al. J. Exp. Med., 160:695 (1984). Lactoferrin,
B12-binding protein, eosinophil cationic protein, and
eosinphil-derived neurotoxin are described in Elsbach, et al.,
Inflammation: Basic Principles and Clinical Correlates, Gallin,
et al., eds.; pgs. 445-471 (1988). The above articles are hereby
incorporated by reference.
The term ion channel-forming proteins includes the basic
structures of the ion channel-forming proteins as well as
analogues or derivatives.
In accordance with another aspect of the present invention,
there is provided a method of antagonizing the biological
activity of a biologically active amphiphilic peptide or protein,
such as those hereinabove described, in a host. The method
comprises administering to a host (human or animal) that is being
treated with a biologically active peptide or protein an ion
selected from the group consisting of calcium ions and magnesium
ions. The ion is administered in an amount effective to
antagonize the biological activity of the peptide or protein.
Such a method is particularly applicable to a method of
selective treatment. For example, one may wish for the peptide
or protein to be biologically effective in one or more body parts
or organs of a host, but not in others. In such cases, the ion
would be administered to those body parts or organs in which one
did not desire the peptide or protein to have biological effect.
As an illustrative example, if one desired the peptide or protein
to be effective in the stomach of the host but not in the
SUBSTITUT~ SHEET
2125~94
WO93/11783 PCT/US92/10427
-31- -
bloodstream, one would administer the ion to the bloodstream in
an amount effective in antagonizing the biological activity of
the peptide or protein in the bloodstream. The ion may be
administered in an amount up to about 2 grams/kg of host weight.
The invention will now be described with respect to the
following examples; however, the scope of the present invention
is not intended to be limited thereby.
Example 1
The procedure for the following antibacterial assay is based
upon the guidelines of the National Committee for Clinical
Laboratory Standards, Document M7-T2, Volume 8, No. 8, 1988.
Stock solutions of the following Peptides (1) through ~3~,
wherein:
Peptide (1) is (SEQ ID N0:27)
Peptide (2) is (SEQ ID N0:97) amide-terminated; and
Peptide (3) is (Lys Ile Ala Gly Lys Ile Ala)3,
wherein each amino acid residue of Peptide (3) i8 a D-amino
acid residue or glycine, were prepared at a concentration of 512
~g/ml in sterile deionized distilled water and stored at -70C.
The stock peptide solutions are diluted in serial dilutions
(1:2) down the wells of a microtiter plate so that the final
concentrations of peptides in the wells are 0.25, 0.50, 1, 2, 4,
8, 16, 32, 64, 12B, and 256 ~g/ml. 1-5 x 105 CFUs/ml of P.
aeruqinosa ATCC 27853 were added to the wells in full strength or
half-strength Mueller Hinton broth (MHB), or in half-strength MHB
broth to which 0.002 M Ca2 , or 0.002 M Mg2 , or 0.001 M Ca2 and
O.OOlM Mg2 has been added. The organisms are from a mid-log
culture. The inoculum is standardized spectrophotometrically at
60Cnm and is verified by colony counts. The pla~s are incubated
for 16-20 hours at 37C, and the minimal inhibitcry concentration
(MIC) for each peptide is determined. Minimal inhibitory
concentration is defined as the lowest conce~.ration of peptide
SUBSTITUTE SHEtT
W093/11783 212 i ~ 32- PCT/US92/10427
which produces a clear well in the microtiter plate. The MIC
values are given in Table I below.
Table~I
MIC (~g/ml)
MHB 1/2 MHB 1/2 MHB 1/2 MHB 1/2 plus
(half- plus plus O.OOlM Ca2
strength) 0.002M 0.002M and O.OOlMMHB full
ca2+ Mg2+ Mg2+strength
Peptide (1) 8 256 128 128 64
Peptide (2) 4,8 32 8 16,32 8,16
Peptide (3) 4,8 128,256 64 128 64
The above results indicate that calcium and magnesium ions
decrease the activity of Peptides (1) through (3) against P.
aeruginosa.
Example 2
The procedure of Example 1 was repeated, except that the
assays were carried out in full strength Mueller Hinton Broth
(MHB) or cation-adjusted Mueller Hinton Broth (CAMHB). Peptides
(1) and (2) were tested for MIC against P. aeruginosa 27853 in
each of these broths without further additives, and in each of
these broths to which was added 0.05mM, 0.5mM, or 5mM of the
calcium channel blockers verapamil or diltiazem. The results are
given in Table II below.
S~lBSTITl~TE SHEET
W093~11783 2 1 2 ~ ~ 3 ~ PCT/US92/1~27
-33-
Table ~I
- MIC ~g/ml)
CAr~B
Verapamil Diltiazem
Peptide 0 0.OSmM 0.5mM 5mM Q 0.05mM 0.5mM 5mM
(1) 256 128-256 128 128 256 128 128 128-256
(2) }6 8/32 16 8 16 8-16 8 32
MHB
Verapamil Diltiazem
Peptide 0 0.05mM 0.5mM 5mM 0 0.OSmM 0.5mM 5mM
(1) 32 16-32 32 16 32 32 16 16-32
12) 4 4 4/16 4 4 4 2-4 8
SUBSTITUT~ SH~ET
2125~4
WO93/11783 PCT/US92/10427
-34-
The above results indicate that the known calcium channel
blockers verapamil and diltiazem have little or no effect upon
the activity of Peptides (l) or (2) against P. aeruqinosa.
Example 3
Peptides (l), (2), (4), (5), and (6) were tested for
activity against P. aeruqinosa strain 27853 in full or
half-strength Mueller Hinton Broth according to the procedure of
Example l, and in Mueller Hinton Broth to which 0.034mM, or
0.34mM, or 3.4mM of ethylene dinitrilo tetraacetic acid (EDTA) or
ethylene glycol bis (B-aminoethyl ether) NNN'N'-tetraacetic acid
is added. Peptide (4) is amide-terminated Magainin II (SEQ ID
N0:7). Peptide (5) has the following structural formula:
Gly Ile Gly D-Lys D-Phe Leu His Ser
Ala D-Lys D-Lys D-Phe Gly D-Lys
Ala D-Phe Val Gly Glu Ile Met Asn Ser-NH2
Peptide (6) i8 SEQ ID N0:21, amide-terminated. The results
of this assay are given in Table III below:
SUBSTITIJTE S~l~ET
2125494
WO 93/11783 PCI/US92/10427
-35-
TABLE III
MIC ( llg/ml )
SUBSTITUTE Sf~ET
21254~
WO93/11783 PCT/US92/10427
-36-
The above results indicate that, with the exception of
Peptides (2) and (5), EGTA at a concentration of 3.4mM enhances
the activity of the peptides against P. aeruginosa.
Example 4
The procedure for this assay, which is a checkerboard assay,
is performed as described in Antibiotics and Laboratory Medicine,
2nd ed., Victor Lorian, M.D. Editor, pgs. 540-546 (1986).
The checkerboard assay was carried out in a 96-well
microtiter plate having 12 rows and 8 columns of wells. 100~1 of
plain broth is added to every row of wells. 100~1 of the desired
peptide at 512 ~g/ml is added to the top well, and serially
diluted through row 11. 50~1 of the EDTA in varying
concentrations (obtained through serial dilutions), are added to
each appropriate column of wells. 50 ~l of P. aeruginosa strain
27853 is then added to each well. The plate is then incubated at
35-37C for 18 to 24 hours, and the wells are then examined for
the presence of visible growth; i.e., turbidity.
SUBSTITUTE S~EET
WO93/11783 2 1 2 5 4 g 4 PCT/US92/10427
-37-
Peptides (1) and (2) were tested alone or in combination
with EDTA for activity against P. aeruqinosa strain 27853. In
the checkerboard assay in which Peptide (1) and EDTA were tested
for activity against P. aeruqinosa, the MIC of Peptide (1) was
64~g/ml, and the MIC of EDTA was 4mM.
Combinations of varying amounts of Peptide (1) and EDTA
which resulted in inhibition of the growth of P. aeruqinosa, and
the FIC values of each combination, were then determined.
FIC, or Fractional Inhibitory Index, is determined as
follows:
FIC = MIC of peptide in combination MIC of EDTA in combination
MIC of peptide alone MIC of EDTA alone
An FIC value of 0.5 or less is indicative of synergy, a value of
greater than 0.5 but less than 2 is indicative of indifference,
and a value greater than 2 is indicative of antagonism. The
following combinations of Peptide (1) and EDTA were found to be
inhibitory, and the FIC values of each combination are given
herewith.
Combination FIC
16~g/ml Peptide (1) plus 0.125mM EDTA 0.28
4~g/ml Peptide (1) plus 0.25mM EDTA 0.125
2~g/ml Peptide (1) plus 0.5mM EDTA 0.156
l~g/ml Peptide (1) plus lmM EDTA 0.27
l~g/ml Peptide (1) plus 2mM EDTA 0.52
The assay was also performed to determine the MIC values of
Peptide (2), EDTA, and inhibitory combinations thereof. The MIC
of Peptide (2) against P. aeruginosa was 4~g/ml, and the MIC of
EDTA was 8mM. The following combinations of Peptide (2) and EDTA
8UBSMTUTE SHEE~
W093/11783 2 1 2 S ~ ~ ~ PCT/US92/1~27
-38-
were also found to be inhibitory, and FIC values are given
therefor:
U~ST~T~T!E~ S~
W0 93/11783 2 1 2 5 4 9 ~ PCI/US92/10427
-39-
Combination FIC
2~1g/ml Peptide (2) plus 0.25mM EDTA 0.53
2~1g/ml Peptide (2) plus 0.5mM EDTA 0.56
2,ug/ml Peptide (2) plus lmM EDTA 0.63
l,ug/ml Peptide (2) plus 2mM EDTA 0.50
0.5~g/ml Peptide (2) plus 4mM EDTA 0.63
The above results suggest that there is a synergistic
interaction between Peptide (l) and EDTA when employed in
combination to inhibit growth of P. aeruqinosa.
Example 5
Peptides (l) and (2) were tested for MIC against P.
aeruginosa strain 27853, as employed alone or in combination with
3mM, or 4mM or 5mM EGTA. The procedure followed was that of
Example l, and all testing was done using P. aeruginosa grown in
full strength MHB broth. The results of this assay are given in
Table IV below.
Table IV
MIC ~lq/ml
Peptide MHB MHB plus MHB plusMHB plus
3mM 4mM EGTA5mM EGTA
EGTA
(l) 32,64 2 2 2
(2) 4 4 2 2,4
Example 6
The procedure of Example 5 was repeated, except the MIC
values of Peptides (l) and (2) were determined for Peptides (l)
and (2) alone and in combination with 0.0625mM, 0.125mM, 0.25mM,
O.5mM, lmM, 2mM, or 3mM EGTA. The results of this assay are
given in Table V below.
SU~STITUTE SH~ET
21~ I J~
WO 93/11783 40 PCI/US92/10427
-- E 3
:c E
~ E ~
E~
~g ,,
~ ~ 3
E 3 ~,
S E 3
A ~IqeT.
2125~9~
WO93/11783 PCT/US92/10427
-41-
It is to be understood, however, that the scope of the
present invention is not to be limited to the specific
embodiments described above. The invention may be practiced
other than as particularly described and still be within the
scope of the accompanying claims.
SUBSTlTlJTE S~iEET
212~4
WOg3/11783 PCT/US92/10427
-42-
SEQUENCE LISTING
(l) GENERAL INFORMATION:
(i) APPLICANT: Berkowitz, Barry ~
(ii) TITLE OF INVENTION: Composition and Treatment with
Biologically Active Peptides and Chelating Agents.
(iii) NUMBER OF SEQUENCES: 97
(iv) CORRESPONDENCE ADDRESS:
(A) ADDRESSEE: Carella, Byrne, Bain, Gilfillan,
Cecchi & Stewart
(B) STREET: 6 Becker Farm Road
(C) CITY: Roseland
(D) STATE: New Jersey
(E) COUNTRY: USA
(F) ZIP: 07068
(v) COMPUTER READABLE FORM:
(A) MEDIUM TYPE: 3.5 inch diskette
(B) COMPUTER: IBM PS/2
(C) OPERATING SYSTEM: PC-DOS
(D) SOFTWARE: DW4.V2
(vi) CURRENT APPLICATION DATA:
(A) APPLICATION NUMBER: 07/803,629
(B) FILING DATE: 9-DEC-l99l
(C) CLASSIFICATION:
(vii) PRIOR APPLICATION DATA:
SlJBSTlTUTt SH~ET
21254~ l
WO93/11783 PCT/US92/1~27
-43-
(A) APPLICATION NUMBER:
(B) FILING DATE:
-- (viii) ATTORNEY/AGENT INFORMATION:
(A) NAME: Olstein, Elliot M.
