Note: Descriptions are shown in the official language in which they were submitted.
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CHLAMYDIA ANTIGENS AND CORRESPONDING DNA FRAGMENTS
AND USES THEREOF
RELATED U.S. APPLICATION
The present patent application claims priority to the following United States
provisional
patent applications: U.S.S.Ns. 60/097,187, 60/097,188, 60/097,189, 60/097,190,
60/097,195,
60/097,196, and 60/097,197, filed August 20, 1998, and U.S.S.N. 60/097,191,
filed August 27,
1998.
FIELD OF THE INVENTION
The present invention relates to Chlamydia antigens and corresponding DNA
molecules,
1 o which can be used in methods to prevent and treat disease caused by
Chlamydia infection in
mammals, such as humans.
BACKGROUND OF THE INVENTION
Chlamydiae are prokaryotes. They exhibit morphologic and structural
similarities to Gram
negative bacteria including a trilaminar outer membrane, which contains
lipopolysaccharide and
15 several membrane proteins. Chlamydiae are differentiated from other
bacteria by their morphology
and by a unique developmental cycle. They are obligate intracellular parasites
with a unique
biphasic life cycle consisting of a metabolically inactive but infectious
extracellular stage and a
replicating but non-infectious intracellular stage. The replicative stage of
the life-cycle takes place
within a membrane-bound inclusion which sequesters the bacteria away from the
cytoplasm of the
2o infected host cell.
Because chlamydiae are small and multiply only within susceptible cells they
were long
thought to be viruses. However, they have many characteristics in common with
other bacteria:
(1) they contain both DNA and RNA, (2) they divide by binary fission, (3)
their cell envelopes
resemble those of other Gram-negative bacteria, (4) they contain ribosomes
similar to those of
25 other bacteria, and (5) they are susceptible to various antibiotics.
Chlamydiae can be seen in the
light microscope, and the genome is about one-third the size of the
Escherichia toll genome.
Many different strains of chlamydiae have been isolated from birds, man, and
other
mammals, and these strains can be distinguished on the basis of host range,
virulence,
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CONFIRMATION COPT
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pathogenesis, and antigenic composition. There is strong homology of DNA
within each species,
but surprisingly little between species, suggesting long-standing evolutionary
separation.
C. trachomatis has a high degree of host specificity, being almost completely
limited to
man; it causes ocular and genitourinary infections of widely varying severity.
In contrast,
C. psittaci strains are rare in man but are found in a wide range of birds and
also in wild, domestic,
and laboratory mammals, where they multiply in cells of many organs.
C. pneumoniae is a common human pathogen, originally described as the TWAR
strain of
C. psittaci, but subsequently recognized to be a new species. C. pneumoniae is
antigenically,
genetically, and morphologically distinct from other Chlamydia species (C.
trachomatis,
1 o C. pecorum and C. psittaci). It shows 10% or less DNA sequence homology
with either of
C. trachomatis or C. psittaci and so far appears to consist of only a single
strain, TWAR.
C. pneumoniae is a common cause of community acquired pneumonia, less frequent
only
than Streptococcus pneumoniae and Mycoplasma pneumoniae. Grayston et al., J.
Infect. Dis. 168:
1231 (1995); Campos et al., Invest. Ophthalmol. Vis. Sci. 36: 1477 (1995),
each incorporated
herein by reference. It can also cause upper respiratory tract symptoms and
disease, including
bronchitis and sinusitis. See, e.g., Grayston et al., J. Infect. Dis. 168:
1231 (1995); Campos et al.,
Invest. Ophthalmol. Vis. Sci. 36: 1477 (1995); Grayston et al., J. Infect.
Dis. 161: 618 (1990);
Marrie, Clin. Infect. Dis. 18: 501 (1993). The great majority of the adult
population (over 60%)
has antibodies to C. pneumoniae (Wang et al., Chlamydial Infections, Cambridge
University Press,
2o Cambridge, p. 329 (1986)), indicating past infection which was unrecognized
or asymptomatic.
C. pneumoniae infection usually presents as an acute respiratory disease
(i.e., cough, sore
throat, hoarseness, and fever; abnormal chest sounds on auscultation). For
most patients, the
cough persists for 2 to 6 weeks, and recovery is slow. In approximately 10% of
these cases, upper
respiratory tract infection is followed by bronchitis or pneumonia.
Furthermore, during a
C. pneumoniae epidemic, subsequent co-infection with pneumococcus has been
noted in about half
of these pneumonia patients, particularly in the infirm and the elderly. As
noted above, there is
more and more evidence that C. pneumoniae infection is also linked to diseases
other than
respiratory infections.
The reservoir for the organism is presumably people. In contrast to C.
psittaci infections,
3o there is no known bird or animal reservoir. Transmission has not been
clearly defined. It may
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result from direct contact with secretions, from formites, or from airborne
spread. There is a long
incubation period, which may last for many months. Based on analysis of
epidemics,
C. pneumoniae appears to spread slowly through a population (case-to-case
interval averaging
30 days) because infected persons are inefficient transmitters of the
organism. Susceptibility to
C. pneumoniae is universal. Reinfections occur during adulthood, following the
primary infection
as a child. C. pneumoniae appears to be an endemic disease throughout the
world, noteworthy for
superimposed intervals of increased incidence (epidemics) that persist for 2
to 3 years. C.
trachomatis infection does not confer cross-immunity to C. pneumoniae.
Infections are easily
treated with oral antibiotics, tetracycline or erythromycin (2 g/day, for at
least 10 to 14 days). A
to recently developed drug, azithromycin, is highly effective as a single-dose
therapy against
chlamydial infections.
In most instances, C.'. pneumoniae infection is mild and without
complications, and up to
90% of infections are subacute or unrecognized. Among children in
industrialized countries,
infections have been thought to be rare up to the age of five years, although
a recent study has
15 reported that many children in this age group show PCR evidence of
infection despite being
seronegative, and estimates a prevalence of 17-19% in 2-4 years old. See,
Normann et al., Acta
Paediatrica, 87: 23-27 ( 1998). In developing countries, the seroprevalence of
C. pneumoniae
antibodies among young children is elevated, and there are suspicions that C.
pneumoniae may be
an important cause of acute lower respiratory tract disease and mortality for
infants and children in
2o tropical regions of the world.
From seroprevalence studies and studies of local epidemics, the initial C.
pneumoniae
infection usually happens between the ages of 5 and 20 years. In the USA, fox
example, there are
estimated to be 30,000 cases of childhood pneumonia each year caused by C.
pneumoniae.
Infections may cluster among groups of children or young adults (e.g., school
pupils or military
25 conscripts).
C. pneumoniae causes 10 to 25% of community-acquired lower respiratory tract
infections
(as reported from Sweden, Italy, Finland, and the USA). During an epidemic, C.
pneumonia
infection may account for 50 to 60% of the cases of pneumonia. During these
periods, also, more
episodes of mixed infections with S. pneumoniae have been reported.
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Reinfection during adulthood is common; the clinical presentation tends to be
milder.
Based on population seroprevalence studies, there tends to be increased
exposure with age, which
is particularly evident among men. Some investigators have speculated that a
persistent,
asymptomatic C. pneumoniae infection state is common.
In adults of middle age or older, C. pneumoniae infection may progress to
chronic
bronchitis and sinusitis. A study in the USA revealed that the incidence of
pneumonia caused by
C. pneumoniae in persons younger than 60 years is 1 case per 1,000 persons per
year; but in the
elderly, the disease incidence rose three-fold. C pneumoniae infection rarely
leads to
hospitalization, except in patients with an underlying illness.
to Of considerable importance is the association of atherosclerosis and C.
pneumoniae
infection. There are several epidemiological studies showing a correlation of
previous infections
with C. pneumoniae and heart attacks, coronary artery and carotid artery
disease. See, Saikku et
al., Lancet 2: 983 (1988); Thom et al., JAMA 268: 68 (1992); Linnanmaki et
al., Circulation 87:
1030 (1993); Saikku et al., Annals Int. Med. 116: 273 (1992); Melnick et al.,
Am. J. Med. 95: 499
i 5 ( 1993). Moreover, the organisms has been detected in atheromas and fatty
streaks of the coronary,
carotid, peripheral arteries and aorta. See, Shor et al., South African Med.
J. 82: 158 (1992); Kuo
et al., J. Infect. Dis. 167: 841 (i993); Kuo et al., Arteriosclerosis and
Thrombosis 13: 1500 (1993);
Campbell et al., J. Infect. Dis. 172: 585 (1995); Chiu et al., Circulation 96:
2144-2148 (1997).
Viable C. pneumoniae has been recovered from the coronary and carotid artery.
Ramirez et al.,
2o Annals Int. Med. 125: 979 (1996); Jackson et al., Abst. K121, p272, 36th
ICAAC, New Orleans
( 1996). Furthermore, it has been shown that C. pneumoniae can induce changes
of atherosclerosis
in a rabbit model. See, Fong et al., (1997) Journal of Clinical Microbiololo~
35: 48. Taken
together, these results indicate that it is highly probable that C. pneumoniae
can cause
atherosclerosis in humans, though the epidemiological importance of chlamydial
atherosclerosis
25 remains to be demonstrated.
A number of recent studies have also indicated an association between C.
pneumoniae
infection and asthma. Infection has been linked to wheezing, asthmatic
bronchitis, adult-onset
asthma and acute exacerbation of asthma in adults, and small-scale studies
have shown that
prolonged antibiotic treatment was effective at greatly reducing the severity
of the disease in some
3o individuals. Hahn et al., Ann Allergy Asthma Immunol. 80: 45-49 (1998);
Hahn et al., Epidemiol
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WO 00111183 PCT/IB99/01449
Infect. 117: 513-517 (1996); Bjornsson et al., Scand J Infect. Dis. 28: 63-69
(1996); Hahn, J. Fam.
Pract. 41: 345-351 (1995); Allegra et al., Eur. Respir. J. 7: 2165-2168
(1994); Hahn et al., JAMA
266: 225-230 (1991).
In light of these results, a protective vaccine against disease caused by C.
pneumoniae
s infection would be of considerable importance. There is not yet an effective
vaccine for human
C. pneumoniae infection. Nevertheless, studies with C. trachomatis and G
psittaci indicate that
this is an attainable goal. For example, mice which have recovered from a lung
infection with
C. trachomatis are protected from infertility induced by a subsequent vaginal
challenge. Pal et al.,
Infection and Immunity 64: 5341 (1996). Similarly, sheep immunized with
inactivated C. psittaci
t0 were protected from subsequent chlamydial-induced abortions and
stillbirths. Jones et al., Iraccine
13: 715 ( 1995). Protection from chlamydial infections has been associated
with Th 1 immune
responses, particularly the induction of INFy-producing CD4+ T cells.
Igietsemes et al.,
Immunology 5: 317 (1993). The adoptive transfer of CD4+ cell lines or clones
to nude or SCID
mice conferred protection from challenge or cleared chronic disease (Igietseme
et al., Regional
~5 Immunology S: 317 (1993); Magee et al., Regional Immunology 5: 305 (1993)),
and in vivo
depletion of CD4+ T cells exacerbated disease post-challenge (Larders et al.,
Infection 8c
Immunity 59: 3774 (1991); Magee et al., Infection ~c Immunity 63: 516 (1995)).
However, the
presence of sufficiently high titres of neutralizing antibody at mucosal
surfaces can also exert a
protective effect. Cotter et al., Infection and Immunity 63: 4704 (1995).
2o The extent of antigenic variation within the species C. pneumoniae is not
well
characterized. Serovars of C. trachomatis are defined on the basis of
antigenic variation in major
outer membrane proteins (MOMP), but published C. pneumoniae MOMP gene
sequences show no
variation between several diverse isolates of the organism. See, Campbell et
al., Infection and
Immunity 58: 93 (1990); McCafferty et al., Infection and Immunity 63: 2387-9
(1995); Knudsen et
2s al., Third Meeting of the European Society for Chlamydia Research, Vienna
(1996). Regions of
the protein known to be conserved in other chlamydial MOMPs are conserved in
C. pneumoniae.
See, Campbell et al., Infection and Immunity 58: 93 (1990); McCafferty et al.,
Infection and
Immunity 63: 2387-9 (1995). One study has described a strain of C. pneumoniae
with a MOMP of
greater that usual molecular weight, but the gene for this has not been
sequenced. Grayston et al.,
3o J. Infect. Dis. 168: 1231 (1995). Partial sequences of outer membrane
protein 2 from nine diverse
isolates were also found to be invariant. Ramirez et al., Annals Int. Med.
125: 979 (1996). The
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genes for HSP60 and HSP70 show little variation from other chlamydial species,
as would be
expected. The gene encoding a 76 kDa antigen has been cloned from a single
strain of
C. pneumoniae. It has no significant similarity with other known chlamydial
genes. Marrie, Clin.
Infect. Dis. 18: 501 (1993).
Many antigens recognized by immune sera to C. pneumoniae are conserved across
all
chlamydiae, but 98kDa, 76 kDa and S4 kDa proteins may be C. pneumoniae-
specific. Campos et
al., Invest. Ophthalmol. Vis. Sci. 36: 1477 (1995); Marrie, Clin. Infect. Dis.
18: 501 (1993);
Wiedmann-AI-Ahmad et al., Clin. Diagn. Lab. Immunol. 4: 700-704 (1997).
Immunoblotting of
isolates with sera from patients does show variation of blotting patterns
between isolates,
1o indicating that serotypes C. pneumoniae may exist. Grayston et al., J.
Infect. Dis. 168: 1231
(1995); Ramirez et al., Annals Int. Med. 125: 979 (1996). However, the results
are potentially
confounded by the infection status of the patients, since immunoblot profiles
of a patient's sera
change with time post-infection. An assessment of the number and relative
frequency of any
serotypes, and the defining antigens, is not yet possible.
Thus, a need remains for effective compositions for preventing, treating, and
diagnosing
Chlamydia infections.
SUMMARY OF THE INVENTION
In one aspect, the present invention provides purified and isolated DNA
molecules, which
encode Chlamydia that can be used in methods to prevent, treat, and diagnose
Chlamydia
infection. The present invention provides eight separate preferred DNA
molecules, each
individually and separately defined by one SEQ ID NOS: 1, 3, S, 7, 9, 11, 13,
or 15. Further
provided in the present invention are eight separate preferred polypeptides,
each individually and
separately defined by one of SEQ ID NOS: 2, 4, 6, 8, 10, 12, 14, or 16. Those
skilled in the art
will appreciate that the invention also includes DNA molecules that encode
mutants, variants, and
derivatives of such polypeptides, which result from the addition, deletion, or
substitution of
non-essential amino acids as described herein. The invention also includes RNA
molecules
corresponding to the DNA molecules of the invention.
In addition to the DNA and RNA molecules, the invention includes the
corresponding
polypeptides and monospecific antibodies that specifically bind to such
polypeptides.
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The present invention has wide application and includes expression cassettes,
vectors, and
cells transformed or transfected with the polynucleotides of the invention.
Accordingly, the
present invention provides: (i) a method for producing a polypeptide of the
invention in a
recombinant host system and related expression cassettes, vectors, and
transformed or transfected
cells; (ii) a live vaccine vectors such as viral or bacterial live vaccine
vectors, including, pox virus,
alphavirus, Salmonella typhimurium, or Vibrio cholerae vector, containing a
polynucleotide of the
invention, such vaccine vectors being useful for, e.g., preventing and
treating Chlamydia infection,
in combination with a diluent or carrier, and related pharmaceutical
compositions and associated
therapeutic and/or prophylactic methods; (iii) a therapeutic and/or
prophylactic method involving
administration of an RNA or DNA molecule of the invention, either in a naked
form or formulated
with a delivery vehicle, a polypeptide or combination of polypeptides, or a
monospecific antibody
of the invention, and related pharmaceutical compositions; (iv) a method for
diagnosing the
presence of Chlamydia in a biological sample, which can involve the use of a
DNA or RNA
molecule, a monospecific antibody, or a polypeptide of the invention; and (v)
a method for
purifying a polypeptide of the invention by antibody-based affinity
chromatography.