(B) REGISTRATION NUMBER: 24,025
(C) REFERENCE/DOCKET NUMBER: 421250-146
(ix) TELECOMMUNICATION INFORMATION:
(A) TELEPHONE: 201-994-1700
(B) TELEFAX: 201-994-1744
(2) INFORMATION FOR SEQ ID NO:l:
(i) SEQUENCE CHARACTERISTICS
(A) LENGTH: 20 amino acids
(B) TYPE: amino acid
(C) STRANDEDNESS:
(D) TOPOLOGY: linear
(ii) MOLECULE TYPE: peptide
(ix) FEATURE:
(D) OTHER INFORMATION: amide- or carboxy-terminated
SUBSTITUT~ SH~ET
212~4~
WO93/11783 PCT/US92/10427
-44-
(xi) SEQUENCE DESCRIPTION: SEQ ID NO:l:
Ala Phe Ser Lys Ala Phe Ser Lys
Ala Phe Ser Lys Ala Phe Ser Lys Ala
Phe Ser Lys
(2) INFORMATION FOR SEQ ID NO:2:
(i) SEQUENCE CHARACTERISTICS:
(A) LENGTH: 24 amino acids
(B) TYPE: amino acid
(C) STRANDEDNESS:
(D) TOPOLOGY: linear
(ii) MOLECULE TYPE: peptide
(ix) FEATURE:
(D) OTHER INFORMATION: amide- or carboxy-terminated
(xi) SEQUENCE DESCRIPTION: SEQ ID NO:2:
Ala Phe Ser Lys Ala Phe Ser Lys Ala Phe
Ser Lys Ala Phe Ser Lys Ala Phe Ser Lys
Ala Phe Ser Lys
(2) INFORMATION FOR SEQ ID NO:3:
(i) SEQUENCE CHARACTERISTICS:
(A) LENGTH: 16 amino acids
(B) TYPE: amino acid
(C~ STRANDEDNESS:
- (D) TOPOLOGY: linear
SIJBST~TUTE SH~T
WO93/11783 2 1 2 5 ~ 9 ~ PCT/US92/1~27
-45-
(ii) MOLECULE TYPE: peptide
(ix) FEATURE:
(D) OTHER INFORMATION: amide- or carboxy-terminated
(xi) SEQUENCE DESCRIPTION: SEQ ID NO:3:
Phe Ser Lys Ala Phe Ser Lys Ala Phe Ser
Lys Ala Phe Ser Lys Ala
(2) INFORMATION FOR SEQ ID NO:4:
(i) SEQUENCE CHARACTERISTICS
(A) LENGTH: 20 amino acids
(B) TYPE: amino acid
(C) STRANDEDNESS:
(D) TOPOLOGY: linear
(ii) MOLECULE TYPE: peptide
(ix) FEATURE:
(D) OT~:R INFORMATION: amide- or carboxy-terminated
(xi) SEQUENCE DESCRIPTION: SEQ ID NO:4:
Ser Lys Ala Phe Ser Lys Ala Phe Ser Lys
Ala Phe Ser Lys Ala Phe Ser Lys Ala Phe
(2) INFORMATION FOR SEQ ID NO:5:
(i) SEQUENCE CHARACTERISTICS
(A) LENGTH: .16 amino acids
- (B) TYPE: amino acid
(C) STRANDEDNESS:
SUBSTITUTE Sl IEET
WO 93/ll783 2 1 2 5 4 ~ 4 PCT/US92/10427
-46-
(D) TOPOLOGY: linear
(ii) MOLECULE TYPE: peptide
(ix) FEATURE:
(D) OTHER INFORMATION: amide- or carboxy-terminated
(xi) SEQUENCE DESCRIPTION: SEQ ID NO:5:
Lys Ala Phe Ser Lys Ala Phe Ser Lys Ala
Phe Ser Lys Ala Phe Ser
(2) INFORMATION FOR SEQ ID NO:6:
(i) SEQUENCE CHARACTERISTICS
(A) LENGTH: 23 amino acids
(B) TYPE: amino acid
(C) STRANDEDNESS:
(D) TOPOLOGY: linear
(ii) MOLECULE TYPE: peptide
(ix) FEATURE:
(A) NAME/KEY: Magainin I peptide.
(D) OTHER INFORMATION: amide- or carboxy- terminated
(x) PUBLICATION INFORMATION:
(A) AUTHOR: Zasloff, Michael
(C) JOURNAL: Proceedings of the National Academy
of Sciences
(D) VOLUME: 84
(F) PAGES: 5449-5453
(G) DATE: AUG - l987
(H) DOCUMENT NUMBER: US 4810777
SIJBSTITUTE Sl lEET
W093/11783 2 1 2 S ~ ~ 4 PCT/US92/10427
-47-
(I) FILING DATE: 04-MAR-1987
(J) PUBLICATION DATE: 07-MAR-1989
(xi) SEQUENCE DESCRIPTION: SEQ ID NO:6
Gly Ile Gly Lys Phe Leu His Ser Ala Gly
Lys Phe Gly Lys Ala Phe Val Gly Glu Ile
Met Lys Ser
(2) INFORMATION FOR SEQ ID NO:7:
(i) SEQUENCE CHARACTERISTICS
(A) LENGTH: 23 amino acids
(B) TYPE: amino acid
(C) STRANDEDNESS:
(D) TOPOLOGY: linear
(ii) MOLECULE TYPE: peptide
(ix) FEATURE:
(A) NAME/KEY: Magainin II peptide.
(D) OTHER INFORMATION: amide- or carboxy- terminated
(x) PUBLICATION INFORMATION:
(A) AUTHOR: Zasloff, Michael
(C) JOURNAL: Proceedinqs of the National Academy
of Sciences
(D) VOLUME: 84
(F) PAGES: 5449-S453
(G) DATE: AUG - 1987
(H) DOCUMENT NUMBER: US 4810777
(I) FILING DATE: 04-MAR-1987
(J) PUBLICATION DATE: 07-MAR-1989
SlJBSTlTUTE Sl IEET
W093/11783 2 1 2 S 4 9 ~ PCT/US92/10427
-48-
(xi) SEQUENCE DESCRIPTION: SEQ ID NO:7
Gly Ile Gly Lys Phe Leu His Ser Ala Lys
Lys Phe Gly Lys Ala Phe Val Gly Glu Ile
Met Asn Ser
(2) INFORMATION FOR SEQ ID NO:8:
(i) SEQUENCE CHARACTERISTICS
(A) LENGTH: 22 amino acids
(B) TYPE: amino acid
(C) STRANDEDNESS:
(D) TOPOLOGY: linear
(ii) MOLECULE TYPE: peptide
(ix) FEATURE:
(A) NAME/KEY: Magainin III peptide.
(D) OTHER INFORMATION: amide- or carboxy- terminated
(x) PUBLICATION INFORMATION:
(A) AUTHOR: Zasloff, Michael
(C) JOURNAL: Proceedings of the National Academy
of Sciences
(D) VOLUME: 84
(F) PAGES: 5449-5453
(G) DATE: AUG - 1987
(H) DO~uM~NT NUMBER: US 4810777
(I) FILING DATE: 04-MAR-1987
(J) PUBLICATION DATE: 07-MAR-1989
SlJBSTITUTE S~I~ET
2125494
W093/11783 PCT/US92/1~27
-49-
(xi) SEQUENCE DESCRIPTION: SEQ ID NO:8
Gly Ile Gly Lys Phe Leu Hi 8 Ser Ala Lys
Lys Phe Gly Lys Ala Phe Val Gly Glu Ile
Met Asn
(2) INFORMATION FOR SEQ ID NO:9:
(i) SEQUENCE CHARACTERISTICS
(A) LENGTH: 22 amino acids
(B) TYPE: amino acid
(C) STRANDEDNESS:
(D) TOPOLOGY: linear
(ii) MOLECULE TYPE: peptide
(ix) FEATURE:
(A) NAME/KEY: maqainin peptide.
(D) OTHER INFORMATION: amide- or carboxy- terminated
(x) PUBLICATION INFORMATION:
(A) AUTHOR: Zasloff, Michael
(C) JOURNAL: Proceedinqs of the National Academy
of Sciences
(D) VOLUME: 84
(F) PAGES: 5449-5453
(G) DATE: AUG - 1987
(H) DOCUMENT NUMBER: US 4810777
(I) FILING DATE: 04-MAR-1987
(J) PUBLICATION DATE: 07-MAR-1989
SUBSTlTUTE S~E~T
W093/11783 2 1 2 5 4 ~ ~ PCT/US92/10427
-50-
(xi) SEQUENCE DESCRIPTION: SEQ ID NO:9
Ile Gly Lys Phe Leu His Ser Ala Lys Lys
Phe Gly Lys Ala Phe Val Gly Glu Ile Met -
Asn Ser
(2) INFORMATION FOR SEQ ID NO:10:
(i) SEQUENCE CHARACTERISTICS
(A) LENGTH: 21 amino acids
(B) TYPE: amino acid
(C) STRANDEDNESS:
(D) TOPOLOGY: linear
(ii) MOLECULE TYPE: peptide
(ix) FEATURE:
(A) NAME/KEY: magainin peptide.
(D) OTHER INFORMATION: amide- or carboxy- terminated
(x) PUBLICATION INFORMATION:
(A) AUTHOR: Zasloff, Michael
(C) JOURNAL: Proceedings of the National Academy
of Sciences
(D) VOLUME: 84
(F) PAGES: 5449-5453
(G) DATE: AUG - 1987
(H) DOCUMENT NUMBER: US 4810777
(I) FILING DATE: 04-MAR-1987
(J) PUBLICATION DATE: 07-MAR-1989
S~JBSTITUTE SH~ET
2125~
WO93/11783 PCT/US92/10427
-51-
(xi) SEQUENCE DESCRIPTION: SEQ ID NO:10
Gly Lys Phe Leu His Ser Ala Lys Lys Phe
5 10
Gly Lys Ala Phe Val Gly Glu Ile Met Asn
15 20
- Ser
(2) INFORMATION FOR SEQ ID NO:ll:
(i) SEQUENCE CHARACTERISTICS
(A) LENGTH: 20 amino acids
(B) TYPE: amino acid
(C) STRAN,-~EDNESS:
(D) TOPOLOGY: linear
(ii) MOLECULE TYPE: peptide
(ix) FEATURE:
(A) NAME/KEY: magainin peptide.
(D) OTHER INFORMATION: amide- or carboxy- terminated
(x) PUBLICATION INFORMATION:
(A) AUTHOR: Zasloff, Michael
(C) JOURNAL: Proceedings of the National Academy
of Sciences
(D) VOLUME: 84
(F) PAGES: 5449-5453
(G) DATE: AUG - 1987
(H) DOCUMENT NUMBER: US 4810777
(I) FILING DATE: 04-MAR-1987
(J) PUBLICATION DATE: 07-MAR-1989
SUBSTITUTE SH~ET
WO93/11783 ~ 9 4 -52- PCT/US92/1~27
(xi) SEQUENCE DESCRIPTION: SEQ ID NO:ll
Lys Phe Leu His Ser Ala Lys Lys Phe Gly
Lys Ala Phe Val Gly Glu Ile Met Asn Ser
(2) INFORMATION FOR SEQ ID NO:12:
(i) SEQUENCE CHARACTERISTICS
(A) LENGTH: 21 amino acids
(B) TYPE: amino acid
(C) STRANDEDNESS:
(D) TOPOLOGY: linear
(ii) MOLECULE TYPE: peptide
(ix) FEATURE:
(A) NAME/KEY: PGLa peptide.
(D) O-l~R INFORMATION: amide-terminated
(x) PUBLICATION INFORMATION:
(A) AUTHOR: Hoffman, et al.
(C) JOURNAL: EMBO J.
(D) VOLUME: 2
(F) PAGES: 711-714
(G) DATE: 1983
(A) AUTHOR: Andreu, et al.
(C) JOURNAL: Journal of Biochemistry
(D) VOLUME: 149
(F) PAGES: 531-535
(G) DATE: 1985
(A) AUTHOR: Gibson, et al.
(C) JOURNAL: J. Biol. Chem.
(D) VOLUME: 261
(F) PAGES: 5341-5349
SUBSTITUTE SHEET
W093~11783 2 1 2 5 4 ~ 4 PCT/US92/1~27
-53-
(G) DATE: 1986
(A) AUTHOR: Giovannini, et al.
(C) JOURNAL: Biochem J.
(D) VOLUME: 243
(F) PAGES: 113-120
(G) DATE: 1987
(xi) SEQUENCE DESCRIPTION: SEQ ID NO:12
Gly Met Ala Ser Lys Ala Gly Ala Ile Ala
Gly Lys Ile Ala Lys Val Ala Leu Lys Ala
Leu
(2) INFORMATION FOR SEQ ID NO:13:
(i) SEQUENCE CHARACTERISTICS
(A) LENGTH: 25 amino acids
(B) TYPE: amino acid
(C) STRANDEDNESS:
(D) TOPOLOGY: linear
(ii) MOLECULE TYPE: peptide
(ix) FEATURE:
(A) NAME/KEY: XPF peptide.
(x) PUBLICATION INFORMATION:
(A) AUTHOR: Hoffman, et al.l
(C) JOURNAL: EMBO J.
(D) VOLUME: 2
(F) PAGES: 711-714
(G~ DATE: 1983
(A) AUTHOR: Andreu, et al.
(C) JOURNAL: Journal of BiochemistrY
SlJBSTlTU TE SHEET
~12S4~4
WO93/11783 PCT/US92/10427
-54-
(D) VOLUME: 149
(F) PAGES: 531-535
(G) DATE: 1985
(A) AUTHOR: Gibson, et al.
(C) JOURNAL: J. Biol. Chem.
(D) VOLUME: 261
(F) PAGES: 5341-5349
(G) DATE: 1986
(A) AUTHOR: Giovannini, et al.
(C) JOURNAL: Biochem J.
(D) VOLUME: 243
(F) PAGES: 113-120
(G) DATE: 1987
(xi) SEQUENCE DESCRIPTION: SEQ ID NO:13
Gly Trp Ala Ser Lys Ile Gly Gln Thr Leu
Gly Lys Ile Ala Lys Val Gly Leu Lys Glu
Leu Ile Gln Pro Lys
(2) INFORMATION FOR SEQ ID NO:14:
(i) SEQUENCE CHARACTERISTICS
(A) LENGTH: 27 amino acids
(B) TYPE: amino acid
(C) STRANDEDNESS:
(D) TOPOLOGY: linear
(ii) MOLECULE TYPE: peptide
(ix) FEATURE:
(A) NAME/KEY: CPF peptide.
(D) OTHER INFORMATION: amide- or carboxy- terminated
SU~STITU I E Sl lEET
212~494
W O 93/11783 PC~r/US92/10427
-55-
(x) PUBLICATION INFORMATION:
(A) AUTHOR: Richter, K.
Egger, R.
Kreil
(C) JOURNAL: J. Biol. Chem.
(D) VOLUME: 261
(F) PAGES: 3676-3680
(G) DATE: 1986
(A) AUTHOR: Wakabayashi, T.
Kato, H.
Tachibaba, S.
(C) JOURNAL: Nucleic Acids Research
(D) VOLUME: 13
(F) PAGES: 1817-1828
(G) DATE: 1985
(A) AUTHOR: Gibson, B.W.
Poulter, L.
Williams, D.H.
Maggio, J.E.
(C) JOURNAL: J. Biol. Chem.
(D) VOLUME: 261
(F) PAGES: 5341-5349
(G) DATE: 1986
(H) DOCUMENT NUMBER: W090/04407
(I) FILING DATE: 16-OCT-1989
(J) PUBLICATION DATE: 03-MAY-l99O
(xi) SEQUENCE DESCRIPTION: SEQ ID NO:14
Gly Phe Gly Ser Phe Leu Gly Leu Ala Leu
Lys Ala Ala Leu Lys Ile Gly Ala Asn Ala
Leu Gly Gly Ala Pro Gln Gln
S~JBSTITUTE SHE~
2125~4
WOg3/11783 PCT/US92/10427
-56-
(2) INFORMATION FOR SEQ ID NO:15:
(i) SEQUENCE CHARACTERISTICS
(A) LENGTH: 27 amino acids
(B) TYPE: amino acid
(C) STRANDEDNESS:
(D) TOPOLOGY: linear
(ii) MOLECULE TYPE: peptide
(ix) FEATURE:
(A) NAME/KEY: CPF peptide.
(D) OTHER INFORMATION: amide- or carboxy- terminated
(x) PUBLICATION INFORMATION:
(A) AUTHOR: Richter, K
Egger, R.
Kreil
(C) JOURNAL: J. Biol. Chem.
(D) VOLUME: 261
(F) PAGES: 3676-3680
(G) DATE: 1986
(A) AUTHOR: Wakabayashi, T.
Kato, H.
Tachibaba, S.
(C) JOURNAL: Nucleic Acids Research
(D) VOLUME: 13
(F) PAGES: 1817-1828
(G) DATE: 1985
(A) AUTHOR: Gibson, B.W.
Poulter, L.
Williams, D.H.
- Maggio, J.E.
(C) JOURNAL: J. Biol. Chem.
(D) VOLUME: 261
SUBSTITUTE SH~ET
W093/11783 212 ~ 4 9 4 PCT/US92/10427
-57-
(F) -PAGES: 5341-5349
(G) DATE: 1986
(H) DOCUMENT NUMBER: WO90/04407
(I) FILING DATE: 16-OCT-1989
(J) PUBLICATION DATE: 03-MAY-l99O
(xi) SEQUENCE DESCRIPTION: SEQ ID NO:15
Gly Leu Ala Ser Phe Leu Gly Lys Ala Leu
Lys Ala Gly Leu Lys Ile Gly Ala His Leu
Leu Gly Gly Ala Pro Gln Gln
(2) INFORMATION FOR SEQ ID NO:16:
(i) SEQUENCE CHARACTERISTICS
(A) LENGTH: 27 amino acids
(B) TYPE: amino acid
(C) STRANDEDNESS:
(D) TOPOLOGY: linear
(ii) MOLECULE TYPE: peptide
(ix) FEATURE:
(A) NAME/KEY: CPF peptide.
(D) OTHER INFORMATION: amide- or carboxy- terminated
(x) PUBLICATION INFORMATION:
(A) AUTHOR: Richter, K.
Egger, R.
Kreil
(C) JOURNAL: J. Biol. Chem.
(D) VOLUME: 261
(F) PAGES: 3676-3680
SUBSTITUTE StlEET
212~434
WO93/11783 PCT/US92/10427
-58-
(G) DATE: 1986
(A) AUTHOR: Wakabayashi, T.
Kato, H.
Tachibaba, S.
(C) JOURNAL: Nucleic Acids Research
(D) VOLUME: 13
(F) PAGES: 1817-1828
(G) DATE: 1985
(A) AUTHOR: Gibson, B.W.
Poulter, L.
Williams, D.H.
Maggio, J.E.
(C) JOURNAL: J. Biol. Chem.
(D) VOLUME: 261
(F) PAGES: 5341-5349
(G) DATE: 1986
(H) DOCUMENT NUMBER: W090/04407
(I) FILING DATE: 16-OCT-1989
(J) PUBLICATION DATE: 03-MAY-1990
(xi) SEQUENCE DESCRIPTION: SEQ ID NO:16
Gly Leu Ala Ser Leu Leu Gly Lys Ala Leu
Lys Ala Gly Leu Lys Ile Gly Thr His Phe
Leu Gly Gly Ala Pro Gln Gln
(2) INFORMATION FOR SEQ ID NO:17:
(i) SEQUENCE CHARACTERISTICS
(A) LENGTH: 27 amino acids
(B) TYPE: amino, acid
(C) STRANDEDNESS:
(D) TOPOLOGY: linear
SlJBSTITUT~ S~
212~4~4
W093/11783 PCT/US92/1~27
-59-
(ii) MOLECULE TYPE: peptide
(ix) FEATURE:
(A) NAME/KEY: CPF peptide.
(D) OTHER INFORMATION: amide- or carboxy- terminated
(x) PUBLICATION INFORMATION:
(A) AUTHOR: Richter, K.
Egger, R.
Kreil
(C) JOURNAL: J. Biol. Chem.
(D) VOLUME: 261
(F) PAGES: 3676-3680
(G) DATE: 1986
(A) AUTHOR: Wakabayashi, T.
Kato, H.
Tachibaba, S.
(C) JOURNAL: Nucleic Acids Research
(D) VOLUME: 13
(F) PAGES: 1817-1828
(G) DATE: 1985
(A) AUTHOR: Gibson, B.W.
Poulter, L.
Williams, D.H.
Maggio, J.E.
(C) JOURNAL: J. Biol. Chem.
(D) VOLUME: 261
(F) PAGES: 5341-5349
(G) DATE: 1986
(H) DOCUMENT NUMBER: WO90/04407
(I) FILING DATE: 16-OCT-1989
(J) PUBLICATION DATE: 03-MAY-l99O
Sl lBSTITUTE SHFET
212~
W093/l1783 PCT/US92/10427
-60-
(xi) SEQUENCE DESCRIPTION: SEQ ID NO:17
Gly Leu Ala Ser Leu Leu Gly Lys Ala Leu
Lys Ala Thr Leu Lys Ile Gly Thr His Phe
Leu Gly Gly Ala Pro Gln Gln
(2) INFORMATION FOR SEQ ID NO:18:
(i) SEQUENCE CHARACTERISTICS
(A) LENGTH: 27 amino acids
(B) TYPE: amino acid
(C) STRANDEDNESS:
(D) TOPOLOGY: linear
(ii) MOLECULE TYPE: peptide
(ix) FEATURE:
(A) NAME/KEY: CPF peptide.
(D) OTHER INFORMATION: amide- or carboxy- terminated
(x) PUBLICATION INFORMATION:
(A) AUTHOR: Richter, K.
Egger, R.
Kreil
(C) JOURNAL: J. Biol. Chem.
(D) VOLUME: 261
(F) PAGES: 3676-3680
(G) DATE: 1986
(A) AUTHOR: Wakabayashi, T.
Kato, H.
. Tachibaba, S. . .
(C) JOURNAL: Nucleic Acids Research
(D) VOLUME: 13
SUBSTITUTE SH~ET
2125494
W093/11783 PCT/US92/10427
-61-
(F) PAGES: 1817-1828
(G) DATE: 1985
(A) AUTHOR: Gibson, B.W.
Poulter, L.
Williams, D.H.
Maggio, J.E.
(C) JOURNAL: J. Biol. Chem.
(D) VOLUME: 261
(F) PAGES: 5341-5349
(G) DATE: 1986
(H) DOCUMENT NUMBER: WO90/04407
(I) FILING DATE: 16-OCT-1989
(J) PUBLICATION DATE: 03-MAY-1990
(xi) SEQUENCE DESCRIPTION: SEQ ID NO:18
Gly Phe Ala Ser Phe Leu Gly Lys Ala Leu
Lys Ala Ala Leu Lys Ile Gly Ala Asn Met
20eu Gly Gly Thr Pro Gln Gln
2) INFORMATION FOR SEQ ID NO:l9:
(i) SEQUENCE CHARACTERISTICS
(A) LENGTH: 27 amino acids
(B) TYPE: amino acid
(C) STRANDEDNESS:
(D) TOPOLOGY: linear
(ii) MOLECULE TYPE: peptide
(ix) FEATURE:
(A) NAME/KEY: CPF peptide.
(D) OTHER INFORMATION: amide- or carboxy- terminated
SUBSTITUTE SH~ET
WO93/1178~ 1 2 5 ~ ~ ~ PCT/US92/10427
-62-
(x) PUBLICATION INFORMATION:
(A) AUTHOR: Richter, K.
Egger, R.
Kreil
(C) JOURNAL: J. Biol. Chem.
(D) VOLUME: 261
(F) PAGES: 3676-3680
(G) DATE: 1986
(A) AUTHOR: Wakabayashi, T.
Kato, H.
Tachibaba, S.
(C) JOURNAL: Nucleic Acids Research -
(D) VOLUME: 13
(F) PAGES: 1817-1828
(G) DATE: 1985
(A) AUTHOR: Gibson, B.W.
Poulter, L.
Williams, D.H.
Maggio, J.E.