BRIEF DESCRIPTION OF THE DRAWINGS
The present invention will be fuxther understood from the following
description with
reference to the drawings, in which:
FIG. 1 shows the nucleotide sequence (top sequence) and the deduced amino acid
sequence
of the full-length (middle sequence) and processed (bottom sequence) CPN
100111 polypeptide
from Chlamydia pneumoniae.
FIG. 2 shows the restriction enzyme analysis of the nucleotide sequence
encoding
C. pneumoniae CPN100111.
FIG. 3 shows the nucleotide sequence (top sequence) and the deduced amino acid
sequence
of the full-length (middle sequence) and processed (bottom sequence) CPN
100224 polypeptide
from Chlamydia pneumoniae.
FIG. 4 shows the restriction enzyme analysis of the nucleotide sequence
encoding the
C. pneumoniae CPN 100224.
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FIG. 5 shows the nucleotide sequence (top sequence) and the deduced amino acid
sequence
(bottom sequence) of the CPN100230 polypeptide from Chlamydia pneumoniae.
FIG. 6 shows the restriction enzyme analysis of the nucleotide sequence
encoding the
C. pneumoniae CPN100230.
FIG. 7 shows the nucleotide sequence (top sequence) and the deduced amino acid
sequence
(bottom sequence) of the CPN100231 polypeptide from Chlamydia pneumoniae.
FIG. 8 shows the restriction enzyme analysis of the nucleotide sequence
encoding the
C. pneumoniae CPN 100231.
FIG. 9 shows the nucleotide sequence (top sequence) and the deduced amino acid
sequence
(bottom sequence) of the CPN100232 polypeptide from Chlamydia pneumoniae.
FIG. 10 shows the restriction enzyme analysis of the nucleotide sequence
encoding the
C. pneumoniae CPN 100232.
FIG. 11 shows the nucleotide sequence (top sequence) and the deduced amino
acid
sequence (bottom sequence) of the CPN100233 polypeptide from Chlamydia
pneumoniae.
15 FIG. 12 shows the restriction enzyme analysis of the nucleotide sequence
encoding the
C. pneumoniae CPN100233.
FIG. 1'3 shows the nucleotide sequence (top sequence) and the deduced amino
acid
sequence of the full-length (middle sequence) and processed (bottom sequence)
CPN100394
polypeptide from Chlamydia pneumoniae.
2o FIG. 14 shows the restriction enzyme analysis of the nucleotide sequence
encoding the
C. pneumoniae CPN100394.
FIG. 1 S shows the nucleotide sequence (top sequence) and the deduced amino
acid
sequence (bottom sequence) of the CPN100395 polypeptide from Chlamydia
pneumoniae.
FIG. 16 shows the restriction enzyme analysis of the nucleotide sequence
encoding the
25 C. pneumoniae CPN 100395.
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DETAILED DESCRIPTION OF THE INVENTION
In the C. pneumoniae genome, open reading frames (ORFs) encoding chlamydial
polypeptides have been identified. These polypeptides include polypeptides
permanently found in
the bacterial membrane structure, polypeptides that are present in the
external vicinity of the
bacterial membrane, polypeptides permanently found in the inclusion membrane
structure,
polypeptides that are present in the external vicinity of the inclusion
membrane, and polypeptides
that are released into the cytoplasm of the infected cell. These polypeptides
can be used in
vaccination methods for preventing and treating Chlamydia infection.
According to a first aspect of the invention, there are provided isolated
polynucleotides
encoding the precursor and mature forms of Chlamydia polypeptides.
An isolated polynucleotide of the invention encodes a polypeptide having an
amino acid
sequence that is homologous to a Chlamydia amino acid sequence, the Chlamydia
amino acid
sequence being selected from the group consisting of the amino acid sequences
as shown in SEQ
ID NOS: 2, 4, 6, 8, 10, 12, 14, or 16.
The term "isolated polynucleotide" is defined as a polynucleotide removed from
the
environment in which it naturally occurs. For example, a naturally-occurring
DNA molecule
present in the genome of the bacteria is not isolated, but the same molecule
separated from the
remaining part of the bacterial genome, as a result of, e.g., a cloning event
(amplification), is
isolated. Typically, an isolated DNA molecule is free from DNA regions (e.g.,
coding regions)
2o with which it is immediately contiguous at the S' or 3' end, in the
naturally occurring genome.
Such isolated polynucleotides could be part of a vector or a composition and
still be isolated in that
such a vector or composition is not part of its natural environment.
A polynucleotide of the invention can be in the form of RNA or DNA (e.g.,
cDNA,
genomic DNA, or synthetic DNA), or modifications or combinations thereof. The
DNA can be
double-stranded or single-stranded, and, if single-stranded, can be the coding
strand or the
non-coding (anti-sense) strand. The sequence that encodes a polypeptide of the
invention as
shown in SEQ ID NOS: 2, 4, 6, 8, 10, 12, 14 or 16 can be: (a) the coding
sequence as shown in
either SEQ ID NOS: 1, 3, S, 7, 9, 11, 13 and 15; or (b) a ribonucleotide
sequence derived by
transcription of (a); or (c) a different coding sequence; this latter, as a
result of the redundancy or
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degeneracy of the genetic code, encodes the same polypeptides as the DNA
molecules of which the
nucleotide sequences are illustrated in SEQ ID NOS: l, 3, 5, 7, 9, 11, 13, and
15..
By "homologous amino acid sequence" is meant an amino acid sequence that
differs from
an amino acid sequence shown in SEQ ID NOS: 2, 4, 6, 8, 10, 12, 14 or 16, only
by one or more
conservative amino acid substitutions, or by one or more non-conservative
amino acid
substitutions, deletions, or additions located at positions at which they do
not destroy the specific
antigenicity of the polypeptide.
Preferably, such a sequence is at least 75%, more preferably 80%, and most
preferably 90%
identical to an amino acid sequence shown in SEQ ID NOS: 2, 4, 6, 8, 10, 12,
14 or 16.
l0 Homologous amino acid sequences include sequences that are identical or
substantially
identical to an amino acid sequence as shown in SEQ ID NOS: 2, 4, 6, 8, 10,
12, 14 and 16. By
"amino acid sequence substantially identical" is meant a sequence that is at
least 90%, preferably
95%, more preferably 97%, and most preferably 99% identical to an amino acid
sequence of
reference and that preferably differs from the sequence of reference, if at
all, by a majority of
t 5 conservative amino acid substitutions.
Conservative amino acid substitutions typically include substitutions among
amino acids of
the same class. These classes include, for example, (a) amino acids having
uncharged polar side
chains, such as asparagine, glutamine, serine, threonine, and tyrosine; (b)
amino acids having basic
side chains, such as lysine, arginine, and histidine; (c) amino acids having
acidic side chains, such
20 as aspartic acid and glutamic acid; and (d) amino acids having nonpolar
side chains, such as
glycine, alanine, valine, leucine, isoleucine, proline, phenylalanine,
methionine, tryptophan, and
cysteine.
Homology is typically measured using sequence analysis software (e.g.,
Sequence Analysis
Software Package of the Genetics Computer Group, University of Wisconsin
Biotechnology
25 Center, 1710 University Avenue, Madison, WI 53705). Similar amino acid
sequences are aligned
to obtain the maximum degree of homology (i. e., identity). To this end, it
may be necessary to
introduce gaps into the sequence. Once the optimal alignment has been set up,
the degree of
homology (i.e., identity) is established by recording all of the positions in
which the amino acids of
both sequences are identical, relative to the total number of positions.
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Alternatively, homology can be determined by aligning the candidate sequence
and the
reference sequence using an alignment tool, such as the dynamic programming
algorithm
described in Needleman et al., J. Mol. Blol. 48: 443 -( 1970), and the Align
Program, a commercial
software package produced by DNAstar, Inc., the teachings of which are
incorporated by reference
herein. After the initial alignment is made, it can be refined by comparison
to a multiple sequence
alignment of a family of related proteins. Once the alignment between the
candidate and reference
sequences is made and refined, a percent homology score is calculated. The
individual amino
acids of each sequence are compared sequentially according to their similarity
to each other.
Similarity factors include similar size, shape and electrical charge. One
particularly
1 o preferred method of determining amino acid similarities is the PAM250
matrix described in
Dayhoff et al., 5 ATLAS OF PROTEIN SEQUENCE AND STRUCTURE 345-352 (1978 &
Supp.),
incorporated by reference herein. A similarity score is first calculated as
the sum of the aligned
pairwise amino acid similarity scores. Insertions and deletions are ignored
for the purposes of
percent homology and identity. Accordingly, gap penalties are not used in this
calculation. The
15 raw score is then normalized by dividing it by the geometric mean of the
scores of the candidate
compound and the reference sequence. The geometric mean is the square root of
the product of
these scores. The normalized raw score is the percent homology.
Preferably, a homologous sequence is one that is at least 45%, more preferably
60%, and
most preferably 85% identical to a coding sequence of SEQ ID NOS: 1, 3, 5, 7,
9, 11, 13, and 15.
2o Polypeptides having a sequence homologous to one of the sequences shown in
SEQ ID
NOS: 2, 4, 6, 8, 10, 12, 14 or 16 include naturally-occurring allelic
variants, as well as mutants and
variants or any other non-naturally-occurring variants that are analogous in
terms of antigenicity,
to a polypeptide having a sequence as shown in SEQ ID NOS: 2, 4, 6, 8, 10, 12,
14 or 16.
An allelic variant is an alternate form of a polypeptide that is characterized
as having a
25 substitution, deletion, or addition of one or more amino acids that does
not substantially alter the
biological function of the polypeptide. By "biological function" is meant the
function of the
polypeptide in the cells in which it naturally occurs, even-if the function is
not necessary for the
growth or survival of the cells. For example, the biological function of a
porin is to allow the entry
into cells of compounds present in the extracellular medium. The biological
function is distinct
3o from the antigenic function. A polypeptide can have more than one
biological function.
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Allelic variants are very common in nature. For example, a bacterial species,
e.g.,
C. pneumoniae, is usually represented by a variety of strains that differ from
each other by minor
allelic variations. Indeed, a polypeptide that fulfills the same biological
function in different
strains can have an amino acid sequence that is not identical in each of the
strains. Such an allelic
variation may be equally reflected at the polynucleotide level.
Support for the use of allelic variants of polypeptide antigens comes from,
e.g., studies of
the Chlamydial MOMP antigen. The amino acid sequence of the MOMP varies from
strain to
strain, yet cross-strain antibody binding plus neutralization of infectivity
occurs, indicating that the
MOMP, when used as an immunogen, is tolerant of amino acid variations.
to Polynucleotides, e.g., DNA molecules, encoding allelic variants can easily
be retrieved by
polymerise chain reaction (PCR) amplification of genomic bacterial DNA
extracted by
conventional methods. This involves the use of synthetic oligonucleotide
primers matching
upstream and downstream of the 5' and 3' ends of the encoding domain. Suitable
primers can be
designed according to the nucleotide sequence information provided in SEQ ID
NOS: 1, 3, 5, 7, 9,
15 11, 13 and 15. Typically, a primer can consist of 10 to 40, preferably 15
to 25 nucleotides. It may
be also advantageous to select primers containing C and G nucleotides in a
proportion sufficient to
ensure efficient hybridization; e.g., an amount of C and G nucleotides of at
least 40%, preferably
50% of the total nucleotide amount.
Useful homologs that do not naturally occur can be designed using known
methods for
2o identifying regions of an antigen that are likely to be tolerant of amino
acid sequence changes
and/or deletions. For example, sequences of the antigen from different species
can be compared to
identify conserved sequences.
Polypeptide derivatives that are encoded by polynucleotides of the invention
include, e.g.,
fragments, polypeptides having large internal deletions derived from full-
length polypeptides, and
25 fusion proteins.
Polypeptide fragments of the invention can be derived from a polypeptide
having a
sequence homologous to any of the sequences shown in SEQ ID NOS: 2, 4, 6, 8,
10, 12, 14 or 16,
to the extent that the fragments retain the desired substantial antigenicity
of the parent polypeptide
(specific antigenicity). Polypeptide derivatives can also be constructed by
large internal deletions
3o that remove a substantial part of the parent polypeptide, while retaining
the desired specific
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antigenicity. Generally, polypeptide derivatives should be about at least 12
amino acids in length
to maintain the antigenicity. Advantageously, they can be at least 20 amino
acids, preferably_at
least SO amino acids, more preferably at least 75 amino acids, and most
preferably at least
100 amino acids in length.
Useful polypeptide derivatives, e.g., polypeptide fragments, can be designed
using
computer-assisted analysis of amino acid sequences in order to identify sites
in protein antigens
having potential as surface-exposed, antigenic regions. Hughes et al., Infect.
Immun. 60: 3497
( 1992).
Polypeptide fragments and polypeptides having large internal deletions can be
used for
revealing epitopes that are otherwise masked in the parent polypeptide and
that may be of
importance for inducing, for example, a protective T cell-dependent immune
response. Deletions
can also remove inununodorninant regions of high variability among strains.
It is an accepted practice in the field of immunology to use fragments and
variants of
protein immunogens as vaccines and immunogens, as all that is required to
induce an immune
response to a protein may be a small (e.g., 8 to 10 amino acid) region of the
protein. This has been
done for a number of vaccines against pathogens other than Chlamydia. For
example, short
synthetic peptides corresponding to surface-exposed antigens of pathogens such
as murine
mammary tumor virus, peptide containing 11 amino acids (Dion et al., Virology
179: 474-477
(1990)); Semliki Forest virus, peptide containing 16 amino acids (Snijders et
al., J. Gen. Virol. 72:
557-565 (1991)); and canine parvovirus, two overlapping peptides, each
containing 15 amino acids
(Langeveld et al., Vaccine 12: 1473-1480 (1994)) have been shown to be
effective vaccine
antigens against their respective pathogens.
Polynucleotides encoding polypeptide fragments and polypeptides having large
internal
deletions can be constructed using standard methods (see, e.g., Ausubel et
al., CURRENT
PROTOCOLS IN MOLECULAR BIOLOGY, John Wiley & Sons Inc. (1994)); for example,
by PCR,
including inverse PCR, by restriction enzyme treatment of the cloned DNA
molecules, or by the
method of Kunkel et al. (Proc. Natl. Acad. Sci. USA 82: 448 ( 1985));
biological material available
at Stratagene.
A polypeptide derivative can also be produced as a fusion polypeptide that
contains a
3o polypeptide or a polypeptide derivative of the invention fused, e.g., at
the N- or C-terminal end, to
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any other polypeptide. For construction of DNA encoding the amino acid
sequence corresponding
to hybrid fusion proteins, a first DNA encoding amino acid sequence
corresponding to any of the
portions of CPN100111, CPN 100224, CPN 100230, CPN 100231, CPN 100232, CPN
100233,
CPN 100394, or CPN 100395 nucleotide sequence (SEQ ID NOS: l, 3, 5, 7, 9, 11,
13 or 15) is
joined to a second DNA using methods described in, for example, U.S. Patent
5,844,095,
incorporated herein by reference. A product can then be easily obtained by
translation of the
genetic fusion. Vectors for expressing fusion polypeptides are commercially
available, such as the
pMal-c2 or pMal-p2 systems of New England Biolabs, in which the fusion peptide
is a maltose
binding protein, the glutathione-S-transferase system of Pharmacia, or the His-
Tag system
Io available from Novagen. These and other expression systems provide
convenient means for
further purification of polypeptides and derivatives of the invention.
Another particular example of fusion polypeptides included in the invention
includes a
polypeptide or polypeptide derivative of the invention fused to a polypeptide
having adjuvant
activity, such as, e.g., the subunit B of either cholera toxin or E. coli heat-
labile toxin. Several
possibilities are can be used for achieving fusion. First, the polypeptide of
the invention can be
fused to the N-, or preferably, to the C-terminal end of the polypeptide
having adjuvant activity.
Second, a polypeptide fragment of the invention can be fused within the amino
acid sequence of
the polypeptide having adjuvant activity.
As stated above, the polynucleotides of the invention encode Chlamydia
polypeptides in
2o precursor or mature form. They can also encode hybrid precursors containing
heterologous signal
peptides, which can mature into polypeptides of the invention. By
"heterologous signal peptide" is
meant a signal peptide that is not found in the naturally-occurnng precursor
of a polypeptide of the
invention.