(C) JOURNAL: J. Biol. Chem.
(D) VOLUME: 261
(F) PAGES: 5341-5349
(G) DATE: 1986
(H) DOCUMENT NUMBER: W090/04407
(I) FILING DATE: 16-OCT-1989
(J) PUBLICATION DATE: 03-MAY-1990
(xi) SEQUENCE DESCRIPTION: SEQ ID NO:l9
Gly Phe Gly Ser Phe Leu Gly Lys Ala Leu
Lys Ala Ala Leu Lys Ile Gly Ala Asn Ala
Leu Gly Gly Ala Pro Gln Gln
SVBSTITUTE SHFET
WO93/11783 2 1 2 5 ~ 9 4 PCT/US92/10427
- -63-
(2) INFORMATION FOR SEQ ID NO:20:
(i) SEQUENCE CHARACTERISTICS
(A) LENGTH: 27 amino acids
(B) TYPE: amino acid
(C) STRANDEDNESS:
(D) TOPOLOGY: linear
(ii) MOLECULE TYPE: peptide
(ix) FEATURE:
(A) NAME/KEY: CPF peptide.
(D) OTHER INFORMATION: amide- or carboxy- terminated
(x) PUBLICATION INFORMATION:
(A) AUTHOR: Richter, K.
Egger, R.
Kreil
(C) JOURNAL: J. Biol. Chem.
(D) VOLUME: 261
(F) PAGES: 3676-3680
(G) DATE: 1986
(A) AUTHOR: Wakabayashi, T.
Kato, H.
Tachibaba, S.
(C) JOURNAL: Nucleic Acids Research
(D) VOLUME: 13
(F) PAGES: 1817-1828
(G) DATE: 1985
(A) AUTHOR: Gibson, B.W.
Poulter, L.
Williams, D.H.
Maggio, J.E.
(C) JOURNAL: J. Biol. Chem.
(D) VOLUME: 261
SIJBSTITUTF SHFET
212~q~4
WO93/11783 PCT/US92/10427
-64-
(F) PAGES: 5341-5349
(G) DATE: 1986
(H) DOCUMENT NUMBER: W090/04407
(I) FILING DATE: 16-OCT-1989
(J) PUBLICATION DATE: 03-MAY-1990
(xi) SEQUENCE DESCRIPTION: SEQ ID NO:20
Gly Phe Gly Ser Phe Leu Gly Lys Ala Leu
Lys Ala Ala Leu Lys Ile Gly Ala Asn Ala
Leu Gly Gly Ser Pro Gln Gln
(2) INFORMATION FOR SEQ ID NO:21:
(i) SEQUENCE CHARACTERISTICS
(A) LENGTH: 27 amino acids
(B) TYPE: amino acid
(C) STRANDEDNESS:
(D) TOPOLOGY: linear
(ii) MOLECULE TYPE: peptide
(ix) FEATURE:
(A) NAME/KEY: CPF peptide.
(D) OTHER INFORMATION: amide- or carboxy- terminated
(x) PUBLICATION INFORMATION:
(A) AUTHOR: Richter, K.
Egger, R.
Kreil
(C) JOURNAL: J. Biol. Chem.
(D) VOLUME: 261
(F) PAGES: 3676-36~30
SUBSTITUTE SHEET
W093/11783 2 1 2 ~ 4 9 4 PCT/US92/10427
-65-
(G) DATE: 1986
(A) AUTHOR: Wakabayashi, T.
Kato, H.
Tachibaba, S.
(C) JOURNAL: Nucleic Acids Research
(D) VOLUME: 13
(F) PAGES: 1817-1828
(G) DATE: 1985
(A) AUTHOR: Gibson, B.W.
Poulter, L.
Williams, D.H.
Maggio, J.E.
(C) JOURNAL: J. Biol. Chem.
(D) VOLUME: 261
(F) PAGES: 5341-5349
(G) DATE: 1986
(H) DOCUMENT NUMBER: WO90/04407
(I) FILING DATE: 16-OCT-1989
(J) PUBLICATION DATE: 03-MAY-l99O
(xi) SEQUENCE DESCRIPTION: SEQ ID NO:21
Gly Phe Ala Ser Phe Leu Gly Lys Ala Leu
Lys Ala Ala Leu Lys Ile Gly Ala Asn Leu
Leu Gly Gly Thr Pro Gln Gln
(2) INFORMATION FOR SEQ ID NO:22:
(i) SEQUENCE CHARACTERISTICS
(A) LENGTH: 27 amino acids
(B) TYPE: amino acid
(C3 STRANDEDNESS:
(D) TOPOLOGY: linear
SUBSTtTlJTE S~
212S~494
W093/11783 PCT/US92/10427
-66-
(ii) MOLECULE TYPE: peptide
(ix) FEATURE:
(A) NAME/KEY: CPF peptide.
(D) OTHER INFORMATION: amide- or carboxy- terminated
(x) PUBLICATION INFORMATION:
(A) AUTHOR: Richter, K.
Egger, R.
Kreil
(C) JOURNAL: J. Biol. Chem.
(D) VOLUME: 261
(F) PAGES: 3676-3680
(G) DATE: 1986
(A) AUTHOR: Wakabayashi, T.
Kato, H.
Tachibaba, S.
(C) JOURNAL: Nucleic Acids Research
(D) VOLUME: 13
(F) PAGES: 1817-1828
(G) DATE: 1985
(A) AUTHOR: Gibson, B.W.
Poulter, L.
Williams, D.H.
Maggio, J.E.
(C) JOURNAL: J. Biol. Chem.
(D) VOLUME: 261
(F) PAGES: 5341-5349
(G) DATE: 1986
SUBSTITUTE SHEET
212S494
W093/11783 PCT/US92/10427
-67-
(xi) SEQUENCE DESCRIPTION: SEQ ID NO:22
Gly Phe Ala Ser Phe Leu Gly Lys Ala Leu
Lys Ala Ala Leu Lys Ile Gly Ala Asn Ala
lS 20
Leu Gly Gly Ala Pro Gln Gln
(2) INFORMATION FOR SEQ ID NO:23:
(i) SEQUENCE CHARACTERISTICS
(A) LENGTH: 27 amino acids
(B) TYPE: amino acid
(C) STRANDEDNESS:
(D) TOPOLOGY: linear
(ii) MOLECULE TYPE: peptide
(ix) FEATURE:
(A) NAME/KEY: CPF peptide.
(D) OTn~R INFORMATION: amide- or carboxy- terminated
(x) PUBLICATION INFORMATION:
(A) AUTHOR: Richter, K.
Egger, R.
Kreil
(C) JOURNAL: J. Biol. Chem.
(D) VOLUME: 261
(F) PAGES: 3676-3680
(G) DATE: 1986
(A) AUTHOR: Wakabayashi, T.
Kato, H.
Tachibaba, S.
(C) JOURNAL: Nucleic Acids Research
(D) VOLUME: 13
SUBSTITUTE SH~ET
212~494
WO93/11783 PCT/US92/10427
-68-
(F) PAGES: 1817-1828
(G) DATE: 1985
(A) AUTHOR: Gibson, B.W.
Poulter, L.
Williams, D.H.
Maggio, J.E.
(C) JOURNAL: J. Biol. Chem.
(D) VOLUME: 261
(F) PAGES: 5341-5349
(G) DATE: 1986
(xi) SEQUENCE DESCRIPTION: SEQ ID NO:23 -
Gly Phe Ala Ser Phe Leu Gly Lys Ala Leu
Lys Ala Ala Leu Lys Ile Gly Ala Asn Met
Leu Gly Gly Ala Pro Gln Gln
(2) INFORMATION FOR SEQ ID NO:24:
(i) SEQUENCE CHARACTERISTICS
(A) LENGTH: 27 amino acids
(B) TYPE: amino acid
(C) STRANDEDNESS:
(D) TOPOLOGY: linear
(ii) MOLECULE TYPE: peptide
(ix) FEATURE:
(A) NAME/KEY: CPF peptide.
(D) OTHER INFORMATION: amide- or carboxy- terminated
.
(x) PUBLICATION INFORMATION:
(A) AUTHOR: Richter, K.
SUBSTITlJTE SHEET
WO93/11783 2 1 ~ 5 4 9 ~ PCT/US92/1~27
-69-
Egger, R.
Kreil
(C) JOURNAL: J. Biol. Chem.
(D) VOLUME: 261
(F) PAGES: 3676-3680
(G) DATE: 1986
(A) AUTHOR: Wakabayashi, T.
Kato, H.
Tachibaba, S.
(C) JOURNAL: Nucleic Acids Research
(D) VOLUME: 13
(F) PAGES: 1817-1828
(G) DATE: 1985
(A) AUTHOR: Gibson, B.W.
Poulter, L.
Williams, D.H.
Maggio, J.E.
(C) JOURNAL: J. Biol. Chem.
(D) VOLUME: 261
(F) PAGES: 5341-5349
(G) DATE: 1986
(xi) SEQUENCE DESCRIPTION: SEQ ID NO:24
Gly Phe Gly Ser Phe Leu Gly Lys Ala Leu
Lys Ala Ala Leu Lys Ile Gly Ala Asn Ala
Leu Gly Gly Ser Leu Gln Gln
(2) INFORMATION FOR SEQ ID NO:25:
(i) SEQUENCE CHARACTERISTICS
(A) LENGTH: 27 amino acids
(B) TYPE: amino acid
8U8STITUTE SHFE~
212~4
W093/11783 PCT/US92/10427
-70-
(C) STRANDEDNESS:
(D) TOPOLOGY: linear
(ii) MOLECULE TYPE: peptide
(ix) FEATURE:
(A) NAME/KEY: CPF peptide.
(D) OTHER INFORMATION: amide- or carboxy- terminated
(x) PUBLICATION INFORMATION:
(A) AUTHOR: Richter, K.
Egger, R.
Kreil
(C) JOURNAL: J. Biol. Chem.
(D) VOLUME: 261
(F) PAGES: 3676-3680
(G) DATE: 1986
(A) AUTHOR: Wakabayashi, T.
Kato, H.
Tachibaba, S.
(C) JOURNAL: Nucleic Acids Research
(D) VOLUME: 13
(F) PAGES: 1817-1828
(G) DATE: 1985
(A) AUTHOR: Gibson, B.W.
Poulter, L.
Williams, D.H.
Maggio, J.E.
(C) JOURNAL: J. Biol. Chem.
(D) VOLUME: 261
(F) PAGES: 5341-5349
(G) DATE: 1986
8UBSTITUTE SHEET
WO93/11783 2 1 2 ~ 4 g I PCT/US92/10427
-71-
I
(xi) ~QU~-NCE DESCRIPTION: SEQ ID NO:25
Gly Phe Gly Ser Phe Leu Gly Lys Ala Leu
Lys Ala Gly Leu Lys Ile Gly Thr Asn Phe
Leu Gly Gly Ala Pro Gln Gln
(2) INFORMATION FOR SEQ ID NO:26:
(i) SEQUENCE CHARACTERISTICS
(A) LENGTH: 27 amino acids
(B) m E: amino acid
(C) STRANDEDNESS:
(D) TOPOLOGY: linear
(ii) MOLECULE TYPE: peptide
(ix) FEATURE:
(A) NAME/KEY: CPF peptide.
(D) OTHER INFORMATION: amide- or carboxy- terminated
(x) PUBLICATION INFORMATION:
(A) AUTHOR: Richter, K
Egger, R.
Kreil
(C) JOURNAL: J. Biol. Chem.
(D) VOLUME: 261
(F) PAGES: 3676-3680
(G) DATE: 1986
(A) AUTHOR: Wakabayashi, T.
Kato, H.
Tachibaba, S. . .
(C) JOURNAL: Nucleic Acids Research
(D) VOLUME: l3
SUBSTITUTF SH~ET
21254~4
W093/11783 PCT/US92/1~27
-72-
(F) PAGES: 1817-1828
(G) DATE: 1985
(A) AUTHOR: Gibson, B.W.
Poulter, L.
Williams, D.H.
Maggio, J.E.
(C) JOURNAL: J. Biol. Chem.