A polynucleotide of the invention, having_a homologous coding sequence,
hybridizes,
preferably under stringent conditions, to a polynucleotide having a sequence
as shown in SEQ ID
NOS: 2, 4, 6, 8, 10, 12, 14 and i6. Hybridization procedures are described in,
e.g., Ausubel et al.,
CURRENT PROTOCOLS IN MOLECULAR BIOLOGY, John Wiley & Sons Inc. (1994); Silhavy
et al.,
EXPERIMENTS WITH GENE FUSIONS, Cold Spring Harbor Laboratory Press (1984);
Davis et al., A
MANUAL FOR GENETIC ENGINEERING: ADVANCED BACTERIAL GENETICS, Cold Spring
Harbor
3o Laboratory Press (1980), each incorporated herein by reference. Important
parameters that can be
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WO 00/11183 PCT/IB99/01449
considered for optimizing hybridization conditions are reflected in a formula
that allows
calculation of a critical value, the melting temperature above which two
complementary DNA
strands separate from each other. Casey and Davidson, Nucl. Acid Res. 4: 1539
(1977). This
formula is as follows:
Tm = 81.5 + 0.5 x (% G+C) + 1.6 log (positive ion concentration) - 0.6 x (%
formamide).
Under appropriate stringency conditions, hybridization temperature (Th) is
approximately
20-40°C, 20-25°C or, preferably, 30-40°C below the
calculated Tm. Those skilled in the art will
understand that optimal temperature and salt conditions can be readily
determined empirically in
preliminary experiments using conventional procedures.
to For example, stringent conditions can be achieved, both for pre-hybridizing
and
hybridizing incubations, (i) within 4-16 hours at 42°C, in 6xSSC
containing 50% formamide or (ii)
within 4-16 hours at 65°C in an aqueous 6xSSC solution (1 M NaCI, 0.1 M
sodium citrate
(pH 7.0)).
For polynucleotides containing 30 to 600 nucleotides, the above formula is
used and then is
15 corrected by subtracting (600/polynucleotide size in base pairs).
Stringency conditions are defined
by a Th that is 5 to 10°C below Tm.
Hybridization conditions with oligonucleotides shorter than 20-30 bases do not
exactly
follow the rules set forth above. In such cases, the formula for calculating
the Tm is as follows:
Tm = 4 x (G+C) + 2 (A+T).
20 For example, an 18 nucleotide fragment of 50% G+C would have an approximate
Tm of 54°C.
A polynucleotide molecule of the invention, containing RNA, DNA, or
modifications or
combinations thereof, can have various applications. For example, a DNA
molecule can be used:
(i) in a process for producing the encoded polypeptide in a recombinant host
system, (ii) in the
construction of vaccine vectors such as poxviruses, which are further used in
methods and
25 compositions for preventing and/or treating Chlamydia infection, (iii) as a
vaccine agent (as well
as an RNA molecule), in a naked form or formulated with a delivery vehicle
and, (iv) in the
construction of attenuated Chlamydia strains that can overexpress a
polynucleotide of the
invention or express it in a modified, mutated form, such as a non-toxic form,
if appropriate.
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For vaccine compositions and uses of the proteins and peptides and encoding
nucleotides of the
present invention for protection against diseases caused by Chlamydia, it is
not preferred to use
naked DNA encoding the protein or peptides and administering these nucleotides
intranasally or
intramuscularly. For these proteins, it is preferred to administer the
encoding nucleic acids by
other routes such as intradermally and/or to formulate the encoding nucleic
acids to improve (or
adjuvant) the immune response. It is also preferred to include the encoding
nucleic acid as part of
a recombinant live vector, such as a viral or bacterial vector for use as the
immunizing agent. It is
also preferred to immunize with vaccine formulations comprising the proteins
or peptides of the
invention themselves. These vaccine formulations may include the use of
adjuvants.
to According to a second aspect of the invention, there is therefore provided:
(i) an
expression cassette containing a DNA molecule of the invention placed under
the control of the
elements required for expression, in particular under the control of an
appropriate promoter; (ii) an
expression vector containing an expression cassette of the invention; (iii) a
prokaryotic or
eukaryotic cell transformed or transfected with an expression cassette and/or
vector of the
15 invention, as well as (iv) a process for producing a polypeptide or
polypeptide derivative encoded
by a polynucleotide of the invention, which involves culturing a prokaryotic
or eukaryotic cell
transformed or transfected with an expression cassette and/or vector of the
invention, under
conditions that allow expression of the DNA molecule of the invention and,
recovering the
encoded polypeptide or polypeptide derivative from the cell culture.
2o A recombinant expression system can be selected from prokaryotic and
eukaryotic hosts.
Eukaryotic hosts include yeast cells (e.g., Saccharomyces cerevisiae or Pichia
pastoris),
mammalian cells (e.g., COS1, NIH3T3, or JEG3 cells), arthropods cells (e.g.,
Spodoptera
frugiperda (SF9) cells), and plant cells. Preferably, a prokaryotic host such
as E. toll is used.
Bacterial and eukaryotic cells are available from a number of different
sources to those skilled in
25 the art, e.g., the American Type Culture Collection (ATCC; Rockville,
Maryland).
The choice of the expression system depends on the features desired for the
expressed
polypeptide. For example, it may be useful to produce a polypeptide of the
invention in a
particular lipidated form or any other form.
The choice of the expression cassette will depend on the host system selected
as well as the
3o features desired for the expressed polypeptide. Typically, an expression
cassette includes a
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WO 00/11183 PCT/IB99/01449
promoter that is functional in the selected host system and can be
constitutive or inducible; a
ribosome binding site; a start codon (ATG) if necessary, a region encoding a
signal peptide, e.g., a
lipidation signal peptide; a DNA molecule of the invention; a stop codon; and
optionally a 3'
terminal region (translation and/or transcription terminator}. The signal
peptide encoding region is
adjacent to the polynucleotide of the invention and placed in proper reading
frame. The signal
peptide-encoding region can be homologous or heterologous to the DNA molecule
encoding the
mature polypeptide and can be specific to the secretion apparatus of the host
used for expression.
The open reading frame constituted by the DNA molecule of the invention,
solely or together with
the signal peptide, is placed under the control of the promoter so that
transcription and translation
occur in the host system. Promoters, signal peptide encoding regions are
widely known and
available to those skilled in the art and includes, for example, the promoter
of Salmonella
typhimurium {and derivatives) that is inducible by arabinose (promoter araB)
and is functional in
Gram-negative bacteria such as E coli (as described in U.S. Patent 5,028,530
and in Cagnon et al.,
(Cagnon et al., Protein Engineering ~: 843 ( 1991 )); the promoter of the gene
of bacteriophage T7
IS encoding RNA polymerase, that is functional in a number of E. coli strains
expressing T7
polymerase (described in U.S. Patent 4,952,496); OspA lipidation signal
peptide; and RIpB
lipidation signal peptide (Takase et al., J. Bact. 169: 5692 (1987)).
The expression cassette is typically part of an expression vector, which is
selected for its
ability to replicate in the chosen expression system. Expression vectors
(e.g., plasmids or viral
vectors) can be chosen from those described in Pouwels et al. (CLONING
VECTORS: LABORATORY
MANUAL, 85, Supp. 1987). They can be purchased from various commercial
sources.
Methods for transforming/transfecting host cells with expression vectors will
depend on the
host system selected as described in Ausubel et al., CURRENT PROTOCOLS IN
MOLECULAR
BIOLOGY, John Wiley & Sons Inc. (1994).
Upon expression, a recombinant polypeptide of the invention (or a polypeptide
derivative)
is produced and remains in the intracellular compartment, is secreted/excreted
in the extracellular
medium or in the periplasmic space, or is embedded in the cellular membrane.
The polypeptide
can then be recovered in a substantially purified form from the cell extract
or from the supernatant
after centrifugation of the recombinant cell culture. Typically, the
recombinant polypeptide can be
3o purified by antibody-based affinity purification or by any other method
that can be readily adapted
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WO 00/11183 PCT/IB99/01449
by a person skilled in the art, such as by genetic fusion to a small affinity
binding domain.
Antibody-based affinity purification methods are also available for purifying
a polypeptide of the
invention extracted from a Chlamydia strain. Antibodies useful for purifying
by immunoaffinity
the polypeptides of the invention can be obtained as described below.
A polynucleotide of the invention can also be useful in the vaccine field,
e.g., for achieving
DNA vaccination. There are two major possibilities, either using a viral or
bacterial host as gene
delivery vehicle (live vaccine vector) or administering the gene in a free
form, e.g., inserted into a
plasmid. Therapeutic or prophylactic efficacy of a polynucleotide of the
invention can be
evaluated as described below.
1 o Accordingly, in a third aspect of the invention, there is provided: (i) a
vaccine vector such
as a poxvirus, containing a DNA molecule of the invention, placed under the
control of elements
required for expression; (ii) a composition of matter containing a vaccine
vector of the invention,
together with a diluent or earner; particularly, (iii) a pharmaceutical
composition containing a
therapeutically or prophylactically effective amount of a vaccine vector of
the invention; (iv) a
15 method for inducing an immune response against Chlamydia in a mammal (e.g.,
a human;
alternatively, the method can be used in veterinary applications for treating
or preventing
Chlamydia infection of animals, e.g., cats or birds), which involves
administering to the mammal
an immunogenically effective amount of a vaccine vector of the invention to
elicit an immune
response, e.g., a protective or therapeutic immune response to Chlamydia; and
particularly, (v) a
2o method for preventing and/or treating a Chlamydia (e.g., C. trachomatis, C.
psittaci,
C. pneumonia, C. pecorum) infection, which involves administering a
prophylactic or therapeutic
amount of a vaccine vector of the invention to an individual in need.
Additionally, the third aspect
of the invention encompasses the use of a vaccine vector of the invention in
the preparation of a
medicament for preventing and/or treating Chlamydia infection.
25 A vaccine vector of the invention can express one or several polypeptides
or derivatives of
the invention, as well as at least one additional Chlamydia antigen, fragment,
homolog, mutant, or
derivative thereof. In addition, it can express a cytokine, such as
interleukin-2 (IL-2) or
interleukin-12 (IL-12), that enhances the immune response (adjuvant effect).
Thus, a vaccine
vector can include an additional DNA sequence encoding, e.g., a chlamydial
antigen , or a
3o cytokine, placed under the control of elements required for expression in a
mammalian cell.
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Alternatively, a composition of the invention can include several vaccine
vectors, each of
them being capable of expressing a polypeptide or derivative of the invention.
A composition can
also contain a vaccine vector capable of expressing an additional Chlamydia
antigen, or a subunit,
fragment, homolog, mutant, or derivative thereof; or a cytokine such as IL-2
or IL-12.
In vaccination methods for treating or preventing infection in a mammal, a
vaccine vector
of the invention can be administered by any conventional route in use in the
vaccine field,
particularly, to a mucosal (e.g., ocular, intranasal, oral, gastric,
pulmonary, intestinal, rectal,
vaginal, or urinary tract) surface or via the parenteral route (e.g.,
subcutaneous, intradermal,
intramuscular, intravenous, or intraperitoneal). Preferred routes depend upon
the choice of the
vaccine vector. The administration can be achieved in a single dose or
repeated at intervals. The
appropriate dosage depends on various parameters understood by skilled
artisans such as the
vaccine vector itself, the route of administration or the condition of the
mammal to be vaccinated
(weight, age and the like).
Live vaccine vectors available in the art include viral vectors such as
adenoviruses,
alphavirus, and poxviruses as well as bacterial vectors, e.g., Shigella,
Salmonella, Vibrio cholerae,
Lactobacillus, Bacille bike de Calmette-Guerin (BCG), and Streptococcus.
An example of an adenovirus vector, as well as a method for constructing an
adenovirus
vector capable of expressing a DNA molecule of the invention, are described in
U.S. Patent
4,920,209. Poxvirus vectors that can be used include, e.g., vaccinia and
canary pox virus,
2o described in U.S. Patent 4,722,848 and U.S. Patent 5,364,773, respectively
(also see, e.g., Tartaglia
et al., Virology 188: 217 (1992)) for a description of a vaccinia virus
vector; and Taylor et al,
Vaccine 13: S39 (1995) for a reference of a canary pox). Poxvirus vectors
capable of expressing a
polynucleotide of the invention can be obtained by homologous recombination as
described in
Kieny et al., Nature 312: 163 ( 1984) so that the polynucleotide of the
invention is inserted in the
viral genome under appropriate conditions for expression in mammalian cells.
Generally, the dose
of vaccine viral vector, for therapeutic or prophylactic use, can be of from
about 1x10" to about
1 x 10", advantageously from about 1 x 10' to about 1 x 10' °,
preferably of from about 1 x 10' to about
1x109 plaque-forming units per kilogram. Preferably, viral vectors are
administered parenterally,
for example, in three doses, four weeks apart. Those skilled in the art
recognize that it is
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WO 00/11183 PCT/IB99/01449
preferable to avoid adding a chemical adjuvant to a composition containing a
viral vector of the
invention and thereby minimizing the immune response to the viral vector
itself.
Non-toxicogenic Vibrio cholerae mutant strains that are useful as a live oral
vaccine are
described in Mekalanos et al., Nature 306: 551 (1983) and U.S. Patent
4,882,278 (strain in which a
substantial amount of the coding sequence of each of the two ctxA alleles has
been deleted so that
no functional cholerae toxin is produced); WO 92/11354 (strain in which the
irgA locus is
inactivated by mutation; this mutation can be combined in a single strain with
ctxA mutations); and
WO 94/1533 (deletion mutant lacking functional ctxA and attRSl DNA sequences).
These strains
can be genetically engineered to express heterologous antigens, as described
in WO 94/19482. An
to effective vaccine dose of a Vibrio cholerae strain capable of expressing a
polypeptide or
polypeptide derivative encoded by a DNA molecule of the invention can contain,
e.g., about 1x105
to about 1 x 1 O9, preferably about 1 x 1 O6 to about 1 x 108 viable bacteria
in an appropriate volume for
the selected route of administration. Preferred routes of administration
include all mucosal routes,
most preferably, these vectors are administered intranasally or orally.
Attenuated Salmonella typhimurium strains, genetically engineered for
recombinant
expression of heterologous antigens or not, and their use as oral vaccines are
described in
Nakayama et al., BiolTechnolog~ 6: 693 (1988) and WO 92/11361. Preferred
routes of
administration include all mucosal routes; most preferably, these vectors are
administered
intranasally or orally.
2o Others bacterial strains useful as vaccine vectors are described in High et
al., EMBO 11:
1991 (1992); Sizemore et al., Science 270: 299 (1995) (ShigellaJlexneri);
Medaglini et al., Proc.
Natl. Acad Sci. USA 92: 6868 (1995) (Streptococcusgordonii); and Flynn, Cell.
Mol. Biol. 40: 31
(1994), WO 88/6626, WO 90/0594, WO 91/13157, WO 92/1796, and WO 92/21376
(Bacille
Calmette Guerin).
In bacterial vectors, polynucleotide of the invention can be inserted into the
bacterial
genome or can remain in a free state, carried on a plasmid.
An adjuvant can also be added to a composition containing a vaccine bacterial
vector. A
number of adjuvants are known to those skilled in the art. Preferred adjuvants
can be selected
from the list provided below.
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WO 00/11183 PCT/IB99/01449
According to a fourth aspect of the invention, there is also provided: (i) a
composition of
matter containing a polynucleotide of the invention, together with a diluent
or carrier; (ii) a
pharmaceutical composition containing a therapeutically or prophylacticaily
effective amount of a
polynucleotide of the invention; (iii) a method for inducing an immune
response against
Chlamydia, in a mammal, by administering to the mammal, an imrnunogenically
effective amount
of a polynucleotide of the invention to elicit an immune response, e.g., a
protective immune
response to Chlamydia; and particularly, (iv) a method for preventing and/or
treating a Chlamydia
(e.g., C. trachomatis, C. psittaci, C. pneumoniae, or C. pecorum) infection,
by administering a
prophylactic or therapeutic amount of a polynucleotide of the invention to an
individual in need.
Additionally, the fourth aspect of the invention encompasses the use of a
polynucleotide of the
invention in the preparation of a medicament for preventing and/or treating
Chlamydia infection.
The fourth aspect of the invention preferably includes the use of a DNA
molecule placed under
conditions for expression in a mammalian cell, e.g., in a plasmid that is
unable to replicate in
mammalian cells and to substantially integrate in a mammalian genome.
Polynucleotides (DNA or RNA) of the invention can also be administered as such
to a
mammal for vaccine, e.g., therapeutic or prophylactic, purpose. When a DNA
molecule of the
invention is used, it can be in the form of a plasmid that is unable to
replicate in a mammalian cell
and unable to integrate in the mammalian genome. Typically, a DNA molecule is
placed under the
control of a promoter suitable for expression in a mammalian cell. The
promoter can function
2o ubiquitously or tissue-specifically. Examples of non-tissue specific
promoters include the early
Cytomegalovirus (CMV) promoter (described in U.S. Patent 4,168,062) and the
Rous Sarcoma
Virus promoter (described in Norton & Coffin, Molec. Cell Biol. ~: 281(1985)).
The desmin
promoter (Li et al., Gene 78: 243 (1989), Li & Paulin, J. Biol. Chem. 266:
6562 (1991), and Li &
Paulin, J. Biol. Chem. 268: 10403 (1993)) is tissue-specific and drives
expression in muscle cells.
More generally, useful vectors are described, i.a., WO 94/21797 and Hartikka
et al., Human Gene
Therapy 7: 1205 (1996).
For DNA/RNA vaccination, the polynucleotide of the invention can encode a
precursor or a
mature form. When it encodes a precursor form, the precursor form can be
homologous or
heterologous. In the latter case, a eukaryotic leader sequence can be used,
such as the leader
3o sequence of the tissue-type plasminogen factor (tPA).
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WO 00/11183 PCT/IB99/01449
A composition of the invention can contain one or several polynucleotides of
the invention.
It can also contain at least one additional polynucleotide encoding another
Chlamydia antigen or a
fragment, derivative, mutant, or analog thereof. A polynucleotide encoding a
cytokine, such as
interleukin-2 (IL-2) or interleukin-12 (IL-12), can also be added to the
composition so that the
immune response is enhanced. These additional polynucleotides are placed under
appropriate
control for expression. Advantageously, DNA molecules of the invention and/or
additional DNA
molecules to be included in the same composition, can be carried in the same
plasmid.
Standard techniques of molecular biology for preparing and purifying
polynucleotides can
be used in the preparation of poiynucleotide therapeutics of the invention.
For use as a vaccine, a
l0 polynucleotide of the invention can be formulated according to various
methods.
First, a polynucleotide can be used in a naked form, free of any delivery
vehicles, such as
anionic liposomes, cationic lipids, microparticles, e.g., gold microparticles,
precipitating agents,
e.g., calcium phosphate, or any other transfection-facilitating agent. In this
case, the
polynucleotide can be simply diluted in a physiologically acceptable solution,
such as sterile saline
15 or sterile buffered saline, with or without a carrier. When present, the
earner preferably is isotonic,
hypotonic, or weakly hypertonic, and has a relatively low ionic strength, such
as provided by a
sucrose solution, e.g., a solution containing 20% sucrose.
Alternatively, a polynucleotide can be associated with agents that assist in
cellular uptake.
It can be, i.a., (i) complemented with a chemical agent that modifies the
cellular permeability, such
2o as bupivacaine (see, e.g., WO 94/16737), (ii) encapsulated into liposomes,
or (iii) associated with
cationic lipids or silica, gold, or tungsten microparticles.
Anionic and neutral liposomes are well-known in the art (see, e.g., LIPOSOMES:
A
PRACTICAL APPROACH, RPC New Ed, IRL press (1990)), for a detailed description
of methods for
making liposomes) and are useful for delivering a. large range of products,
including
25 polynucleotides.
Cationic lipids are also known in the art and are commonly used for gene
delivery. Such
lipids include LipofectinT'~'' also known as DOTMA (N-[1-(2,3-
dioleyloxy)propyl]-N,N,N-
trimethylammonium chloride), DOTAP (1,2-bis(oleyloxy)-3-
(trimethylammonio)propane), DDAB
(dimethyldioctadecylammonium bromide), DOGS (dioctadecylamidologlycyl
spermine) and
3o cholesterol derivatives such as DC-Chol (3 beta-(N-(N',N'-dimethyl
aminomethane)-carbamoyl)
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WO 00/11183 PCT/IB99/01449
cholesterol). A description of these cationic lipids can be found in EP
187,702, WO 90/11092,
U.S. Patent 5,283,185, WO 91/15501, WO 95/26356, and U.S. Patent 5,527,928.
Cationic lipids
for gene delivery are preferably used in association with a neutral lipid such
as DOPE (dioleyl
phosphatidylethanolamine), as described in, e.g., WO 90/11092.
Other transfection-facilitating compounds can be added to a formulation
containing
cationic liposomes. A number of them are described in, e.g., WO 93/18759, WO
93/19768,
WO 94/25608, and WO 95/2397. They include, i.a., spermine derivatives useful
for facilitating
the transport of DNA through the nuclear membrane (see, for example, WO
93/18759) and
membrane-permeabilizing compounds such as GALA, Gramicidine S, and cationic
bile salts (see,
to for example, WO 93/19768).
Gold or tungsten microparticles can also be used for gene delivery, as
described in
WO 91/359, WO 93/17706, and Tang et al. (Nature 356: 152 (1992)). In this
case, the
microparticle-coated polynucleotides can be injected via intradermal or intra-
epidermal routes
using a needleless injection device ("gene gun"), such as those described in
U.S. Patents 4,945,050
15 and 5,015,580, and WO 94/24263.
The amount of DNA to be used in a vaccine recipient depends, e.g., on the
strength of the
promoter used in the DNA construct, the immunogenicity of the expressed gene
product, the
condition of the mammal intended for administration (e.g., the weight, age,
and general health of
the mammal), the mode of administration, and the type of formulation. In
general, a
2o therapeutically or prophylactically effective dose from about 1 ~g to about
1 mg, preferably, from
about 10 pg to about 800 ~g and, more preferably, from about 25 ~g to about
250 fig, can be
administered to human adults. The administration can be achieved in a single
dose or repeated at
intervals.
The route of administration can be any conventional route used in the vaccine
field. As
25 general guidance, a polynucleotide of the invention can be administered via
a mucosal surface,
e.g., an ocular, intranasal, pulmonary, oral, intestinal, rectal, vaginal, and
urinary tract surface; or
via a parenteral route, e.g., by an intravenous, subcutaneous,
intraperitoneal, intradermal, intra-
epidermal, or intramuscular route. The choice of the administration route will
depend on, e.g., the
formulation that is selected. A polynucleotide formulated in association with
bupivacaine is
3o advantageously administered into muscles. When a neutral or anionic
liposome or a cationic lipid,
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WO OO/I1183 PCT/IB99/01449
such as DOTMA or DC-Chol, is used, the formulation can be advantageously
injected via
intravenous, intranasal (aerosolization), intramuscular, intradermal, and
subcutaneous routes. . A
polynucleotide in a naked form can advantageously be administered via the
intramuscular,
intradermal, or subcutaneous routes.
Although not absolutely required, such a composition can also contain an
adjuvant. If so, a
systemic adjuvant that does not require concomitant administration in order to
exhibit an adjuvant
effect is preferable such as, e.g., QS21, which is described in U.S. Patent
5,057,546.
The sequence information provided in the present application enables the
design of specific
nucleotide probes and primers that can be useful in diagnosis. Accordingly, in
a fifth aspect of the
t0 invention, there is provided a nucleotide probe or primer having a sequence
found in or derived by
degeneracy of the genetic code from a sequence shown in SEQ ID NOS: 1, 3, 5,
7, 9, 1 l, 13 and
15.
The term "probe" as used in the present application refers to DNA (preferably
single
stranded) or RNA molecules (or modifications or combinations thereof) that
hybridize under the
15 stringent conditions, as defined above, to nucleic acid molecules having
sequences homologous to
those shown in SEQ ID NOS: 1, 3, 5, 7, 9, 11, 13 and 15, or to a complementary
or anti-sense
sequence. Generally, probes are significantly shorter than full-length
sequences shown in SEQ ID
NOS: 1, 3, 5, 7, 9, 11, 13 and 15; for example, they can contain from about 5
to about 100,
preferably from about 10 to about 80 nucleotides. In particular, probes have
sequences that are at
20 least 75%, preferably at least 85%, more preferably 95% homologous to a
portion of a sequence as
shown in SEQ ID NOS: 1, 3, 5, 7, 9, 11, 13 and 15 or that are complementary to
such sequences.
Probes can contain modified bases such as inosine, methyl-5-deoxycytidine,
deoxyuridine,
dimethylamino-5-deoxyuridine, or diamino-2, 6-purine. Sugar or phosphate
residues can also be
modified or substituted. For example, a deoxyribose residue can be replaced by
a polyamide
25 (Nielsen et al., Science 254: 1497 ( 1991 )) and phosphate residues can be
replaced by ester groups
such as diphosphate, alkyl, arylphosphonate and phosphorothioate esters. In
addition, the
2'-hydroxyl group on ribonucleotides can be modified by including, e.g., alkyl
groups.
Probes of the invention can be used in diagnostic tests, as capture or
detection probes.
Such capture probes can be conventionally immobilized on a solid support,
directly or indirectly,
3o by covalent means or by passive adsorption. A detection probe can be
labelled by a detection
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marker selected from radioactive isotopes; enzymes such as peroxidase,
alkaline phosphatase, and
enzymes able to hydrolyze a chromogenic, fluorogenic, or luminescent
substrate; compounds.that
are chromogenic, fluorogenic, or luminescent; nucleotide base analogs; and
biotin.
Probes of the invention can be used in any conventional hybridization
technique, such as
dot blot (Maniatis et al., MOLECULAR CLONING: A LABORATORY MANUAL (1982) Cold
Spring
Harbor Laboratory Press, Cold Spring Harbor, New York), Southern blot
(Southern, J. Mol. Biol.
98: 503 (1975)), northern blot (identical to Southern blot to the exception
that RNA is used as a
target), or the sandwich technique (Dune et al., Cell 12: 23 ( 1977)). The
latter technique involves
the use of a specific capture probe and/or a specific detection probe with
nucleotide sequences that
I o at least partially differ from each other.
A primer is usually a probe of about 10 to about 40 nucleotides that is used
to initiate
enzymatic polymerization of DNA in an amplification process (e.g., PCR), in an
elongation
process, or in a reverse transcription method. In a diagnostic method
involving PCR, primers can
be labelled.
I S Thus, the invention also encompasses: (i) a reagent containing a probe of
the invention for
detecting and/or identifying the presence of Chlamydia in a biological
material; (ii) a method for
detecting and/or identifying the presence of Chlamydia in a biological
material, in which (a) a
sample is recovered or derived from the biological material, (b) DNA or RNA is
extracted from the
material and denatured, and (c) exposed to a probe of the invention, for
example, a capture,
2o detection probe or both, under stringent hybridization conditions, such
that hybridization is
detected; and (iii) a method for detecting and/or identifying the presence of
Chlamydia in a
biological material, in which (a) a sample is recovered or derived from the
biological material,
(b) DNA is extracted therefrom, (c) the extracted DNA is primed with at least
one, and preferably
two, primers of the invention and amplified by polymerase chain reaction, and
(d) the amplified
25 DNA fragment is produced.
As previously mentioned, polypeptides that can be produced upon expression of
the newly
identified open reading frames are useful vaccine agents.
Therefore, a sixth aspect of the invention features a substantially purified
polypeptide or
polypeptide derivative having an amino acid sequence encoded by a
polynucleotide of the
30 invention.
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A "substantially purified polypeptide" is defined as a polypeptide that is
separated from the
environment in which it naturally occurs and/or that is free of the majority
of the polypeptides that
are present in the environment in which it was synthesized. For example, a
substantially purified
polypeptide is free from cytoplasmic polypeptides. Those skilled in the art
will understand that the
polypeptides of the invention can be purified from a natural source, i.e., a
Chlamydia strain, or can
be produced by recombinant means.
Homologous polypeptides or polypeptide derivatives encoded by polynucleotides
of the
invention can be screened for specific antigenicity by testing cross-
reactivity with an antiserum
raised against the polypeptide of reference having an amino acid sequence as
shown in SEQ ID
NOS: 2, 4, 6, 8, 10, 12, 14 and 16. Briefly, a monospecific hyperimmune
antiserum can be raised
against a purified reference polypeptide as such or as a fusion polypeptide,
for example, an
expression product of MBP, GST, or His-tag systems or a synthetic peptide
predicted to be
antigenic. The homologous polypeptide or derivative screened for specific
antigenicity can be
produced as such or as a fusion polypeptide. In this latter case and if the
antiserum is also raised
against a fusion polypeptide, two different fusion systems are employed.
Specific antigenicity can
be determined according to a number of methods, including Western blot (Towbin
et al., Proc.
Natl. Acad. SCI. USA 76: 4350 (1979)), dot blot, and ELISA, as described
below.
In a Western blot assay, the product to be screened, either as a purified
preparation or a
total E. toll extract, is submitted to SDS-Page electrophoresis as described
by Laemmli, Nature
227: 680 (1970). After transfer to a nitrocellulose membrane, the material is
further incubated
with the monospecific hyperimmune antiserum diluted in the range of dilutions
from about 1:5 to
about 1:5000, preferably from about 1:100 to about 1:500. Specific
antigenicity is shown once a
band corresponding to the product exhibits reactivity at any of the dilutions
in the above range.
In an ELISA assay, the product to be screened is preferably used as the
coating antigen. A
purified preparation is preferred, although a whole cell extract can also be
used. Briefly, about
100 pl of a preparation at about 10 pg protein/ml are distributed into wells
of a 96-well
polycarbonate ELISA plate. The plate is incubated for 2 hours at 37°C
then overnight at 4°C. The
plate is washed with phosphate buffer saline (PBS) containing 0.05% Tween 20
(PBS/Tween
buffer). The wells are saturated with 250 ~.l PBS containing 1% bovine serum
albumin (BSA) to
3o prevent non-specific antibody binding. After 1 hour incubation at
37°C, the plate is washed with
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PBS/Tween buffer. The antiserum is serially diluted in PBS/Tween buffer
containing 0.5% BSA.
100 ~1 of dilutions are added per well. The plate is incubated for 90 minutes
at 37°C, washed_and
evaluated according to standard procedures. For example, a goat anti-rabbit
peroxidase conjugate
is added to the wells when specific antibodies were raised in rabbits.
Incubation is carried out for
90 minutes at 37°C and the plate is washed. The reaction is developed
with the appropriate
substrate and the reaction is measured by colorimetry (absorbance measured
spectrophotometrically). Under the above experimental conditions, a positive
reaction is shown by
O.D. values greater than a non immune control serum.
In a dot blot assay, a purified product is preferred, although a whole cell
extract can also be
to used. Briefly, a solution of the product at about 100 ~g/ml is serially two-
fold diluted in SO mM
Tris-HC1 (pH 7.5). 100 ~1 of each dilution are applied to a nitrocellulose
membrane 0.45 ~.m set in
a 96-well dot blot apparatus (Biorad). The buffer is removed by applying
vacuum to the system.
Wells are washed by addition of 50 mM Tris-HCl (pH 7.5) and the membrane is
air-dried. The
membrane is saturated in blocking buffer (50 mM Tris-HCl (pH 7.5) 0. i 5 M
NaCI, 10 g/L skim
milk) and incubated with an antiserum dilution from about I :50 to about
1:5000, preferably about
1:500. The reaction is revealed according to standard procedures. For example,
a goat anti-rabbit
peroxidase conjugate is added to the wells when rabbit antibodies are used.
Incubation is earned
out 90 minutes at 37°C and the blot is washed. The reaction is
developed with the appropriate
substrate and stopped. The reaction is measured visually by the appearance of
a colored spot, e.g.,
2o by colorimetry. Under the above experimental conditions, a positive
reaction is shown once a
colored spot is associated with a dilution of at least about 1:5, preferably
of at least about I :500.