(D) VOLUME: 261
(F) PAGES: 5341-5349
(G) DATE: 1986
(xi) SEQUENCE DESCRIPTION: SEQ ID NO:26:
Gly Leu Ala Ser Leu Leu Gly Lys Ala Leu
Lys Ala Ala Leu Lys Ile Gly Ala Asn Ala
Leu Gly Gly Ser Pro Gln Gln
(2) INFORMATION FOR SEQ ID NO:27:
(i) SEQUENCE CHARACTERISTICS
(A) LENGTH: 21 amino acids
(B) TYPE: amino acid
(C) STRANDEDNESS:
(D) TOPOLOGY: linear
(ii) MOLECULE TYPE: peptide
(ix) FEATURE:
(D) OTHER INFORMATION: amide-terminated
(xi) SEQUENCE DESCRIPTION: SEQ ID NO:27:
Lys Ile Ala Gly Lys Ile Ala Lys Ile Ala
SUBSTITlJT~ SH~T
W093/11783 2 1 2 5 4 g ~ PCT/US92/1~27
Gly Lys Ile Ala Lys Ile Ala Gly Lys I le
Ala
(2) INFORMATION FOR SEQ ID NO:28:
(i) SEQUENCE CHARACTERISTICS:
(A) LENGTH: 21 amino acids
(B) TYPE: amino acid
(C) STRANDEDNESS:
(D) TOPOLOGY: linear
(ii) MOLECULE TYPE: peptide
(ix) FEATURE:
(D) OTHER INFORMATION: amide-terminated
(xi) SEQUENCE DESCRIPTION: SEQ ID NO:28:
Lys Ile Ala Lys Ile Ala Gly Lys Ile Ala
Lys Ile Ala Gly Lys Ile Ala Lys Ile Ala
Gly
(2) INFORMATION FOR SEQ ID NO:29:
(i) SEQUENCE CHARACTERISTICS:
(A) LENGTH: 21 amino acids
(B) TYPE: amino acid
(C) STRANDEDNESS:
(D) TOPOLOGY: linear
(ii) MOLECULE TYPE: peptide
(ix) FEATURE:
(D) OTHER INFORMATION: amide-ter.ninated
SUBSTITlJTE SHEET
2125~4
WO93/11783 PCT/US92/10427
-74-
(xi) SEQUENCE DESCRIPTION: SEQ ID NO:29:
Lys Ile Ala Gly Lys Ile Gly Lys Ile Ala
Gly Lys Ile Gly Lys Ile Ala Gly Lys Ile
Gly
(2) INFORMATION FOR SEQ ID NO:30:
(i) SEQUENCE CHARACTERISTICS
(A) LENGTH: 21 amino acids
(B) TYPE: amino acid
(C) STRANDEDNESS:
(D) TOPOLOGY: linear
(ii) MOLECULE TYPE: peptide
(ix) FEATURE:
(D) OTHER INFORMATION: amide-terminated
(xi) SEQUENCE DESCRIPTION: SEQ ID NO:30:
Lys Leu Ala Gly Lys Leu Ala Lys Leu Ala
lOly Lys Leu Ala Lys Leu Ala Gly Lys Leu
20la
2) INFORMATION FOR SEQ ID NO:3l:
(i) SEQUENCE CHARACTERISTICS
(A) LENGTH: 2l amino acids
(B) TYPE: amino acid
(C) STRANDEDNESS:
(D) TOPOLOGY: linear
(ii) MOLECULE TYPE: peptide
8~JBSTITUTE SHEET
WO93/11783 2 1 2 5 ~ 9 ~ PCT/US92/10427
-75-
(ix) FEATURE:
(D) OTHER INFORMATION: amide-terminated
(xi) SEQUENCE DESCRIPTION: SEQ ID NO:31:
Lys Phe Ala Gly Lys Phe Ala Lys Phe Ala
Gly Lys Phe Ala Lys Phe Ala Gly Lys Phe
Ala
(2) INFORMATION FOR SEQ ID NO:32:
(i) SEQUENCE CHARACTERISTICS
(A) LENGTH: 21 amino acids
(B) TYPE: amino acid
(C) STRANDEDNESS:
(D) TOPOLOGY: linear
(ii) MOLECULE TYPE: peptide
(ix) FEATURE:
(D) OTHER INFORMATION: amide-terminated
(xi) SEQUENCE DESCRIPTION: SEQ ID NO:32:
Lys Ala Leu Ser Lys Ala Leu Lys Ala Leu
Ser Lys Ala Leu Lys Ala Leu Ser Lys Ala
Leu
(2) INFORMATION FOR SEQ ID NO:33:
(i) SEQUENCE CHARACTERISTICS
(A) LENGTH: 21 amino acids .
(B) TYPE: amino acid
(C) STRANDEDNESS:
SUBSTITIJTE SHFET
2125~4
WO93/11783 PCT/US92/10427
-76-
(D) TOPOLOGY: linear
~ii) MOLECULE TYPE: peptide
(ix) FEATURE:
(D) OTHER INFORMATION: amide-terminated
(xi) SEQUENCE DESCRIPTION: SEQ ID NO:33:
Lys Leu Leu Lys Ala Leu Gly Lys Leu Leu
Lys Ala Leu Gly Lys Leu Leu Lys Ala Leu
Gly
(2) INFORMATION FOR SEQ ID NO:34:
(i) SEQUENCE CHARACTERISTICS
(A) LENGTH: 2l amino acids
(B) TYPE: amino acid
(C) STRANDEDNESS:
(D) TOPOLOGY: linear
(ii) MOLECULE TYPE: peptide
(ix) FEATURE:
(D) OTHER INFORMATION: amide-terminated
(xi) SEQUENCE DESCRIPTION: SEQ ID NO:34:
Lys Ala Ile Gly Lys Ala Ile Lys Ala Ile
Gly Lys Ala Ile Lys Ala Ile Gly Lys Ala
Ile
(2) INFORMATION FOR SEQ ID NO:35
SUBSTITUTE Sl IFET
WO93/l1783 2 1 2 5 4 9 4 PCT/US92/10427
-77-
(i) SEQUENCE CHARACTERISTICS
(A) LENGTH: 21 amino acids
(B) TYPE: amino acid
(C) STRANDEDNESS:
(D) TOPOLOGY: linear
(ii) MOLECULE TYPE: peptide
(ix) FEATURE:
(D) OTHER INFORMATION: amide-terminated
(xi) SEQUENCE DESCRIPTION: SEQ ID NO:35:
Gly Ile Ala Lys Ile Ala Lys Gly Ile Ala
Lys Ile Ala Lys Gly Ile Ala Lys Ile Ala
Lys
(2) INFORMATION FOR SEQ ID NO:36:
(i) SEQUENCE CHARACTERISTICS
(A) LENGTH: 21 amino acids
(B) TYPE: amino acid
(C) STRANDEDNESS:
(D) TOPOLOGY: linear
(ii) MOLECULE TYPE: peptide
(ix) FEATURE:
(D) OTHER INFORMATION: amide-terminated
(xi) SEQUENCE DESCRIPTION: SEQ ID NO:36:
Lys Ile Ala Lys ~le Phe Gly Lys Ile Ala
SUBSTITUT~ SH~ET
2125~734
WO93/11783 PCT/US92/10427
-78-
Lys Ile Phe Gly Lys Ile Ala Lys Ile Phe
20ly2) INFORMATION FOR SEQ ID NO:37:
(i) SEQUENCE CHARACTERISTICS
(A) LENGTH: 21 amino acids
(B) TYPE: amino acid
(C) STRANDEDNESS:
(D) TOPOLOGY: linear
(ii) MOLECULE TYPE: peptide
(ix) FEATURE:
(D) OTHER INFORMATION: amide-terminated
(xi) SEQUENCE DESCRIPTION: SEQ ID NO:37:ly Ile Ala Arg Ile Ala Lys Gly Ile Ala l0rg Ile Ala Lys Gly Ile Ala Arg Ile Ala
20ys
2) INFORMATION FOR SEQ ID NO:38:
(i) SEQUENCE CHARACTERISTICS
(A) LENGTH: 21 amino acids
(B) TYPE: amino acid
(C) STRANDEDNESS:
(D) TOPOLOGY: linear
(ii) MOLECULE TYPE: peptide
(ix) FEATURE:
(D) OTHER INFORMATION: amide-terminated
SUBSTITIJTE S~ET
WO93/11783 2 1 2 5 ~ 9 1 PCT/US92/10427
-79-
(xi) SEQUENCE DESCRIPTION: SEQ ID NO:38:ys Phe Ala Arg Ile Ala Gly Lys Phe Ala 10rg Ile Ala Gly Lys Phe Ala Arg Ile Ala
20ly2) INFORMATION FOR SEQ ID NO:39:
(i) SEQUENCE CHARACTERISTICS
(A) LENGTH: 21 amino acids
(B) TYPE: amino acid
(C) STRANDEDNESS:
(D) TOPOLOGY: linear
(ii) MOLECULE TYPE: peptide
(ix) FEATURE:
(D) OTHER INFORMATION: amide-terminated
(xi) SEQUENCE DESCRIPTION: SEQ ID NO:39:ly Phe Ala Lys Ile Ala Lys Gly Phe Ala 10ys Ile Ala Lys Gly Phe Ala Lys Ile Ala
20ys
2) INFORMATION FOR SEQ ID NO:40:
(i) SEQUENCE CHARACTERISTICS
(A) LENGTH: 21 amino acids
(B) TYPE: amino acid
(C) STRANDEDNESS:
(D) TOPOLOGY: linear
(ii) MOLECULE TYPE: peptide
SUBSTITUTE Sl l~'E~
212~94
WO93/11783 PCT/US92/10427
-80-
(ix) FEATURE:
(D) OTHER INFORMATION: amide-terminated, Xaa is
ornithine.
(xi) SEQUENCE DESCRIPTION: SEQ ID NO:40:
Lys Ile Ala Gly Xaa Ile Ala Lys Ile Ala
Gly Xaa Ile Ala Lys Ile Ala Gly Xaa Ile
Ala
(2) INFORMATION FOR SEQ ID NO:41:
(i) SEQUENCE CHARACTERISTICS
(A) LENGTH: 21 amino acids
(B) TYPE: amino acid
(C) STRANDEDNESS:
(D) TOPOLOGY: linear
(ii) MOLECULE TYPE: peptide
(ix) FEATURE:
(D) OTHER INFORMATION: amide-terminated
(xi) SEQUENCE DESCRIPTION: SEQ ID NO:41:
Lys Ile Ala Arg Ile Ala Gly Lys Ile Ala
Arg Ile Ala Gly Lys Ile Ala Arg Ile Ala
Gly
(2) INFORMATION FOR SEQ ID NO:42:
Q~:NCE CHARACTERISTICS
(A) LENGTH: 21 amino acids
(B)- TYPE: amino acid
(C) STRANDEDNESS:
(D) TOPOLOGY: linear
SUBSTITUT~ SHFET
WO93/11783 ~ 1 2 5 4 ~ 4 PCT/US92/10427
-81-
(ii) MOLECULE TYPE: peptide
(ix) FEATURE:
(D) OTHER INFORMATION: amide-terminated, Xaa is
ornithine.
(xi) SEQUENCE DESCRIPTION: SEQ ID NO:42:
Xaa Ile Ala Gly Lys Ile Ala Xaa Ile Ala
Gly Lys Ile Ala Xaa Ile Ala Gly Lys Ile
Ala
(2) INFORMATION FOR SEQ ID NO:43:
(i) SEQUENCE CHARACTERISTICS
(A) LENGTH: 21 amino acids
(B) TYPE: amino acid
(C) STRANDEDNESS:
(D) TOPOLOGY: linear
(ii) MOLECULE TYPE: peptide
(ix) FEATURE:
(D) OTHER INFORMATION: amide-terminated
(xi) SEQUENCE DESCRIPTION: SEQ ID NO:43:
Gly Ile Ala Arg Ile Phe Lys Gly Ile Ala
Arg Ile Phe Lys Gly Ile Ala Arg Ile Phe
Lys
(2) INFORMATION FOR SEQ ID NO:44: . .
(i) SEQUENCE CHARACTERISTICS
(A) LENGTH: 21 amino acids
SUBSTITUT~ SI~ET
212~494
WO93/11783 PCT/US92/10427
-82-
(B) TYPE: amino acid
(C) STRANDEDNESS:
(D) TOPOLOGY: linear
(ii) MOLECULE TYPE: peptide
(ix) FEATURE:
(D) OTHER INFORMATION: amide-terminated, Xaa is
norleucine
(xi) SEQUENCE DESCRIPTION: SEQ ID NO:44:
Lys Xaa Ala Gly Lys Xaa Ala Lys Xaa Ala
Gly Lys Xaa Ala Lys Xaa Ala Gly Lys Xaa
Ala
(2) INFORMATION FOR SEQ ID NO:45:
(i) SEQUENCE CHARACTERISTICS
(A) LENGTH: 21 amino acids
(B) TYPE: amino acid
(C) STRANDEDNESS:
(D) TOPOLOGY: linear
(ii) MOLECULE TYPE: peptide
(ix) FEATURE:
(D) OTHER INFORMATION: amide-terminated, Xaa is
norleucine
(xi) SEQUENCE DESCRIPTION: SEQ ID NO:45:
Lys Xaa Ala Gly Lys Ile Ala Lys Xaa Ala
S~BSTITUTE ~ET
WO93/11783 2 1 2 5 4 ~ 4 PCT/US92/10427
-83-
Gly Lys Ile Ala Lys Xaa Ala Gly Lys Ile
Ala
(2) INFORMATION FOR SEQ ID NO:46:
(i) SEQUENCE CHARACTERISTICS
(A) LENGTH: 21 amino acids
(B) TYPE: amino acid
(C) STRANDEDNESS:
(D) TOPOLOGY: linear
(ii) MOLECULE TYPE: peptide
(ix) FEATURE:
(D) OTHER INFORMATION: amide-terminated, Xaa is
norleucine
(xi) SEQUENCE DESCRIPTION: SEQ ID NO:46:
Lys Ile Ala Gly Lys Xaa Ala Lys Ile Ala
S 10
Gly Lys Xaa Ala Lys Ile Ala Gly Lys Xaa
Ala
(2) INFORMATION FOR SEQ ID NO:47:
(i) SEQUENCE CHARACTERISTICS
(A) LENGTH: 21 amino acids
(B) TYPE: amino acid
(C) STRANDEDNESS:
(D) TOPOLOGY: linear
(ii) MOLECULE TYPE: peptide
(ix) FEATURE:
SUBSTITUTE ~ ET
2125ll~4
WO93/11783 PCT/US92/10427
-84-
(D) OTHER INFORMATION: amide-terminated, Xaa is
norvaline
(xi) SEQUENCE DESCRIPTION: SEQ ID NO:47:
Lys Xaa Ala Gly Lys Xaa Ala Lys Xaa Ala
Gly Lys Xaa Ala Lys Xaa Ala Gly Lys Xaa
Ala
(2) INFORMATION FOR SEQ ID NO:48:
(i) SEQUENCE CHARACTERISTICS
(A) LENGTH: 2l amino acids
(B) TYPE: amino acid
(C) STRANDEDNESS:
(D) TOPOLOGY: linear
(ii) MOLECULE TYPE: peptide
(ix) FEATURE:
(D) OTHER INFO~MATION: amide-terminated, Xaa is
norvaline
(xi) SEQUENCE DESCRIPTION: SEQ ID NO:48:
Lys Xaa Ala Gly Lys Ile Ala Lys Xaa Ala
Gly Lys Ile Ala Lys Xaa Ala Gly Lys Xaa
Ala
(2) INFORMATION FOR SEQ ID NO:49:
(i) SEQUENCE CHARACTERISTICS
(A) LENGIH: 21 amino acids
(B) TYPE: amino acid
(C) STRANDEDNESS:
~UE~STITUTE SHEET
W093/11783 2 1 % ~ 4 ~ 1 PCT/US92/10427
-85-
(D) TOPOLOGY: linear
(ii) MOLECULE TYPE: peptide
(ix) FEATURE:
(D) OTHER INFORMATION: amide-terminated
(xi) SEQUENCE DESCRIPTION: SEQ ID NO:49:ys Leu Leu Ser Lys Leu Gly Lys Leu Leu 10er Lys Leu Gly Lys Leu Leu Ser Lys Leu
20ly2) INFORMATION FOR SEQ ID NO:50:
(i) SEQUENCE CHARACTERISTICS
(A) LENGTH: 21 amino acids
(B) TYPE: amino acid
(C) STRANDEDNESS:
(D) TOPOLOGY: linear
(ii) MOLECULE TYPE: peptide
(ix) FEATURE:
(D) OTHER INFORMATION: amide-terminated
(xi) SEQUENCE DESCRIPTION: SEQ ID NO:50:ys Leu Leu Ser Lys Phe Gly Lys Leu Leu 10er Lys Phe Gly Lys Leu Leu Ser Lys Phe
20ly
(2) INFORMATION FOR SEQ ID NO:51
(i) SEQUENCE CHARACTERISTICS
SUBSTITUTE SHEET
21~S4~
W093/11783 PCT/US92/10427
-86-
(A) LENGTH: 2l amino acids
(B) TYPE: amino acid
(C) STRANDEDNESS:
(D) TOPOLOGY: linear
(ii) MOLECULE TYPE: peptide
(ix) FEATURE:
(D) OTHER INFORMATION: amide-terminated, Xaa is
norvaline
(xi) SEQUENCE DESCRIPTION: SEQ ID NO:5l:
Lys Ile Ala Gly Lys Xaa Ala Lys Ile Ala
lOly Lys Xaa Ala Lys Ile Ala Gly Lys Xaa
Ala2) INFORMATION FOR SEQ ID NO:52:
(i) S~:Q~:NCE CHARACTERISTICS
(A) LENGTH: 21 amino acids
(B) TYPE: amino acid
(C) STRANDEDNESS:
(D) TOPOLOGY: linear
(ii) MOLECULE TYPE: peptide
(ix) FEATURE:
(D) OTHER INFORMATION: amide-terminated
(xi) SEQUENCE DESCRIPTION: SEQ ID NO:52:
His Ile Ala Gly His Ile Ala His Ile Ala
. lO
SUBSTITU~E SI~EET
WO93/11783 2 1 2 ~ 4 9 4 PCT/US92/10427
-87-
Gly His Ile Ala His Ile Ala Gly His Ile
Ala
(2) INFORMATION FOR SEQ ID NO:53:
(i) SEQUENCE CHARACTERISTICS
(A) LENGTH: 21 amino acids
(B) TYPE: amino acid
(C) STRANDEDNESS:
(D) TOPOLOGY: linear
(ii) MOLECULE TYPE: peptide
(ix) FEATURE:
(D) OTHER INFORMATION: amide-terminated
(xi) SEQUENCE DESCRIPTION: SEQ ID NO:53:
Ala Gly Lys Ile Ala Lys Ile Ala Gly Lys
Ile Ala Lys Ile Ala Gly Lys Ile Ala Lys
Ile
(2) INFORMATION FOR SEQ ID NO:54:
(i) SEQUENCE CHARACTERISTICS
(A) LENGTH: 2l amino acids
(B) TYPE: amino acid
(C) STRANDEDNESS:
(D) TOPOLOGY: linear
(ii) MOLECULE TYPE: peptide
(ix) FEATURE:
(D) OTHER INFORMATION: amide-terminated
SUBSTITUTE SHEET
212S~
W093/11783 PCT/US92/10427
-88-
(xi) SEQUENCE DESCRIPTION: SEQ ID NO:54:le Ala Lys Ile Ala Gly Lys Ile Ala Lys lOle Ala Gly Lys Ile Ala Lys Ile Ala Gly
20ys2) INFORMATION FOR SEQ ID NO:55:
(i) SEQUENCE CHARACTERISTICS
(A) LENGTH: 21 amino acids
(B) TYPE: amino acid
(C) STRANDEDNESS:
(D) TOPOLOGY: linear
(ii) MOLECULE TYPE: peptide
(ix) FEATURE:
(D) OTHER INFORMATION: amide-terminated
(xi) SEQUENCE DESCRIPTION: SEQ ID NO:55:ys Ile Ala Gly Arg Ile Ala Lys Ile Ala lOly Arg Ile Ala Lys Ile Ala Gly Arg Ile
20la2) INFORMATION FOR SEQ ID NO:56:
(i) SEQUENCE CHARACTERISTICS
(A) LENGTH: 21 amino acids
(B) TYPE: amino acid
(C) STRANDEDNESS:
(D) TOPOLOGY: linear
(ii) MOLECULE TYPE: peptide
(ix) FEATURE:
SUBSTITUT~ SHErT
W093/11783 2 1 2 5 ~ 9 ~ PCT/US92/10427
-89
(D) OTHER INFORMATION: amide-terminated
(xi) SEQUENCE DESCRIPTION: SEQ ID NO:56:
Arg Ile Ala Gly Arg Ile Ala Arg Ile Ala
Gly Arg Ile Ala Arg Ile Ala Gly Arg Ile
Ala
(2) INFORMATION FOR SEQ ID NO:57:
(i) SEQUENCE CHARACTERISTICS
(A) LENGTH: 2l amino acids
(B) TYPE: amino acid
(C) STRANDEDNESS:
(D) TOPOLOGY: linear
(ii) MOLECULE TYPE: peptide
(ix) FEATURE:
(D) OTHER INFORMATION: amide-terminated
(xi) SEQUENCE DESCRIPTION: SEQ ID NO:57:
Lys Val Ala Gly Lys Ile Ala Lys Val Ala
Gly Lys Ile Ala Lys Val Ala Gly Lys Ile
Ala
(2) INFORMATION FOR SEQ ID NO:58:
(i) SEQUENCE CHARACTERISTICS
(A) LENGTH: 21 amino acids
(B) TYPE: amino acid
(C) STRANDEDNESS:
. (D) TOPOLOGY: linear
(ii) MOLECULE TYPE: peptide
SIJBSTITUTE Sl lFET
212S49~
WO93/11783 PCT/US92/1~27
-90-
(ix) FEATURE:
(D) OTHER INFORMATION: amide-terminated
(xi) SEQUENCE DESCRIPTION: SEQ ID NO:58:
Lys Ile Ala Gly Lys Val Ala Lys Ile Ala
Gly Lys Val Ala Lys Ile Ala Gly Lys Val
Ala
(2) INFORMATION FOR SEQ ID NO:59:
(i) SEQUENCE CHARACTERISTICS
(A) LENGTH: 21 amino acids
(B) TYPE: amino acid
(C) STRANDEDNESS:
(D) TOPOLOGY: linear
(ii) MOLECULE TYPE: peptide
(ix) FEATURE:
(D) OTHER INFORMATION: amide-terminated
(xi) SEQUENCE DESCRIPTION: SEQ ID NO:59:
Ala Lys Ile Ala Gly Lys Ile Ala Lys Ile
Ala Gly Lys Ile Aia Lys Ile Ala Gly Lys
Ile
(2) INFORMATION FOR SEQ ID NO:60:
~~NCE CHARACTERISTICS
- (A~ LENGTH: 21 amino acids
(B) TYPE: amino acid
(C) STRANDEDNESS:
SUBSTITUTE SHEET
212549~
W093/11783 PC. ~92/1~27
-91 -
(D) TOPOLOGY: linear
(ii) MOLECULE TYPE pepti~e
(ix) FEATURE:
(D) OTHER INFORMATION: amide-terminated, Xaa is
ornithine
(xi) SEQUENCE DESCRIPTION: SEQ ID NO:60:
Xaa Ile Ala Gly Xaa Ile Ala Xaa Ile Ala
Gly Xaa Ile Ala Xaa Ile Ala Gly Xaa Ile
Ala
(2) INFORMATION FOR SEQ ID NO:6l:
(i) SEQUENCE CHARACTERISTICS
(A) LENGTH: 21 amino acids
(B) TYPE: amino acid
(C) STRANDEDNESS:
(D) TOPOLOGY: linear
(ii) MOLECULE TYPE: peptide
(ix) FEATURE:
(D) OTHER INFORMATION: amide-terminated
(xi) SEQUENCE DESCRIPTION: SEQ ID NO:61:
Lys Phe Ala Gly Lys Ile Ala Lys Phe Ala
Gly Lys Ile Ala Lys Phe Ala Gly Lys Ile
Ala
SUBSTITVTE Sl I~E~
,~!o192 ~3 4 9 ~
3/11783 PCT/US92/10427
-92-
(2) INFORMATION FOR SEQ ID NO:62:
(i) SEQUENCE CHARACTERISTICS
(A) LENGTH: 21 amino acids
(B) TYPE: amino acid
(C) STRANDEDNESS:
(D) TOPOLOGY: linear
(ii) MOLECULE TYPE: peptide
(ix) FEATURE:
(D) OTHER INFORMATION: amide-terminated
(xi) SEQUENCE DESCRIPTION: SEQ ID NO:62:
Lys Ile Ala Gly Lys Phe Ala Lys Ile Ala
Gly Lys Phe Ala Lys Ile Ala Gly Lys Phe
Ala
(2) INFORMATION FOR SEQ ID NO:63:
(i) SEQUENCE CHARACTERISTICS
(A) LENGTH: 21 amino acids
(B) TYPE: amino acid
(C) STRANDEDNESS:
(D) TOPOLOGY: linear
(ii) MOLECULE TYPE: peptide
(ix) FEATURE:
(D) OTHER INFORMATION: amide-terminated, Xaa is