Therapeutic or prophylactic efficacy of a polypeptide or derivative of the
invention can be
evaluated as described below.
According to a seventh aspect of the invention, there is provided: (i) a
composition of
matter containing a polypeptide of the invention together with a diluent or
earner; in particular,
(ii) a pharmaceutical composition containing a therapeutically or
prophylactically effective amount
of a polypeptide of the invention; (iii) a method for inducing an immune
response against
Chlamydia in a mammal, by administering to the mammal an immunogenically
effective amount
of a polypeptide of the invention to elicit an immune response, e.g., a
protective immune response
3o to Chlamydia; and particularly, (iv) a method for preventing and/or
treating a Chlamydia (e.g.,
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WO 00/11183 PCT/IB99/01449
C. trachomatis. C. psittaci, C. pneumoniae. or C. pecorum) infection, by
administering a
prophylactic or therapeutic amount of a polypeptide of the invention to an
individual in need..
Additionally, the seventh aspect of the invention encompasses the use of a
polypeptide of the
invention in the preparation of a medicament for preventing and/or treating
Chlamydia infection.
The immunogenic compositions of the invention can be administered by any
conventional
route in use in the vaccine field, in particular to a mucosal (e.g., ocular,
intranasal, pulmonary, oral,
gastric, intestinal, rectal, vaginal, or urinary tract) surface or via the
parenteral (e.g., subcutaneous,
intradermal, intramuscular, intravenous, or intraperitoneal) route. The choice
of the administration
route depends upon a number of parameters, such as the adjuvant associated
with the polypeptide.
1o For example, if a mucosal adjuvant is used, the intranasal or oral route
will be preferred and if a
lipid formulation or an aluminum compound is used, the parenteral route will
be preferred. In the
latter case, the subcutaneous or intramuscular route is most preferred. The
choice can also depend
upon the nature of the vaccine agent. For example, a polypeptide of the
invention fused to CTB or
LTB will be best administered to a mucosal surface.
A composition of the invention can contain one or several polypeptides or
derivatives of
the invention. It can also contain at least one additional Chlamydia antigen,
or a subunit, fragment,
homolog, mutant, or derivative thereof.
For use in a composition of the invention, a polypeptide or derivative thereof
can be
formulated into or with liposomes, preferably neutral or anionic liposomes,
microspheres,
2o ISCOMS, or virus-like-particles (VLPs) to facilitate delivery and/or
enhance the immune response.
These compounds are readily available to one skilled in the art; for example,
see LIPOSOMES: A
PRACTICAL APPROACH (supra).
Adjuvants other than liposomes and the like can also be used and are known in
the art. An
appropriate selection can conventionally be made .by those skilled in the art,
for example, from the
list provided below.
Administration can be achieved in a single dose or repeated as necessary at
intervals as can
be determined by one skilled in the art. For example, a priming dose can be
followed by three
booster doses at weekly or monthly intervals. An appropriate dose depends on
various parameters
including the recipient (e.g., adult or infant), the particular vaccine
antigen, the route and
3o frequency of administration, the presence/absence or type of adjuvant, and
the desired effect (e.g.,
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WO 00/11183 PCT/IB99/01449
protection and/or treatment), as can be determined by one skilled in the art.
In general, a vaccine
antigen of the invention can be administered by a mucosal route in an amount
from about 10 ~g to
about 500 mg, preferably from about 1 mg to about 200 mg. For the parenteral
route of
administration, the dose usually should not exceed about 1 mg, preferably
about 100 pg.
s When used as vaccine agents, polynucleotides and polypeptides of the
invention can be
used sequentially as part of a multistep immunization process. For example, a
mammal can be
initially primed with a vaccine vector of the invention such as a pox virus,
e.g., via the parenteral
route, and then boosted twice with the polypeptide encoded by the vaccine
vector, e.g., via the
mucosal route. In another example, liposomes associated with a polypeptide or
derivative of the
to invention can also be used for priming, with boosting being carned out
mucosally using a soluble
polypeptide or derivative of the invention in combination with a mucosal
adjuvant (e.g., LT).
A polypeptide derivative of the invention is also useful as a diagnostic
reagent for detecting
the presence of anti-Chlamydia antibodies, e.g., in a blood sample. Such
polypeptides are about 5
to about 80, preferably about 10 to about 50 amino acids in length and can be
labeled or unlabeled,
I5 depending upon the diagnostic method. Diagnostic methods involving such a
reagent are
described below.
Upon expression of a DNA molecule of the invention, a polypeptide or
polypeptide
derivative is produced and can be purified using known laboratory techniques.
For example, the
polypeptide or polypeptide derivative can be produced as a fusion protein
containing a fused tail
2o that facilitates purification. The fusion product can be used to immunize a
small mammal, e.g., a
mouse or a rabbit, in order to raise antibodies against the polypeptide or
polypeptide derivative
(monospecific antibodies). The eighth aspect of the invention thus provides a
monospecific
antibody that binds to a polypeptide or polypeptide derivative of the
invention.
By "monospecific antibody" is meant an antibody that is capable of reacting
with a unique
25 naturally-occurring Chlamydia polypeptide. An antibody of the invention can
be polyclonal or
monoclonal. Monospecific antibodies can be recombinant, e.g., chimeric (e.g.,
constituted by a
variable region of marine origin associated with a human constant region),
humanized (a-human
immunoglobulin constant backbone together with hypervariable region of animal,
e.g., marine,
origin), and/or single chain. Both polyclonal and monospecific antibodies can
also be in the form
30 of immunoglobulin fragments, e.g., F(ab)'2 or Fab fragments. The antibodies
of the invention can
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WO 00/11183 PCT/IB99/01449
be of any isotype, e.g., IgG or IgA, and polyclonal antibodies can be of a
single isotype or can
contain a mixture of isotypes.
The antibodies of the invention, which are raised to a polypeptide or
polypeptide derivative
of the invention, can be produced and identified using standard immunological
assays, e.g.,
Western blot analysis, dot blot assay, or ELISA (see, e.g., Coligan et al.,
CultttirNT PROTOCOLS 1N
IMMUNOLOGY (1994) John Wiley & Sons, Inc., New York, NY). The antibodies can
be used in
diagnostic methods to detect the presence of a Chlamydia antigen in a sample,
such as a biological
sample. The antibodies can also be used in affinity chromatography methods for
purifying a
polypeptide or polypeptide derivative of the invention. As is discussed
further below, such
t 0 antibodies can be used in prophylactic and therapeutic passive
immunization methods.
Accordingly, a ninth aspect of the invention provides: (i) a reagent for
detecting the
presence of Chlamydia in a biological sample that contains an antibody,
polypeptide, or
polypeptide derivative of the invention; and {ii) a diagnostic method for
detecting the presence of
Chlamydia in a biological sample, by contacting the biological sample with an
antibody, a
polypeptide, or a polypeptide derivative of the invention, such that an immune
complex is formed,
and by detecting such complex to indicate the presence of Chlamydia in the
sample or the
organism from which the sample is derived.
Those skilled in the art will understand that the immune complex is formed
between a
component of the sample and the antibody, polypeptide, or polypeptide
derivative, whichever is
2o used, and that any unbound material can be removed prior to detecting the
complex. As can be
easily understood, a polypeptide reagent is useful for detecting the presence
of anti-Chlamydia
antibodies in a sample, e.g., a blood sample, while an antibody of the
invention can be used for
screening a sample, such as a gastric extract or biopsy, for the presence of
Chlamydia
polypeptides.
For use in diagnostic applications, the reagent (i.e., the antibody,
polypeptide, or
polypeptide derivative of the invention) can be in a free state or immobilized
on a solid support,
such as a tube, a bead, or any other conventional support used in the field.
Immobilization can be
achieved using direct or indirect means. Direct means include passive
adsorption (non-covalent
binding) or covalent binding between the support and the reagent. By "indirect
means" is meant
3o that an anti-reagent compound that interacts with a reagent is first
attached to the solid support.
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WO 00/11183 PCT/IB99/01449
For example, if a polypeptide reagent is used, an antibody that binds to it
can serve as an
anti-reagent, provided that it binds to an epitope that is not involved in the
recognition of
antibodies in biological samples. Indirect means can also employ a ligand-
receptor system, for
example, a molecule such as a vitamin can be grafted onto the polypeptide
reagent and the
corresponding receptor can be immobilized on the solid phase. This is
illustrated by the
biotin-streptavidin system. Alternatively, indirect means can be used, e.g.,
by adding to the
reagent a peptide tail, chemically or by genetic engineering, and immobilizing
the grafted or fused
product by passive adsorption or covalent linkage of the peptide tail.
According to a tenth aspect of the invention, there is provided a process for
purifying, from
to a biological sample, a polypeptide or polypeptide derivative of the
invention, which involves
carrying out antibody-based affinity chromatography with the biological
sample, wherein the
antibody is a monospecific antibody of the invention.
For use in a purification process of the invention, the antibody can be
polyclonal or
monospecific, and preferably is of the IgG type. Purified IgGs can be prepared
from an antiserum
t5 using standard methods (see, e.g., Coligan et al., supra). Conventional
chromatography supports,
as well as standard methods for grafting antibodies, are disclosed in, e.g.,
ANTIBODIES: A
LABORATORY MANUAL, D. Lane, E. Harlow, Eds. ( 1988).
Briefly, a biological sample, such as an C. pneumoniae extract, preferably in
a buffer
solution, is applied to a chromatography material, preferably equilibrated
with the buffer used to
2o dilute the biological sample so that the polypeptide or polypeptide
derivative of the invention (i.e.,
the antigen) is allowed to adsorb onto the material. The chromatography
material, such as a gel or
a resin coupled to an antibody of the invention, can be in batch form or in a
column. The unbound
components are washed off and the antigen is then eluted with an appropriate
elution buffer, such
as a glycine buffer or a buffer containing a chaotropic agent, e.g., guanidine
HCI, or high salt
25 concentration (e.g., 3 M MgCl2). Eluted fractions are recovered and the
presence of the antigen is
detected, e.g., by measuring the absorbance at 280 nm.
An antibody of the invention can be screened for therapeutic efficacy as
described as
follows. According to an eleventh aspect of the invention, there is provided:
(i) a composition of
matter containing a monospecific antibody of the invention, together with a
diluent or carrier; (ii) a
3o pharmaceutical composition containing a therapeutically or prophylactically
effective amount of a
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WO 00/11183 PCT/IB99/01449
monospecific antibody of the invention, and (iii) a method for treating or
preventing a Chlamydia
(e.g., C. trachomatis, C. psittaci, C. pneumoniae or C. pecorum) infection, by
administering a
therapeutic or prophylactic amount of a monospecific antibody of the invention
to an individual in
need. Additionally, the eleventh aspect of the invention encompasses the use
of a monospecific
antibody of the invention in the preparation of a medicament for treating or
preventing Chlamydia
infection.
To this end, the monospecific antibody can be polyclonal or monoclonal,
preferably of the
IgA isotype (predominantly). In passive immunization, the antibody can be
administered to a
mucosal surface of a mammal, e.g., the gastric mucosa, e.g., orally or
intragastrically,
advantageously, in the presence of a bicarbonate buffer. Alternatively,
systemic administration,
not requiring a bicarbonate buffer, can be carried out. A monospecific
antibody of the invention
can be administered as a single active component or as a mixture with at least
one monospecific
antibody specific for a different Chlamydia polypeptide. The amount of
antibody and the
particular regimen used can be readily determined by one skilled in the art.
For example, daily
15 administration of about 100 to 1,000 mg of antibodies over one week, or
three doses per day of
about 100 to 1,000 mg of antibodies over two or three days, can be an
effective regimens for most
purposes.
Therapeutic or prophylactic efficacy can be evaluated using standard methods
in the art,
e.g., by measuring induction of a mucosal immune response or induction of
protective and/or
2o therapeutic immunity, using, e.g., the C. pneumoniae mouse model. Those
skilled in the art will
recognize that the C. pneumoniae strain of the model can be replaced with
another Chlamydia
strain. For example, the efficacy of DNA molecules and polypeptides from C.
pneumoniae is
preferably evaluated in a mouse model using an C. pneumoniae strain.
Protection can be
determined by comparing the degree of Chlamydia infection to that of a control
group. Protection
25 is shown when infection is reduced by comparison to the control group. Such
an evaluation can be
made for polynucleotides, vaccine vectors, polypeptides and derivatives
thereof, as well as
antibodies of the invention.
Adjuvants useful in any of the vaccine compositions described above are as
follows.
Adjuvants for parenteral administration include aluminum compounds, such as
aluminum
3o hydroxide, aluminum phosphate, and aluminum hydroxy phosphate. The antigen
can be
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WO 00/11183 PCT/IB99/01449
precipitated with, or adsorbed onto, the aluminum compound according to
standard protocols.
Other adjuvants, such as RIBI (ImmunoChem, Hamilton, MT), can be used in
parenteral
administration.
Adjuvants for mucosal administration include bacterial toxins, e.g., the
cholera toxin (CT),
the E. coli heat-labile toxin (LT), the Clostridium di~cile toxin A and the
pertussis toxin (PT), or
combinations, subunits, toxoids, or mutants thereof. For example, a purified
preparation of native
cholera toxin subunit B (CTB) can be of use. Fragments, homologs, derivatives,
and fusions to
any of these toxins are also suitable, provided that they retain adjuvant
activity. Preferably, a
mutant having reduced toxicity is used. Suitable mutants are described, e.g.,
in WO 95/17211
(Arg-7-Lys CT mutant), WO 96/6627 (Arg-192-Gly LT mutant), and WO 95/34323
(Arg-9-Lys
and Glu-129-Gly PT mutant). Additional LT mutants that can be used in the
methods and
compositions of the invention include, e.g., Ser-63-Lys, Ala-69-GIy, Glu-110-
Asp, and
Glu-112-Asp mutants. Other adjuvants, such as a bacterial monophosphoryl lipid
A (MPLA) of,
e.g., E. coli, Salmonella minnesota, Salmonella typhimurium, or Shigella
flexneri; saponins, or
I5 polylactide glycolide (PLGA) microspheres, can also be used in mucosal
administration.
Adjuvants useful for both mucosal and parenteral administrations include
polyphosphazene
(WO 95/2415), DC-chol (3 b-(N-(N',N'-dimethyl aminomethane)-carbamoyl)
cholesterol (U.S.
Patent 5,283,185 and WO 96/14831) and QS-21 (WO 88/9336).
Any pharmaceutical composition of the invention, containing a polynucleotide,
a
polypeptide, a polypeptide derivative, or an antibody of the invention, can be
manufactured in a
conventional manner. In particular, it can be formulated with a
pharmaceutically acceptable
diluent or carrier, e.g., water or a saline solution such as phosphate buffer
saline. In general, a
diluent or carrier can be selected on the basis of the mode and route of
administration, and standard
pharmaceutical practice. Suitable pharmaceutical Garners or diluents, as well
as pharmaceutical
necessities for their use in pharmaceutical formulations, are described in
Remington's
Pharmaceutical Sciences, a standard reference text in this field and in the
USP/NF.
The invention also includes methods in which Chlamydia infection, are treated
by~ oral
administration of a Chlamydia polypeptide of the invention and a mucosal
adjuvant, in
combination with an antibiotic, an antacid, sucralfate, or a combination
thereof. Examples of such
compounds that can be administered with the vaccine antigen and the adjuvant
are antibiotics,
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WO 00/11183 PCT/IB99/01449
including, e.g., macrolides, tetracyclines, and derivatives thereof (specific
examples of antibiotics
that can be used include azithromycin or doxicyclin or immunomodulators such
as cytokines-or
steroids. In addition, compounds containing more than one of the above-listed
components
coupled together, can be used. The invention also includes compositions for
carrying out these
methods, i.e., compositions containing a Chlamydia antigen (or antigens) of
the invention, an
adjuvant, and one or more of the above-listed compounds, in a pharmaceutically
acceptable carrier
or diluent.
Amounts of the above-listed compounds used in the methods and compositions of
the
invention can readily be determined by one skilled in the art. In addition,
one skilled in the art can
1o readily design treatment/immunization schedules. For example, the non-
vaccine components can
be administered on days 1-14, and the vaccine antigen + adjuvant can be
administered on days 7,
14, 21, and 28.
The above disclosure generally describes the present invention. A more
complete
understanding can be obtained by reference to the following specific examples.
These examples
15 are described solely for purposes of illustration and are not intended to
limit the scope of the
invention. Changes in form and substitution of equivalents are contemplated as
circumstances
may suggest or render expedient. Although specific terms have been employed
herein, such terms
are intended in a descriptive sense and not for purposes of limitation.
EQUIVALENTS
20 From the foregoing detailed description of the specific embodiments of the
invention, it
should be apparent that unique Chlamydia antigens have been described.
Although particular
embodiments have been disclosed herein in detail, this has been done by way of
example for
purposes of illustration only, and is not intended to be limiting with respect
to the scope of the
appended claims which follow. In particular, it is contemplated by the
inventor that various
25 substitutions, alterations, and modifications may be made to the invention
without departing from
the spirit and scope of the invention as defined by the claims.
-34-
CA 02341637 2001-02-19
WO 00/11183 PCT/IB99/01449
SEQUENCE LISTING
<110> Murdin, Andrew
<120> CFiLAMYDIA ANTIGENS AND CORRESPONDING DNA FRAGMENTS
THEREOF
<130> Chlamydia antigens
<140>
<141>
<160> 16
<170> Patentln Ver. 2.0
<210> 1
<211> 2550
<212> DNA
<213> Chlamydia sp.
<220>
<221> CDS
<222> (101)..(2461)
<400> 1
taggtcttgt taaaaaactt ctttgtccct atttttgtgc ttacctcatt atctgataaa 60
caagctttct atctagggaa gactcttggg aatgctcatc atg cga aat aaa gtt 115
Met Arg Asn Lys Val
1 5
atc ttg caa ata tct att cta gcg tta atc caa acc cct tta act tta 163
Ile Leu Gln Ile Ser Ile Leu Ala Leu Ile Gln Thr Pro Leu Thr Leu
10 15 20
ttt tct act gaa aaa gtt aaa gaa ggc cat gtg gtg gta gac tct atc 211
Phe Ser Thr Glu Lys Val Lys Glu Gly His Val Val Val Asp Ser Ile
25 30 35
aca atc ata acg gaa gga gaa aat get tca aat aaa cat ccc tta ccc 259
Thr Ile Ile Thr Glu Gly Glu Asn Ala Ser Asn Lys His Pro Leu Pro
40 45 50
aaa tta aag acc aga agt ggg get ctt ttt tct caa tta gat ttt gat 307
Lys Leu Lys Thr Arg Ser Gly Ala Leu Phe Ser Gln Leu Asp Phe Asp
55 60 65
1
CA 02341637 2001-02-19
WO 00/11183 PCT/IB99/01449
gaa ttg attcta aaa tac gactctgtt gagcct aaa 355
gac aga get gaa
Glu Leu IleLeu Lys Tyr AspSerVal GluPro Lys
Asp Arg Ala Glu
70 75 80 85
gta ttt gaaggg act ata gcccttcac ctaata get 403
gaa tct aaa aac
Val Phe GluGly Thr Ile AlaLeuHis LeuIle Ala
Glu Ser Lys Asn
90 95 100
aaa ccc tca att cga aat att cat atc tca gga aat caa gtc gtt cct 451
Lys Pro Ser Ile Arg Asn Ile His Ile Ser Gly Asn Gln Val Val Pro
105 110 115
gaa cat aaa att ctt aaa acc cta caa att tac cgt aat gat ctc ttt 499
Glu His Lys Ile Leu Lys Thr Leu Gln Ile Tyr Arg Asn Asp Leu Phe
120 125 130
gaa cga gaa aaa ttt ctt aag ggt ctt gat gat cta aga acg tat tat 547
Glu Arg Glu Lys Phe Leu Lys Gly Leu Asp Asp Leu Arg Thr Tyr Tyr
135 140 145
ctc aag cga gga tat ttc gca tcc agt gta gac tac agt ctg gaa cac 595
Leu Lys Arg Gly Tyr Phe Ala Ser Ser Val Asp Tyr Sex Leu Glu His
150 155 160 165
aat caa gaa aaa ggt cac atc gat gtt tta att aaa atc aat gaa ggt 643
Asn Gln Glu Lys Gly His Ile Asp Val Leu Ile Lys Ile Asn Glu Gly
170 175 180
cct tgc ggg aaa att aaa cag ctt acg ttc tca gga atc tct cga tca 691
Pro Cys Gly Lys Ile Lys Gln Leu Thr Phe Ser Gly Ile Ser Arg Ser
185 190 195
gaa aaa tca gat atc caa gaa ttt att caa acc aag cag cac tct aca 739
Glu Lys Ser Asp Ile Gln Glu Phe Ile Gln Thr Lys Gln His Ser Thr
200 205 210
act aca agt tgg ttt act gga get gga ctc tat cac cca gat att gtt 787
Thr Thr Ser Trp Phe Thr Gly Ala Gly Leu Tyr His Pro Asp Ile Val
215 220 225
gaa caa gat agc ttg gca att acg aat tac cta cat aat aac ggg tac 835
Glu Gln Asp Ser Leu Ala Ile Thr Asn Tyr Leu His Asn Asn Gly Tyr
230 235 240 245
get gat get ata gtc aac tct cac tat gac ctt gac gac aaa ggg aat 883
Ala Asp Ala Ile Val Asn Ser His Tyr Asp Leu Asp Asp Lys Gly Asn
250 255 260
2
CA 02341637 2001-02-19
WO 00/11183 PCT/IB99/01449
att ctt ctt tac atg gat att gat cga ggg tcg cga tat acc tta gga 931
Ile Leu Leu Tyr Met Asp Ile Asp Arg Gly Ser Arg Tyr Thr Leu Gly
265 270 275
cac gtc cat atc caa ggg ttt gag gtt ttg cca aaa cgc ctt ata gaa 979
His Val His Ile Gln Gly Phe Glu Val Leu Pro Lys Arg Leu Ile Glu
280 285 290
aag caa tcc caa gtc ggc ccc aat gat ctt tat tgc ccc gat aaa ata 1027
Lys Gln Ser Gln Val Gly Pro Asn Asp Leu Tyr Cys Pro Asp Lys Ile
2g5 300 305
tgg gat ggg get cat aag atc aaa caa act tat gca aag tat ggc tac 1075
Trp Asp Gly Ala His Lys Ile Lys Gln Thr Tyr Ala Lys Tyr Gly Tyr
310 315 320 325
atc aat acc aat gta gac gtt ctc ttc atc cct cac gca acc cgc cct 1123
Ile Asn Thr Asn Val Asp Val Leu Phe Ile Pro His Ala Thr Arg Pro
330 335 340
att tat gat gta act tat gag gta agt gaa ggg tct cct tat aaa gtt 1171
Ile Tyr Asp Val Thr Tyr Glu Val Ser Glu Gly Ser Pro Tyr Lys Val
345 350 355
ggg tta att aaa att act ggg aat acc cat aca aaa tct gac gtt att 1219
Gly Leu Ile Lys Ile Thr Gly Asn Thr His Thr Lys Ser Asp Val Ile
360 365 370
tta cac gaa acc agt ctc ttc cca gga gat aca ttc aat cgc tta aag 1267
Leu His Glu Thr Ser Leu Phe Pro Gly Asp Thr Phe Asn Arg Leu Lys
375 380 385
cta gaa gat act gag caa cgt tta aga aat aca ggc tac ttc caa agc 1315
Leu Glu Asp Thr Glu Gln Arg Leu Arg Asn Thr Gly Tyr Phe Gln Ser
390 395 400 405
gtt agt gtc tat aca gtt cgt tct caa ctt gat cct atg ggc aat gcg 1363
Val Ser Val Tyr Thr Val Arg Ser Gln Leu Asp Pro Met Gly Asn Ala
410 415 420
gat caa tac cga gat att ttt gta gaa gtc aaa gaa aca aca aca gga 1411
Asp Gln Tyr Arg Asp Ile Phe Val Glu Val Lys Glu Thr Thr Thr Gly
425 430 435
aac tta ggc tta ttc tta gga ttt agt tct ctt gac aat ctt ttt gga 1459
Asn Leu Gly Leu Phe Leu Gly Phe Ser Ser Leu Asp Asn Leu Phe Gly
440 445 450
3
CA 02341637 2001-02-19
WO 00/11183 PCT/IB99/01449
gga att gaa cta tct gaa agt aat ttt gat cta ttt gga get aga aat 1507
Gly Ile Glu Leu Ser Glu Ser Asn Phe Asp Leu Phe Gly Ala Arg Asn
455 460 465
ata ttt tct aaa ggt ttt cgt tgt cta aga ggc ggt gga gaa cat cta 1555
Ile Phe Ser Lys Gly Phe Arg Cys Leu Arg Gly Gly Gly Glu His Leu
470 475 480 485
ttc tta aaa gcc aac ttc ggg gac aaa gtc aca gac tat act ttg aag 1603
Phe Leu Lys Ala Asn Phe Gly Asp Lys Val Thr Asp Tyr Thr Leu Lys
490 495 500
tgg acc aaa cct cat ttt cta aac act cct tgg att tta gga att gaa 1651
Trp Thr Lys Pro His Phe Leu Asn Thr Pro Trp Ile Leu Gly Ile Glu
505 510 515
tta gat aaa tca att aac aga gca tta tct aaa gat tat get gtc caa 1699
Leu Asp Lys Ser Ile Asn Arg Ala Leu Ser Lys Asp Tyr Ala Val Gln
520 525 530
acc tat ggc ggg aac gtc agc aca acg tat atc ttg aac gaa cac ctg 1747
Thr Tyr Gly Gly Asn Val Ser Thr Thr Tyr Ile Leu Asn Glu His Leu
535 540 545
aaa tac ggt cta ttt tat cga gga agt caa acg agt tta cat gaa aaa 1795
Lys Tyr Gly Leu Phe Tyr Arg Gly Ser Gln Thr Ser Leu His Glu Lys
550 555 560 565
cgt aag ttc ctc cta ggg cca aat ata gac agc aat aaa gga ttt gtc 1843
Arg Lys Phe Leu Leu Gly Pro Asn Ile Asp Ser Asn Lys Gly Phe Val
570 575 580
tct get gca ggt gtc aac ttg aat tac gat tct gta gat agt cct aga 1891
Ser Ala Ala Gly Val Asn Leu Asn Tyr Asp Ser Val Asp Ser Pro Arg
585 590 595
act cca act aca ggg att cgc ggg ggg gtg act ttt gag gtt tct ggt 1939
Thr Pro Thr Thr Gly Ile Arg Gly Gly Val Thr Phe Glu Val Ser Gly
600 605 610
ttg gga gga act tat cat ttt aca aaa ctc tct tta aac agc tct atc 1987
Leu Gly Gly Thr Tyr His Phe Thr Lys Leu Ser Leu Asn Ser Ser Ile
615 620 625
tat aga aaa ctt acg cgt aaa ggt att ttg aaa atc aaa ggg gaa get 2035
Tyr Arg Lys Leu Thr Arg Lys Gly Ile Leu Lys Ile Lys Gly Glu Ala
630 635 640 645
4
CA 02341637 2001-02-19
WO 00/11183 PCT/IB99/01449
caa ttt att aaa ece tat agc aat act aca get gaa gga gat ect gtc 2083
Gln Phe Ile Lys Pro Tyr Ser Asn Thr Thr Ala Glu Gly Asp Pro Val
650 655 660
agt gag cgc ttc ttc cta ggt gga gag act aca gtt cgg gga tat aaa 2131
Ser Glu Arg Phe Phe Leu Gly Gly Glu Thr Thr Val Arg Gly Tyr Lys
665 670 675
tcc ttt att atc ggt cca aaa tac tct get aca gaa cct cag gga gga 2179
Ser Phe Ile Ile Gly Pro Lys Tyr Ser Ala Thr Glu Pro Gln Gly Gly
680 685 690
ctc tct tcg ctc ctt att tca gaa gag ttt caa tac cct ctc atc aga 2227
Leu Ser Ser Leu Leu Ile Ser Glu Glu Phe Gln Tyr Pro Leu Ile Arg
695 700 705
caa cct aat att agt gcc ttt gta ttc tta gac tca ggt ttt gtc ggt 2275
Gln Pro Asn Ile Ser Ala Phe Val Phe Leu Asp Ser Gly Phe Val Gly
710 715 720 725
tta caa gag tat aag att tcg tta aaa gat cta cgt agt agt get gga 2323
Leu Gln Glu Tyr Lys Ile Ser Leu Lys Asp Leu Arg Ser Ser Ala Gly
730 735 740
ttt ggt ctg cgc ttc gat gta atg aat aat gtt cct gtt atg tta gga 2371
Phe Gly Leu Arg Phe Asp Val Met Asn Asn Val Pro Val Met Leu Gly
745 750 755
ttt ggt tgg ccc ttc cgt cca acc gag act ttg aat gga gaa aaa att 2419
Phe Gly Trp Pro Phe Arg Pro Thr Glu Thr Leu Asn Gly Glu Lys Ile
760 765 770
gat gta tct cag cga ttc ttc ttt get tta ggg ggc atg ttc 2461
Asp Val Ser Gln Arg Phe Phe Phe Ala Leu Gly Gly Met Phe
775 780 785
taagatataa attaaggact tatcgaagga aatctttgtt gttttcagaa aaggcttttg 2521
gtaccctttt tcctataccc aaagttagt 2550
<210> 2
<211> 787
<212> PRT
<213> Chlamydia sp.