cyclohexylalanine
(xi) SEQUENCE DESCRIPTION: SEQ ID NO:63:
Lys Xaa Ala Gly Lys Ile Ala Lys Xaa Ala
SUBSTITUTE S~ET
WO93/11783 2 1 2 ~ ~ 9 4 PCT/US92/10427
-93-
Gly Lys Ile Ala Lys Xaa Ala Gly Lys Ile
Ala
(2) INFORMATION FOR SEQ ID NO:64:
(i) SEQUENCE CHARACTERISTICS
(A) LENGTH: 21 amino acids
(B) TYPE: amino acid
(C) STRANDEDNESS:
(D) TOPOLOGY: linear
(ii) MOLECULE TYPE: peptide
(ix) FEATURE:
(D) OTHER INFORMATION: amide-terminated, Xaa is
norleucine
(xi) SEQUENCE DESCRIPTION: SEQ ID NO:64:
Lys Xaa Ala Lys Ile Ala Gly Lys Xaa Ala
Lys Ile Ala Gly Lys Xaa Ala Lys Ile Ala
Gly
(2) INFORMATION FOR SEQ ID NO:65:
(i) SEQUENCE CHARACTERISTICS:
(A) LENGTH: 21 amino acids
(B) TYPE: amino acid
(C) STRANDEDNESS:
(D) TOPOLOGY: linear
(ii) MOLECULE TYPE: peptide
SUBS~ITI~E SHEET
212S4~
W093/11783 PCT/US92/10427
-94-
(ix) FEATURE:
(D) OTHER INFORMATION: amide-terminated
(xi) SEQUENCE DESCRIPTION: SEQ ID NO:65:rg Ile Ala Gly Lys Ile Ala Arg Ile Ala lO:
Gly Lys Ile Ala Arg Ile Ala Gly Lys Ile
Ala
2) INFORMATION FOR SEQ ID NO:66:
(i) SEQUENCE CHARACTERISTICS:
(A) LENGTH: 2l amino acids
(B) TYPE: amino acid
(C) STRANDEDNESS:
(D) TOPOLOGY: linear
(ii) MOLECULE TYPE: peptide
(ix) FEATURE:
(D) OTHER INFORMATION: amide-terminated, Xaa is
homoarginine
(xi) SEQUENCE DESCRIPTION: SEQ ID NO:66:
Xaa Ile Ala Gly Xaa Ile Ala Xaa Ile Ala
Gly Xaa Ile Ala Xaa Ile Ala Gly Xaa Ile
Ala
(2) INFORMATION FOR SEQ ID NO:67:
(i3 SEQUENCE CHARACTERISTICS:
(A) LENGTH: 2l amino acids
(B) TYPE: amino acid
SU~STITUTE SH~ET
WO93/11783 2 1 2 5 4 9 ~ PCT/US92/10427
-95-
(C) STRANDEDNESS:
(D) TOPOLOGY: linear
(ii) MOLECULE TYPE: peptide
(ix) FEATURE: amide-terminated, Xaa is p-aminophenylalanine
(xi) SEQUENCE DESCRIPTION: SEQ ID NO:67:
Xaa Ile Ala Gly Lys Ile Ala Xaa Ile Ala
Gly Lys Ile Ala Xaa Ile Ala Gly Lys Ile
Ala
(2) INFORMATION FOR SEQ ID NO:68:
(i) SEQUENCE CHARACTERISTICS:
(A) LENGTH: 2l amino acids
(B) TYPE: amino acid
(C) STRANDEDNESS:
(D) TOPOLOGY: linear
(ii) MOLECULE TYPE: peptide
(ix) FEATURE: amide-terminated, Xaa is p-aminophenylalanine
(xi) SEQUENCE DESCRIPTION: SEQ In NO:68:
Lys Ile Ala Gly Xaa Ile Ala Lys Ile Ala
Gly Xaa Ile Ala Lys Ile Ala Gly Xaa Ile
Ala
(2) INFORMATION FOR SEQ ID NO:69:
(i) SEQUENCE CHARACTERISTICS
SUBSTITUTE SH~ET
212~4~4
WO93/11783 PCT/US92/10427
-96-
(A) LENGTH: 21 amino acids
(B) TYPE: amino acid
(C) STRANDEDNESS:
(D) TOPOLOGY: linear
(ii) MOLECULE TYPE: peptide
(ix) FEATURE:
(D) OTHER INFORMATION: amide-terminated
(xi) SEQUENCE DESCRIPTION: SEQ ID NO:69:
Lys Leu Ala Ser Lys Ala Gly Lys Ile Ala Gly
Lys Ile Ala Lys Val Ala Leu Lys Ala Leu
(2) INFORMATION FOR SEQ ID NO:70:
(i) SEQUENCE CHARACTERISTICS
(A) LENGTH: 21 amino acids
(B) TYPE: amino acid
(C) STRANDEDNESS:
(D) TOPOLOGY: linear
(ii) MOLECULE TYPE: peptide
(ix) FEATURE:
(D) OTHER INFORMATION: amide-terminated, Xaa is
ornithine
(xi) SEQUENCE DESCRIPTION: SEQ ID NO:70:
Lys Ile Ala Gly Lys Ile Ala Lys Ile Ala Gly
Xaa Ile Ala Lys Ile Ala Gly Lys Ile Ala
SUBSTITIJT~ SHFET
21254~
WO93/11783 PCT/US92/10427
-97-
(2) INFORMATION FOR SEQ ID NO:71:
(i) SEQUENCE CHARACTERISTICS
(A) LENGTH: 21 amino acids
(B) TYPE: amino acid
(C) STRANDEDNESS:
(D) TOPOLOGY: linear
(ii) MOLECULE TYPE: peptide
(ix) FEATURE:
(D) OTHER INFORMATION: amide-terminated
(xi) SEQUENCE DESCRIPTION: SEQ ID NO:71:
Lys Ile Ala Gly Lys Ile Ala Lys Ile Ala
Gly Arg Ile Ala Lys Ile Ala Gly Lys Ile
Ala
(2) INFORMATION FOR SEQ ID NO:72:
(i) SEQUENCE CHARACTERISTICS
(A) LENGTH: 2l amino acids
(P) TYPE: amino acid
(C) STRANDEDNESS:
(D) TOPOLOGY: linear
(ii) M-;,.ECULE TYPE: peptide
(ix) FEATURE:
(D) OTHER INFORMATION: amide-termi,-~ted, Xaa is
norleucine
SUBSTl~J'r~ SHEET
2125~
WO93/11783 PCT/US92/10427
-98-
(xi) SEQUENCE DESCRIPTION: SEQ ID NO:72:
Lys Ile Ala Gly Lys Ile Ala Lys Ile Ala
Gly Xaa Ile Ala Lys Ile Ala Gly Lys Ile
Ala
(2) INFORMATION FOR SEQ ID NO:73:
(i) SEQUENCE CHARACTERISTICS
(A) LENGTH: 21 amino acids
(B) TYPE: amino acid
(C) STRANDEDNESS:
(D) TOPOLOGY: linear
(ii) MOLECULE TYPE: peptide
(ix) FEATURE:
(D) OTHER INFORMATION: amide-terminated, Xaa is
norvaline
(xi) SEQUENCE DESCRIPTION: SEQ ID NO:73:
Lys Ile Ala Gly Lys Ile Ala Lys Ile Ala
Gly Xaa Ile Ala Lys Ile Ala Gly Lys Ile
Ala
(2) INFORMATION FOR SEQ ID NO:74:
(i) SEQUENCE CHARACTERISTICS
(A) LENGTH: 2l amino acids
(B) TYPE: amino acid
(C) STRANDEDNESS:
(D) TOPOLOGY: linear
(ii) MOLECULE TYPE: peptide
~;UBSTITUTE ~3HEET
21254g4
W093/11783 PCT/US92/10427
_99
(ix) FEATURE:
(D) OTHER INFORMATION: amide-terminated, Xaa is
ornithine
(xi) SEQUENCE DESCRIPTION: SEQ ID NO:74:
Lys Phe Ala Gly Lys Phe Ala Lys Phe Ala Gly
Xaa Phe Ala Lys Phe Ala Gly Lys Phe Ala
(2) INFORMATION FOR SEQ ID NO:75:
(i) SEQUENCE CHARACTERISTICS
(A) LENGTH: 21 amino acids
(B) TYPE: amino acid
(C) STRANDEDNESS:
(D) TOPOLOGY: linear
(ii) MOLECULE TYPE: peptide
(ix) FEATURE:
(D) OTHER INFORMATION: amide-terminated, Xaa is
ornithine
(xi) SEQUENCE DESCRIPTION: SEQ ID NO:75:
Lys Ile Ala Gly Lys Phe Ala Lys Ile Ala
Gly Xaa Phe Ala Lys Ile Ala Gly Lys Phe
Ala
(2) INFORMATION FOR SEQ ID NO:76:
(i) SEQUENCE CHARACTERISTICS
(A) LENGTH: 2l amino acids
(B) TYPE: amino acid
SUBSTITUTE SHEET
21~5~4
W093/11783 PCT/US92/10427
-100-
(C) STRANDEDNESS:
(D) TOPOLOGY: linear
(ii) MOLECULE TYPE: peptide
(ix) FEATURE:
(D) OTHER INFORMATION: amide-terminated, Xaa at
residue 6, 13, and 20 is norleucine, Xaa at residue 12 is
ornithine.
(xi) SEQUENCE DESCRIPTION: SEQ ID NO:76:
Lys Ile Ala Gly Lys Xaa Ala Lys Ile Ala
Gly Xaa Xaa Ala Lys Ile Ala Gly Lys Xaa
Ala
(2) INFORMATION FOR SEQ ID NO:77:
(i) SEQUENCE CHARACTERISTICS
(A) LENGTH: 21 amino acids
(B) TYPE: amino acid
(C) STRANDEDNESS:
(D) TOPOLOGY: linear
(ii) MOLECULE TYPE: peptide
(ix) FEATURE:
(D) OTHER INFORMATION: amide-terminated
(xi) SEQUENCE DESCRIPTION: SEQ ID NO:77:
Lys Met Ala Ser Lys Ala Gly Lys Ile Ala
- 5 lO
Gly Lys Ile Ala Lys Val Ala Leu Lys Ala
SUBSTITUTE SH~ET
WO g3/11783 2 1 2 ~ 4 9 4 PCT/US92/10427
-101-
Leu
(2) INFORMATION FOR SEQ ID NO:78:
(i) SEQUENCE CHARACTERISTICS
(A) LENGTH: 21 amino acids
(B) TYPE: amino acid
(C) STRANDEDNESS:
(D) TOPOLOGY: linear
(ii) MOLECULE TYPE: peptide
(ix) FEATURE:
(D) OTHER INFORMATION: amide-terminated
(xi) SEQUENCE DESCRIPTION: SEQ ID NO:78
Lys Ile Ala Ser Lys Ala Gly Lys Ile Ala
Gly Lys Ile Ala Lys Val Ala Leu Lys Ala Leu
(2) INFORMATION FOR SEQ ID NO:79:
(i) SEQUENCE CHARACTERISTICS
(A) LENGTH: 21 amino acids
(B) TYPE: amino acid
(C) STRANDEDNESS:
(D) TOPOLOGY: linear
(ii) MOLECULE TYPE: peptide
(ix) FEATURE:
(D) OTHER INFORMATION: amide-terminated, Xaa is
norleucine
(xi) SEQUENCE DESCRIPTION: SEQ ID NO:79:
SUBSTITUTE SHEET
21254~4
W093/11783 PCT/US92/10427
-102-
Lys Ile Ala Ser Lys Ala Gly Lys Xaa Ala
Gly Lys Ile Ala Lys Val Ala Leu Lys Ala
Leu
(2) INFORMATION FOR SEQ ID NO:80:
(i) SEQUENCE CHARACTERISTICS
(A) LENGTH: 21 amino acids
(B) TYPE: amino acid
(C) STRANDEDNESS:
(D) TOPOLOGY: linear
(ii) MOLECULE TYPE: peptide
(ix) FEATURE:(D) OTHER INFORMATION: amide-terminated, Xaa is
norleucine
(xi) SEQUENCE DESCRIPTION: SEQ ID NO:80:
Lys Leu Ala Ser Lys Ala Gly Lys Xaa Ala
Gly Lys Ile Ala Lys Val Ala Leu Lys Ala
Leu
(2) INFORMATION FOR SEQ ID NO:81:
(i) SEQUENCE CHARACTERISTICS
(A) LENGTH: 21 amino acids
(B) TYPE: amino acid
(C) STRANDEDNESS:
(D) TOPOLOGY: linear
(ii) MOLECULE TYPE: peptide
SUBSTITUTE Sl I~~T
WO93/11783 2 1 2 .~ ~ 9 4 PCT/US92/10427
-103-
(ix) FEATURE:
(D) OTHER INFORMATION: amide-terminated, Xaa is
norleucine
(xi) SEQUENCE DESCRIPTION: SEQ ID NO:81:
Lys Xaa Ala Ser Lys Ala Gly Lys Xaa Ala
Gly Lys Ile Ala Lys Val Ala Leu Lys Ala Leu
(2) INFORMATION FOR SEQ ID NO:82:
(i) SEQUENCE CHARACTERISTICS:
(A) LENGTH: 2l amino acids
(B) TYPE: amino acid
(C) STRANDEDNESS:
(D) TOPOLOGY: linear
(ii) MOLECULE TYPE: peptide
(ix) FEATURE:
(D) OTHER INFORMATION: amide-terminated, Xaa is
p-aminophenylalanine.
(xi) SEQUENCE DESCRIPTION: SEQ ID NO:82:
Lys Ile Ala Gly Lys Ile Ala Lys Ile Ala
Gly Xaa Ile Ala Lys Ile Ala Gly Lys Ile
Ala
(2) INFORMATION FOR SEQ ID NO:83:
(i) SEQUENCE CHARACTERISTICS:
(A) LENGTH: 21 amino acids
(B) TYPE: amino acid
SUBSTIT~ S~
W093/1~132 S 4 ~ 4 PCT/US92/1~27
-104-
(C) STRANDEDNESS:
(D) TOPOLOGY: linear
(ii) MOLECULE TYPE: peptide
(ix) FEATURE:
(D) OTHER INFORMATION: amide-terminated
(xi) SEQUENCE DESCRIPTION: SEQ ID NO:83:ys Ile Ala Gly Ala Ile Ala Lys Ile Ala S 10ly Lys Ile Ala Lys Ile Ala Gly Lys Ile
lS 20la
2) INFORMATION FOR SEQ ID NO:84:
(i) SEQUENCE CHARACTERISTICS:
(A) LENGTH: 21 amino acids
(B) TYPE: amino acid
(C) STRANDEDNESS:
(D) TOPOLOGY: linear
(ii) MOLECULE TYPE: peptide
(ix) FEATURE:
(D) OTHER INFORMATION: amide-terminated
(xi) SEQUENCE DESCRIPTION: SEQ ID NO:84:ys Ile Ala Gly Lys Ile Ala Lys Ile Ala S 10ly Ala Ile Ala Lys Ile Ala Gly Lys Ile
lS 20la
SUBSTITUTE S~ET
WO93/11783 2 1 2 ~ 4 ~ 4 PCT/US92/lW27
-105-
(2) INFORMATION FOR SEQ ID NO:85:
(i) SEQUENCE CHARACTERISTICS:
(A) LENGTH: 21 amino acids
(B) TYPE: amino acid
(C) STRANDEDNESS:
(D) TOPOLOGY: linear
(ii) MOLECULE TYPE: peptide
(ix) FEATURE:
(D) OTHER INFORMATION: amide-terminated
(xi) SEQUENCE DESCRIPTION: SEQ ID NO:85:
Lys Ile Ala Gly Lys Ile Ala Lys Ile Ala
Gly Lys Ile Ala Lys Ile Ala Gly Ala Ile
Ala
(2) INFORMATION FOR SEQ ID NO:86:
(i) SEQUENCE CHARACTERISTICS
(A) LENGTH: 21 amino acids
(B) TYPE: amino acid
(C) STRANDEDNESS:
(D) TOPOLOGY: linear
(ii) MOLECULE TYPE: peptide
(ix) FEATURE:
(D) OTHER INFORMATION: amide-terminated
(xi) SEQUENCE DESCRIPTION: SEQ ID NO:86:
Lys Ile Ala Lys Lys Ile Ala Lys Ile Ala
SIJBST~TUTE ~
212~49~
W093/11783 PCT/US92/10427
-106-
Lys Lys Ile Ala Lys Ile Ala Lys Lys Ile
lS 20
Ala
2) INFORMATION FOR SEQ ID NO:B7:
(i) SEQUENCE CHARACTERISTICS
(A) LENGTH: 21 amino acids
(B) TYPE: amino acid
(C) STRANDEDNESS:
(D) TOPOLOGY: linear
(ii) MOLECULE TYPE: peptide
(ix) EEATURE:
(D) OTHER INFORMATION: amide-terminated
- (xi) SEQUENCE DESCRIPTION: SEQ ID NO:87:
Ala Ile Ala Gly Lys Ile Ala Lys Ile Ala
Gly Lys Ile Ala Lys Ile Ala Gly Lys Ile
Ala
(2) INFORMATION FOR SEQ ID NO:88:
(i) SEQUENCE CHARACTERISTICS
(A) LENGTH: 21 amino acids
(B) TYPE: amino acid
(C) STRANDEDNESS:
(D) TOPOLOGY: linear
(ii) MOLECULE TYPE: peptide
(ix) FEATURE:
(D) OTHER INFORMATION: amide-terminated . .
(xi) SEQUENCE DESCRIPTION: SEQ ID NO:88:
SUBSTITUTE S~lET
WO93/11783 2 1 2 5 ~ 9 4 PCT/US92/10427
-107-
Lys Ile Ala Gly Lys Ile Ala Ala Ile Ala
Gly Lys Ile Ala Lys Ile Ala Gly Lys Ile
Ala
(2) INFORMATION FOR SEQ ID NO:89:
(i) SEQUENCE CHARACTERISTICS
(A) LENGTH: 21 amino acids
(B) TYPE: amino acid
(C) STRANDEDNESS:
(D) TOPOLOGY: linear
(ii) MOLECULE TYPE: peptide
(ix) FEATURE:
(D) OTHER INFORMATION: amide-terminated
(xi) SEQUENCE DESCRIPTION: SEQ ID NO:89:
Lys Ile Ala Gly Lys Ile Ala Lys Ile Ala
Gly Lys Ile Ala Ala Ile Ala Gly Lys Ile
Ala
(2) INFORMATION FOR SEQ ID NO:90:
(i) SEQUENCE CHARACTERISTICS
(A) LENGTH: 21 amino acids
(B) TYPE: amino acid
(C) STRANDEDNESS:
(D) TOPOLOGY: linear
(ii) MOLECULE TYPE: peptide
(ix~ FEATURE:
SUBST,TUTE SHET
WO9~1783 PCT/US92/10427
-108-
(D) OTHER INFORMATION: amide-terminated
(xi) SEQUENCE DESCRIPTION: SEQ ID NO:90:
Gly Met Ala Ser Lys Ala Gly Lys Ile Ala
Gly Lys Ile Ala Lys Val Ala Leu Lys Ala
Leu
(2) INFORMATION FOR SEQ ID NO:9l:
(i) SEQUENCE CHARACTERISTICS:
(A) LENGTH: ll amino acids
(B) TYPE: amino acid
(C) STRANDEDNESS:
(D) TOPOLOGY: linear
(ii) MOLECULE TYPE: peptide
(ix) FEATURE:
(D) OTHER INFORMATION: May be a C-terminal amide,
and/or may be acetylated at N-terminus.
SU8STII UTE SHEE~
WO93/11783 2 1 2 5 4 9 4 PCT/US92/10427
-109-
(xi) SEQUENCE DESCRIPTION: SEQ ID NO:91:
Leu Lys Lys Leu Lys Lys Leu Leu Lys Leu
Leu
(2) INFORMATION FOR SEQ ID NO:92:
(i) SEQUENCE CHARACTERISTICS:
(A) LENGTH: 12 amino acids
(B) TYPE: amino acid
(C) STRANDEDNESS:
(D) TOPOLOGY: linear
(ii) MOLECULE TYPE: peptide
(ix) FEATURE:
(D) OTHER INFORMATION: May be a C-terminal amide,
and/or may be acetylated at N-terminus.
(xi) SEQUENCE DESCRIPTION: SEQ ID NO:92:
Leu Leu Lys Lys Leu Lys Lys Leu Leu Lys
Leu Leu
(2) INFORMATION FOR SEQ ID NO:93:
(i) SEQUENCE CHARACTERISTICS:
(A) LENGTH: 13 amino acids
(B) TYPE: amino acid
(C) STRANDEDNESS:
(D) TOPOLOGY: linear
(ii) MOLECULE TYPE: peptide
(ix) FEATURE:
- (D) OTHER INFORMATION: M~y be a C-terminal amide,
and/or may be acetylated at N-terminus.
SlJBSTlTllTE SH~ET
wo 93,l~2 S 4 ~ ~ PCT/US92/10427
- 110-
(xi) SEQUENCE DESCRIPTION: SEQ ID NO:93:
Lys Leu Leu Lys Lys Leu Lys Lys Leu Leu
Lys Leu Leu
(2) INFORMATION FOR SEQ ID NO:94:
(i) SEQUENCE CHARACTERISTICS:
(A) LENGTH: 14 amino acids
(B) TYPE: amino acid
(C) STRANDEDNESS:
(D) TOPOLOGY: linear
(ii) MOLECULE TYPE: peptide
(ix) FEATURE:
(D) OTHER INFORMATION: May be a C-terminal amide,
and/or may be acetylated at N-terminus.
(xi) SEQUENCE DESCRIPTION: SEQ ID NO:94:
Lys Lys Leu Leu Lys Lys Leu Lys Lys Leu
Leu Lys Leu Leu
(2) INFORMATION FOR.SEQ ID NO:95:
(i) SEQUENCE CHARACTERISTICS:
(A) LENGTH: 16 amino acids
(B) TYPE: amino acid
(C) STRANDEDNESS:
(D) TOPOLOGY: linear
(ii) MOLECULE TYPE: peptide
(ix) FEATURE:
(D) OTHER INFORMATION: May.be a C-terminal amide,
and/or may be acetylated at N-terminus.
~UBST!TtJTE SHEET
WO93/11783 2 1 2 5 ~ ~ I PCT/US92/10427
(xi) SEQUENCE DESCRIPTION: SEQ ID NO:95:
Lyç Lys Leu Leu Lys Lys Leu Lys Lys Leu
Leu Lys Lys Leu Arg Arg
(2) INFORMATION FOR SEQ ID NO:96:
(i) SEQUENCE CHARACTERISTICS:
(A) LENGTH: 16 amino acids
(B) TYPE: amino acid
(C) STRANDEDNESS:
(D) TOPOLOGY: linear
(ii) MOLECULE TYPE: peptide
(ix) FEATURE:
(D) OTHER INFORMATION: May be a C-terminal amide,
and/or may be acetylated at N-terminus.
(xi) SEQUENCE DESCRIPTION: SEQ ID NO:96:
Lys Leu Lys Lys Leu Leu Lys Lys Leu Lys
Lys Leu Leu Lys Leu Leu
(2) INFORMATION FOR SEQ ID NO:97:
(i) SEQUENCE CHARACTERISTICS:
(A) LENGTH: 22 amino acids
(B) TYPE: amino acid
(C) STRANDEDNESS:
(D) TOPOLOGY: linear
(ii) MOLECULE TYPE: peptide
.
(ix) FEATURE:
(D) OTHER INFORMATION: amide- or carboxy-terminated
SlJBSTiTliTE ~ET
~12S~g~
WO93/11783 PCT/US92/1~27
-112-
(xi) SEQUENCE DESCRIPTION: SEQ ID NO:97:
Gly Ile Gly Lys Phe Leu Lys Lys Ala Lys
Lys Phe Gly Lys Ala Phe Val Lys Ile Met
Lys Lys
SUBSTITUTE SH~ET