<400> 2
Met Arg Asn Lys Val Ile Leu Gln Ile Ser Ile Leu Ala Leu Ile Gln
5
CA 02341637 2001-02-19
WO 00/11183 PCT/1B99/01449
1 5 10 15
Thr Pro Leu Thr Leu Phe Ser Thr Glu Lys Val Lys Glu Gly His Val
20 25 30
Val Val Asp Ser Ile Thr Ile Ile Thr Glu Gly Glu Asn Ala Ser Asn
35 40 45
Lys His Pro Leu Pro Lys Leu Lys Thr Arg Ser Gly Ala Leu Phe Ser
50 55 60
Gln Leu Asp Phe Asp Glu Asp Leu Arg Ile Leu Ala Lys Glu Tyr Asp
65 70 75 80
Ser Val Glu Pro Lys Val Glu Phe Ser Glu Gly Lys Thr Asn Ile Ala
85 90 95
Leu His Leu Ile Ala Lys Pro Ser Ile Arg Asn Ile His Ile Ser Gly
100 105 110
Asn Gln Val Val Pro Glu His Lys Ile Leu Lys Thr Leu Gln Ile Tyr
115 120 125
Arg Asn Asp Leu Phe Glu Arg Glu Lys Phe Leu Lys Gly Leu Asp Asp
130 135 140
Leu Arg Thr Tyr Tyr Leu Lys Arg Gly Tyr Phe Ala Ser Ser Val Asp
145 150 155 160
Tyr Ser Leu Glu His Asn Gln Glu Lys Gly His Ile Asp Val Leu Ile
165 170 175
Lys Ile Asn Glu Gly Pro Cys Gly Lys Ile Lys Gln Leu Thr Phe Ser
180 185 190
Gly Ile Ser Arg Ser Glu Lys Ser Asp Ile Gln Glu Phe Ile Gln Thr
195 200 205
Lys Gln His Ser Thr Thr Thr Ser Trp Phe Thr Gly Ala Gly Leu Tyr
2I0 215 220
His Pro Asp Ile Val Glu Gln Asp Ser Leu Ala Ile Thr Asn Tyr Leu
225 230 235 240
His Asn Asn Gly Tyr Ala Asp Ala Ile Val Asn Ser His Tyr Asp Leu
245 250 255
Asp Asp Lys Gly Asn Ile Leu Leu Tyr Met Asp Ile Asp Arg Gly Ser
6
CA 02341637 2001-02-19
WO 00/11183 PCT/IB99/01449
260 265 270
Arg Tyr Thr Leu Gly His Val His Ile Gln Gly Phe Glu Val Leu Pro
275 280 285
Lys Arg Leu Ile Glu Lys Gln Ser Gln Val Gly Pro Asn Asp Leu Tyr
290 295 300
Cys Pro Asp Lys Ile Trp Asp Gly Ala His Lys Ile Lys Gln Thr Tyr
305 310 315 320
Ala Lys Tyr Gly Tyr Ile Asn Thr Asn Val Asp Val Leu Phe Ile Pro
325 330 335
His Ala Thr Arg Pro Ile Tyr Asp Vai Thr Tyr Glu Val Ser Glu Gly
340 345 350
Ser Pro Tyr Lys Val Gly Leu Ile Lys Ile Thr Gly Asn Thr His Thr
355 360 365
Lys Ser Asp Val Ile Leu His Glu Thr Ser Leu Phe Pro Gly Asp Thr
370 375 380
Phe Asn Arg Leu Lys Leu Glu Asp Thr Glu Gln Arg Leu Arg Asn Thr
385 390 395 400
Gly Tyr Phe Gln Ser Val Ser Val Tyr Thr Val Arg Ser Gln Leu Asp
405 410 415
Pro Met Gly Asn Ala Asp Gln Tyr Arg Asp Ile Phe Val Glu Val Lys
420 425 430
Glu Thr Thr Thr Gly Asn Leu Gly Leu Phe Leu Gly Phe Ser Ser Leu
435 440 445
Asp Asn Leu Phe Gly Gly Ile Glu Leu Ser Glu Ser Asn Phe Asp Leu
450 455 460
Phe Gly Ala Arg Asn Ile Phe Ser Lys Gly Phe Arg Cys Leu Arg Gly
46S 470 475 480
Gly Gly Glu His Leu Phe Leu Lys Ala Asn Phe Gly Asp Lys Val Thr
485 490 495
Asp Tyr Thr Leu Lys Trp Thr Lys Pro His Phe Leu Asn Thr Pro Trp
500 505 510
Ile Leu Gly Ile Glu Leu Asp Lys Ser Ile Asn Arg Ala Leu Ser Lys
7
CA 02341637 2001-02-19
WO 00/11183 PCT/IB99/01449
515 520 525
Asp Tyr Ala Val Gln Thr Tyr Gly Gly Asn Val Ser Thr Thr Tyr Ile
530 535 540
Leu Asn Glu His Leu Lys Tyr Gly Leu Phe Tyr Arg Gly Ser Gln Thr
545 550 555 560
Ser Leu His Glu Lys Arg Lys Phe Leu Leu Gly Pro Asn Ile Asp Ser
565 570 575
Asn Lys Gly Phe Val Ser Ala Ala Gly Val Asn Leu Asn Tyr Asp Ser
580 585 590
Val Asp Ser Pro Arg Thr Pro Thr Thr Gly Ile Arg Gly Gly Val Thr
595 600 605
Phe Glu Val Ser Gly Leu Gly Gly Thr Tyr His Phe Thr Lys Leu Ser
610 615 620
Leu Asn Ser Ser Ile Tyr Arg Lys Leu Thr Arg Lys Gly Ile Leu Lys
625 630 635 640
Ile Lys Gly Glu Ala Gln Phe Ile Lys Pro Tyr Ser Asn Thr Thr Ala
645 650 655
Glu Gly Asp Pro Val Sex Glu Arg Phe Phe Leu Gly Gly Glu Thr Thr
660 665 670
Val Arg Gly Tyr Lys Ser Phe Ile Ile Gly Pro Lys Tyr Ser Ala Thr
675 680 685
Glu Pro Gln Gly Gly Leu Ser Ser Leu Leu Ile Ser Glu Glu Phe Gln
690 695 700
Tyr Pro Leu Ile Arg Gln Pro Asn Ile Ser Ala Phe Val Phe Leu Asp
705 710 715 720
Ser Gly Phe Val Gly Leu Gln Glu Tyr Lys Ile Ser Leu Lys Asp Leu
725 730 735
Arg Ser Ser Ala Gly Phe Gly Leu Arg Phe Asp Val Met Asn Asn Val
740 745 750
Pro Val Met Leu Gly Phe Gly Trp Pro Phe Arg Pro Thr Glu Thr Leu
755 760 765
Asn Gly Glu Lys Ile Asp Val Ser Gln Arg Phe Phe Phe Ala Leu Gly
8
CA 02341637 2001-02-19
WO 00/11183 PCT/IB99/01449
770 775 7so
Gly Met Phe
785
<210> 3
<211> 3200
<212> DNA
<213> Chlamydia sp.
<220>
<221> CDS
<222> (101)..(3010)
<400> 3
gggaactttt attcgtttta ttaagtaaga tcgagtatgg tattgagaaa cgttctcgta 60
aaagatggta ttctagtaat tacagtgaat taaacgacat atg tta ggt ttt ctt 115
Met Leu Gly Phe Leu
1 5
aaa cgc ttc ttt ggt tcc tct caa gag cgt att cta aaa aaa ttt caa 163
Lys Arg Phe Phe Gly Ser Ser Gln Glu Arg Ile Leu Lys Lys Phe Gln
10 15 20
aaa ctt gta gat aaa gtg aac att tat gat gaa atg ctc acg cct tta 211
Lys Leu Val Asp Lys Val Asn Ile Tyr Asp Glu Met Leu Thr Pro Leu
25 30 35
tct gat gat gaa ttg cgg aat aag act gcg gaa tta aag cag aga tat 259
Ser Asp Asp Glu Leu Arg Asn Lys Thr Ala Glu Leu Lys Gln Arg Tyr
40 45 50
cag aac gga gag tct ctt gat agc atg ctt cct gaa get tat ggt gtc 307
Gln Asn Gly Glu Ser Leu Asp Ser Met Leu Pro Glu Ala Tyr Gly Val
55 60 65
gtg aaa aac gtt tgt cga cgt tta gca ggc acc cca gtc gaa gtc tcc 355
Val Lys Asn Val Cys Arg Arg Leu Ala Gly Thr Pro Val Glu Val Ser
70 7s eo es
gga tac cat caa aga tgg gat atg gtt cct tat gat gtg cag att cta 403
Gly Tyr His Gln Arg Trp Asp Met Val Pro Tyr Asp Val Gln Ile Leu
90 95 100
ggg gcc att get atg cac aag gga ttt att aca gag atg cag acc ggg 451
Gly Ala Ile Ala Met His Lys Gly Phe Ile Thr Glu Met Gln Thr Gly
9
CA 02341637 2001-02-19
WO 00/11183 PCT/IB99/01449
105 110 115
gag ggg aaa aca ctc act gca gtg atg cct ctg tat tta aat get tta 499
Glu Gly Lys Thr Leu Thr Ala Val Met Pro Leu Tyr Leu Asn Ala Leu
120 125 130
aca ggc aag cca gtg cat ttg gtt act gtt aac gac tat ctt gca caa 547
Thr Gly Lys Pro Val His Leu Val Thr Val Asn Asp Tyr Leu Ala Gln
135 140 145
cga gat tgt gaa tgg gta gga tcg gta ctg cgc tgg tta gga ctt aca 595
Arg Asp Cys Glu Trp Val Gly Ser-Val Leu Arg Trp Leu Gly Leu Thr
150 155 160 165
acg gga gtt ttg gtt tca gga act ctt tta gaa aag cgt aag aaa att 643
Thr Gly Val Leu Val Ser Gly Thr Leu Leu Glu Lys Arg Lys Lys Ile
170 175 180
tat caa tgt gat gtt gtc tat ggt aca gca tct gag ttt ggt ttt gat 691
Tyr Gln Cys Asp Val Val Tyr Gly Thr Ala Ser Glu Phe Gly Phe Asp
185 190 195
tat ttg aga gat aat tct ata get act cgc ctt gaa gag cag gta ggt 739
Tyr Leu Arg Asp Asn Ser Ile Ala Thr Arg Leu Glu Glu Gln Val Gly
200 205 210
aga gga tat tac ttt get atc att gat gaa gtc gac tcg atc tta ata 787
Arg Gly Tyr Tyr Phe Ala Ile Ile Asp Glu Val Asp Ser Ile Leu Ile
215 220 225
gat gaa get aga aca ccc tta att atc tca ggt cct gga gaa aaa cat 835
Asp Glu Ala Arg Thr Pro Leu Ile Ile Ser Gly Pro Gly Glu Lys His
230 235 240 245
aat cca gtc tat ttt gag ctt aaa gaa aaa gtc gca agt cta gtg tat 883
Asn Pro Val Tyr Phe Glu Leu Lys Glu Lys Val Ala Ser Leu Val Tyr
250 255 260
ttg caa aaa gag ctc tgc agc cgt atc gca tta gaa gca cgt cgc ggc 931
Leu Gln Lys Glu Leu Cys Ser Arg Ile Ala Leu Glu Ala Arg Arg Gly
265 270 275
tta gat agc ttt tta gat gtt gat att ctt cct aaa gat aaa aaa gtt 979
Leu Asp Ser Phe Leu Asp Val Asp Ile Leu Pro Lys Asp Lys Lys Val
280 285 290
ctt gaa ggc atc tct gaa ttt tgc cgc agc ctt tgg ttg gta agc aaa 1027
Leu Glu Gly Ile Ser Glu Phe Cys Arg Ser Leu Trp Leu Val Ser Lys
10
CA 02341637 2001-02-19
WO 00/11183 PCT/IB99/01449
295 300 305
gga atg cct ttg aat cgt gtg tta cgt cgt gta cgt gag cac cca gat 1075
Gly Met Pro Leu Asn Arg Val Leu Arg Arg Val Arg Glu His Pro Asp
310 315 320 325
ctt cgt get atg atc gat aaa tgg gat gtt tat tat cat get gag cag 1123
Leu Arg Ala Met Ile Asp Lys Trp Asp Val Tyr Tyr His Ala Glu Gln
330 335 340
aat aaa gaa gag agc cta gag cgt ctt tca gag ctc tac att att gtt 1171
Asn Lys Glu Glu Ser Leu Glu Arg Leu Ser Glu Leu Tyr Ile Ile Val
345 350 355
gat gag cac aat aat gat ttt gag ctt aca gat aaa gga atg cag cag 1219
Asp Glu His Asn Asn Asp Phe Glu Leu Thr Asp Lys Gly Met Gln Gln
360 365 370
tgg gtt gag tat get gga ggc tct acc gaa gag ttc gtg atg atg gat 1267
Trp Val Glu Tyr Ala Gly Gly Ser Thr Glu Glu Phe Val Met Met Asp
375 380 385
atg ggg cat gag tat get ctt ata gaa aat gat gag acc cta tca cct 1315
Met Gly His Glu Tyr Ala Leu Ile Glu Asn Asp Glu Thr Leu Ser Pro
390 395 400 405
gca gat aag atc aat aaa aaa att gca att tct gaa gaa gac acc tta 1363
Ala Asp Lys Ile Asn Lys Lys Ile.Ala Ile Ser Glu Glu Asp Thr Leu
410 415 420
aga aag get cgt get cac gga tta cga cag tta tta cga gcc caa ctt 1411
Arg Lys Ala Arg Ala His Gly Leu Arg Gln Leu Leu Arg Ala Gln Leu
425 430 435
ctc atg gag cgt gat gta gat tat att gtc cgc gac gat cag att gtg 1459
Leu Met Glu Arg Asp Val Asp Tyr Ile Val Arg Asp Asp Gln Ile Val
440 445 450
att atc gat gaa cat aca gga cgt cct caa cct gga cga cgt ttt tct 1507
Ile Ile Asp Glu His Thr Gly Arg Pro Gln Pro Gly Arg Arg Phe Ser
455 460 465
gaa ggc ctc cat caa get atc gaa get aaa gaa cac gtc act atc cgt 1555
Glu Gly Leu His Gln Ala Ile Glu Ala Lys Glu His Val Thr Ile Arg
470 475 480 485
aag gaa tct cag acg ctt get aca gtc acg ttg caa aat ttc ttc cgt 1603
Lys Glu Ser Gln Thr Leu Ala Thr Val Thr Leu Gln Asn Phe Phe Arg
11
CA 02341637 2001-02-19
WO 00/11183 PCT/IB99/01449
490 495 500
cta tat gaa aag ctt gca ggg atg acc gga aca gca att acg gag tct 1651
Leu Tyr Glu Lys Leu Ala Gly Met Thr Gly Thr Ala Ile Thr Glu Ser
505 510 515
cga gag ttt aaa gaa att tat aat ctt tat gtc ctc caa gta ccc acg 1699
Arg Glu Phe Lys Glu Ile Tyr Asn Leu Tyr Val Leu Gln Val Pro Thr
520 525 530
ttc aag ccc ttg tta cgc ata gat cat aat gat gaa ttt tat atg aca 1747
Phe Lys Pro Leu Leu Arg Ile Asp His Asn Asp Glu Phe Tyr Met Thr
535 540 545
gag cgt gag aag tac cac get att gtt aat gag att gcg act att cat 1795
Glu Arg Glu Lys Tyr His Ala Ile Val Asn Glu Ile Ala Thr Ile His
550 555 560 565
ggc aag ggg aac cct att ctt gtt ggt aca gaa tct gta gag gtc tct 1843
Gly Lys Gly Asn Pro Ile Leu Val Gly Thr Glu Ser Val Glu Val Ser
570 575 580
gag aag ctg tct cgg att ttg aga cag aat cgg ata gag cat act gta 1891
Glu Lys Leu Ser Arg Ile Leu Arg Gln Asn Arg Ile Glu His Thr Val
585 590 595
ttg aat get aag aat cat get caa gaa gca gaa att ata gca gga gca 1939
Leu Asn Ala Lys Asn His Ala Gln Glu Ala Glu Ile Ile Ala Gly Ala
600 605 610
ggg aaa tta ggt get gtg act gta get aca aat atg get ggt cga ggc 1987
Gly Lys Leu Gly Ala Val Thr Val Ala Thr Asn Met Ala Gly Arg Gly
615 620 625
aca gac atc aaa cta gat aat gaa get gtg atc gta ggc ggt ctc cat 2035
Thr Asp Ile Lys Leu Asp Asn Glu Ala Val Ile Val Gly Gly Leu His
630 635 640 645
gtg atc ggt act aca cgg cat caa tcc cgt cga att gat aga cag ttg 2083
Val Ile Gly Thr Thr Arg His Gln Ser Arg Arg Ile Asp Arg Gln Leu
650 655 660
cgt ggg cgt tgt get cgt tta gga gac cct ggc get gcg aaa ttc ttt 2131
Arg Gly Arg Cys Ala Arg Leu Gly Asp Pro Gly Ala Ala Lys Phe Phe
665 670 675
tta tct ttt gaa gat cgg ctg atg cga cta ttc gcc tcc cct aaa ttg 2179
Leu Ser Phe Glu Asp Arg Leu Met Arg Leu Phe Ala Ser Pro Lys Leu
12
CA 02341637 2001-02-19
WO 00/11183 PCT/IB99/01449
6gp 685 690
aat acc ctt atc cgt cat ttt cgt cct cca gaa gga gag gcg atg tcg 2227
Asn Thr Leu Ile Arg His Phe Arg Pro Pro Glu Gly Glu Ala Met Ser
695 700 705
gac cct atg ttt aat aga ctc ata gaa aca.gca cag aaa cgt gtc gaa 2275
Asp Pro Met Phe Asn Arg Leu Ile Glu Thr Ala Gln Lys Arg Val Glu
710 715 720 725
ggg aga aac tat act atc cgt aag cat acc tta gag tat gat gat gtc 2323
Gly Arg Asn Tyr Thr Ile Arg Lys His Thr Leu Glu Tyr Asp Asp Val
730 735 740
atg aat aag cag aga caa geg ata tac get tte cgc cat gat gtc tta 2371
Met Asn Lys Gln Arg Gln Ala Ile Tyr Ala Phe Arg His Asp Val Leu
745 750 755
cat gcg gaa tct gtt ttc gat ctt gca aaa gaa att cta tgc cat gtg 2419
His Ala Glu Ser Val Phe Asp Leu Ala Lys Glu Ile Leu Cys His Val
760 765 770
tct ctg atg gta gca tcc tta gtg atg agt gat cgt cag ttc aaa ggg 2467
Ser Leu Met Val Ala Ser Leu Val Met Ser Asp Arg Gln Phe Lys Gly
775 780 785
tgg aca ttg cca aat ctt gaa gaa tgg ata acc tca tct ttc cca ata 2515
Trp Thr Leu Pro Asn Leu Glu Glu Trp Ile Thr Ser Ser Phe Pro Ile
790 795 800 805
gcc tta aat ata gaa gaa ctc aga cag ctt aaa gat aca gat tct att 2563
Ala Leu Asn Ile Glu Glu Leu Arg Gln Leu Lys Asp Thr Asp Ser Ile
810 815 820
get gaa aag atc get get gaa ttg att caa gag ttt caa gta cgc ttt 2611
Ala Glu Lys Ile Ala Ala Glu Leu Ile Gln Glu Phe Gln Val Arg Phe
825 830 835
gat cat atg gta gaa ggg ctc tcc aaa get gga ggg gaa gaa ttg gat 2659
Asp His Met Val Glu Gly Leu Ser Lys Ala Gly Gly Glu Glu Leu Asp
840 845 850
gca tct get att tgt aga gat gtc gtt cgg tct gtc atg gtc atg cat 2707
Ala Ser Ala Ile Cys Arg Asp Val Val Arg Ser Val Met Val Met His
855 860 865
att gat gag cag tgg cgg att cat ctt gta gat atg gac tta cta cgg 2755
Ile Asp Glu Gln Trp Arg Ile His Leu Val Asp Met Asp Leu Leu Arg
13
CA 02341637 2001-02-19
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g7p 875 880 885
agt gaa gtt ggc cta cgt act gta ggg caa aaa gat cct ttg tta gaa 2803
Ser Glu Val Gly Leu Arg Thr Val Gly Gln Lys Asp Pro Leu Leu Glu
890 895 900
ttt aaa cac gag tct ttc tta ctg ttt gag agc ttg att cgt gat att 2851
Phe Lys His Glu Ser Phe Leu Leu Phe Glu Ser Leu Ile Arg Asp Ile
905 910 915
cgt att acg att gcg cgg cat ctt ttc cgt ctt gaa ctg acc gta gag 2899
Arg Ile Thr Ile Ala Arg His Leu Phe Arg Leu Glu Leu Thr Val Glu
920 925 930
cct aat cct cgt gtc aac aac gtg att cct act gta get acg tct ttt 2947
Pro Asn Pro Arg Val Asn Asn Val Ile Pro Thr Val Ala Thr Ser Phe
935 940 945
cat aat aat gtg aat tac ggt ccg tta gag ttg act gta gtt aca gat 2995
His Asn Asn Val Asn Tyr Gly Pro Leu Glu Leu Thr Val Vai Thr Asp
950 955 960 965
tct gaa gat caa gat taagaaataa cctagaagga ggagtccttc taggtatttg 3050
Ser Glu Asp Gln Asp
970
tttcacaggc tacatagttt ttatttttaa ataaatgggg tccattctga ccaaacttgg 3110
tcattatagt agcgaatcca gaattttctg cgattgccgg ctatgaggaa ctgggcatag 3170
tggctcagtg tattgtttgc tatacagaaa 3200
<210> 4
<211> 970
<212> PRT
<213> Chlamydia sp.
<400> 4
Met Leu Gly Phe Leu Lys Arg Phe Phe Gly Ser Ser Gln Glu Arg Ile
1 5 10 15
Leu Lys Lys Phe Gln Lys Leu Val Asp Lys Val Asn Ile Tyr Asp Glu
20 25 30
Met Leu Thr Pro Leu Ser Asp Asp Glu Leu Arg Asn Lys Thr Ala Glu
35 40 45
14
CA 02341637 2001-02-19
WO 00/11183 PCT/IB99/01449
Leu Lys Gln Arg Tyr Gln Asn Gly Glu Ser Leu Asp Ser Met Leu Pro
50 55 60
Glu Ala Tyr Gly Val Val Lys Asn Val Cys Arg Arg Leu Ala Gly Thr
65 70 75 80
Pro Val Glu Val Ser Gly Tyr His Gln Arg Trp Asp Met Val Pro Tyr
85 90 95
Asp Val Gln Ile Leu Gly Ala Ile Ala Met His Lys Gly Phe Ile Thr
100 105 110
Glu Met Gln Thr Gly Glu Gly Lys Thr Leu Thr Ala Val Met Pro Leu
115 120 125
Tyr Leu Asn Ala Leu Thr Gly Lys Pro Val His Leu Val Thr Val Asn
130 135 140
Asp Tyr Leu Ala Gln Arg Asp Cys Glu Trp Val Gly Ser Val Leu Arg
145 150 155 160
Trp Leu Gly Leu Thr Thr Gly Val Leu Val Ser Gly Thr Leu Leu Glu
165 170 175
Lys Arg Lys Lys Ile Tyr Gln Cys Asp Val Val Tyr Gly Thr Ala Ser
180 185 190
Glu Phe Gly Phe Asp Tyr Leu Arg Asp Asn Ser Ile Ala Thr Arg Leu
195 200 205
Glu Glu Gln Val Gly Arg Gly Tyr Tyr Phe Ala Ile Ile Asp Glu Val
210 215 220
Asp Ser Ile Leu Ile Asp Glu Ala Arg Thr Pro Leu Ile Ile Ser Gly
225 230 235 240
Pro Gly Glu Lys His Asn Pro Val Tyr Phe Glu Leu Lys Glu Lys Val
245 250 255
Ala Ser Leu Val Tyr Leu Gln Lys Glu Leu Cys Ser Arg Ile Ala Leu
260 265 270
Glu Ala Arg Arg Gly Leu Asp Ser Phe Leu Asp Val Asp Ile Leu Pro
275 280 285
Lys Asp Lys Lys Val Leu Glu Gly Ile Ser Glu Phe Cys Arg Ser Leu
290 295 300
15
CA 02341637 2001-02-19
WO 00!11183 PCT/IB99/01449
Trp Leu Val Ser Lys Gly Met Pro Leu Asn Arg Val Leu Arg Arg Val
305 310 315 320
Arg Glu His Pro Asp Leu Arg Ala Met Ile Asp Lys Trp Asp Val Tyr
325 330 335
Tyr His Ala Glu Gln Asn Lys Glu Glu Ser Leu Glu Arg Leu Ser Glu
340 345 350
Leu Tyr Ile Ile Val Asp Glu His Asn~Asn Asp Phe Glu Leu Thr Asp
355 360 365
Lys Gly Met Gln Gln Trp Val Glu Tyr Ala Gly Gly Ser Thr Glu Glu
370 375 380
Phe Val Met Met Asp Met Gly His Glu Tyr Ala Leu Ile Glu Asn Asp
385 390 395 400
Glu Thr Leu Ser Pro Ala Asp Lys Ile Asn Lys Lys Ile Ala Ile Ser
405 410 415
Glu Glu Asp Thr Leu Arg Lys Ala Arg Ala His Gly Leu Arg Gln Leu
420 425 430
Leu Arg Ala Gln Leu Leu Met Glu Arg Asp Val Asp Tyr Ile Val Arg
435 440 445
Asp Asp Gln Ile Val Ile Ile Asp Glu His Thr Gly Arg Pro Gln Pro
450 455 460
Gly Arg Arg Phe Ser Glu Gly Leu His Gln Ala Ile Glu Ala Lys Glu
465 470 475 480
His Val Thr Ile Arg Lys Glu Ser Gln Thr Leu Ala Thr Val Thr Leu
485 490 495
Gln Asn Phe Phe Arg Leu Tyr Glu Lys Leu Ala Gly Met Thr Gly Thr
500 505 510
Ala Ile Thr Glu Ser Arg Glu Phe Lys Glu Ile Tyr Asn Leu Tyr Val
515 520 525
Leu Gln Val Pro Thr Phe Lys Pro Leu Leu Arg Ile Asp His Asn Asp
530 535 540
Glu Phe Tyr Met Thr Glu Arg Glu Lys Tyr His Ala Ile Val Asn Glu
545 550 555 560
16
CA 02341637 2001-02-19
WO 00/11183 PCT/IB99/01449
Ile Ala Thr Ile His Gly Lys Gly Asn Pro Ile Leu Val Gly Thr Glu
565 570 575
Ser Val Glu Val Ser Glu Lys Leu Ser Arg Ile Leu Arg Gln Asn Arg
580 585 590
Ile Glu His Thr Val Leu Asn Ala Lys Asn His Ala Gln Glu Ala Glu
595 600 605
Ile Ile Ala Gly Ala Gly Lys Leu Gly Ala Val Thr Val Ala Thr Asn
610 615 620
Met Ala Gly Arg Gly Thr Asp Ile Lys Leu Asp Asn Glu Ala Val Ile
625 630 635 640
Val Gly Gly Leu His Val Ile Gly Thr Thr Arg His Gln Ser Arg Arg
645 650 655
Ile Asp Arg Gln Leu Arg Gly Arg Cys Ala Arg Leu Gly Asp Pro Gly
660 665 670
Ala Ala Lys Phe Phe Leu Ser Phe Glu Asp Arg Leu Met Arg Leu Phe
675 680 685
Ala Ser Pro Lys Leu Asn Thr Leu Ile Arg His Phe Arg Pro Pro Glu
690 695 700
Gly Glu Ala Met Ser Asp Pro Met Phe Asn Arg Leu Ile Glu Thr Ala
705 710 715 720
Gln Lys Arg Val Glu Gly Arg Asn Tyr Thr Ile Arg Lys His Thr Leu
725 730 735
Glu Tyr Asp Asp Val Met Asn Lys Gln Arg Gln Ala Ile Tyr Ala Phe
740 745 750
Arg His Asp Val Leu His Ala Glu Ser Val Phe Asp Leu Ala Lys Glu
755 760 765
Ile Leu Cys His Val Ser Leu Met Val~Ala Ser Leu Val Met Ser Asp
770 775 780
Arg Gln Phe Lys Gly Trp Thr Leu Pro Asn Leu Glu Glu Trp Ile Thr
785 790 795 B00
Ser Ser Phe Pro Ile Ala Leu Asn Ile Glu Glu Leu Arg Gln Leu Lys
805 810 815
17
CA 02341637 2001-02-19
WO 00/11183 PCT/IB99/01449
Asp Thr Asp Sex Ile Ala Glu Lys Ile Ala Ala Glu Leu Ile Gln Glu
820 825 830
Phe Gln Val Arg Phe Asp His Met Val Glu Gly Leu Ser Lys Ala Gly
835 840 845
Gly Glu Glu Leu Asp Ala Ser Ala Ile Cys Arg Asp Val Val Arg Ser
850 855 860
Val Met Val Met His Ile Asp Glu Gln Trp Arg Ile His Leu Val Asp
865 870 875 880
Met Asp Leu Leu Arg Ser'Glu Val Gly Leu Arg Thr Val Gly Gln Lys
885 890 895
Asp Pro Leu Leu Glu Phe Lys His Glu Ser Phe Leu Leu Phe Glu Ser
900 905 910
Leu Ile Arg Asp Ile Arg Ile Thr Ile Ala Arg His Leu Phe Arg Leu
915 920 925
Glu Leu Thr Val Glu Pro Asn Pro Arg Val Asn Asn Val Zle Pro Thr
930 935 940
Val Ala Thr Ser Phe His Asn Asn Val Asn Tyr Gly Pro Leu Glu Leu
945 950 955 960
Thr Val Val Thr Asp Ser Glu Asp Gln Asp
965 970
<210> 5
<211> 1300
<212> DNA
<213> Chlamydia sp.
<220>
<221> CDS
<222> (101)..(1180)
<400> 5
gaacaaaacg tttgggcttc actcttgaga aaaacaagag cataaggtaa tttgcagata 60
attccgaata tagtcctctt aatcgaacct tggcaacagc atg ggt gaa aaa aca 115
Met Gly Glu Lys Thr
1 5
gaa aag gcc acg ccg aag cga ctt aga gat get cgg aaa aaa ggt caa 163
18
CA 02341637 2001-02-19
WO 00/11183 PCT/IB99/01449
Glu Lys Ala Thr Pro Lys Arg Leu Arg Asp Ala Arg Lys Lys Gly Gln
10 15 20
gta gca aaa tct cag gat ttt cct tct gcg gtt acc ttt atc gtc tct 211
Val Ala Lys Ser Gln Asp Phe Pro Ser Ala Val Thr Phe Ile Val Ser
25 30 35
atg ttt acg get tte tec cta tcg acc ttt ttt tte aag cat tta ggt 259
Met Phe Thr Ala Phe Ser Leu Ser Thr Phe Phe Phe Lys His Leu Gly
40 45 50
ggc ttt ctg gtt tcc atg ctc tca caa get ccc act cgc cat gat cct 30?
Gly Phe Leu Val Ser Met Leu Ser Gln Ala Pro Thr Arg His Asp Pro
S5 60 65
gta att acc tta ttt tat ttg aag aac tgt ctt atg ctt att tta aca 355
Val. Ile Thr Leu Phe Tyr Leu Lys Asn Cys Leu Met Leu Ile Leu Thr
70 75 80 85
gca tca ctt cec tta ctg gga get gtt get gtt gtt ggc gtc att gta 403
Ala Ser Leu Pro Leu Leu Gly Ala Val Ala Val Val Gly Val Ile Val
90 95 100
ggt ttt ctt atc gtt ggt cct aca ttt tct acc gaa gtt ttt aaa cca 451
Gly Phe Leu Ile Val Gly Pro Thr Phe Ser Thr Glu Val Phe Lys Pro
I05 110 115
gat atc aag aag ttc aac cct att gag aac atc aaa caa aag ttt aaa 499
Asp Ile Lys Lys Phe Asn Pro Ile Glu Asn Ile Lys Gln Lys Phe Lys
120 125 130
ata aag act ctc ata gag ctg atc aaa tcg att tta aaa att ttt gga 547
Ile Lys Thr Leu Ile Glu Leu Ile Lys Ser Ile Leu Lys Ile Phe Gly
135 140 145
gca gcc tta att tta tac ata acg tta aaa agc aaa gtc tct tta att 595
Ala Ala Leu Ile Leu Tyr Ile Thr Leu Lys Ser Lys Val Ser Leu Ile
150 155 160 165
ata gaa act gca gga gtc tct cct ata att act get caa atc ttc aaa 643
Ile Glu Thr Ala Gly Val Ser Pro Ile Ile Thr Ala Gln Ile Phe Lys
170 175 180
gaa att ttt tat aaa gca gta acc tcg ata gga att ttc ttt ttg att 691
Glu Ile Phe Tyr Lys Ala Val Thr Ser Ile Gly Ile Phe Phe Leu Ile
185 190 195
gtt gcg att ett gac ctt gtc tat cag cge cac aat tte get aaa gaa 739
19
CA 02341637 2001-02-19
WO 00/11183 PCT/IB99/01449
Val Ala Ile Leu Asp Leu Val Tyr Gln Arg His Asn Phe Ala Lys Glu
200 205 210
tta aag atg gag aag ttt gag gtt aag cag gag ttt aaa gac acg gaa 787
Leu Lys Met Glu Lys Phe Glu Val Lys Gln Glu Phe Lys Asp Thr Glu
215 220 225
gga aat cct gag att aaa ggc cgt cgt cga caa att get caa gaa att 835
Gly Asn Pro Glu Ile Lys Gly Arg Arg Arg Gln I1e Ala Gln Glu Ile
230 235 240 245
gcc tat gaa gac tcg tca tca cag gtg aaa cat gca agc acc gta gtc 883
Ala Tyr Glu Asp Ser Ser Ser Gln Val Lys His Ala Ser Thr Val Val
250 255 260
tct aat ccc aaa gat att get gtt get att ggc tac atg cct gaa aaa 931
Ser Asn Pro Lys Asp Ile Ala Val Ala Ile Gly Tyr Met Pro Glu Lys
265 270 275
tat aaa gca cct tgg atc att gcc atg ggc atc aac tta cga get aaa 979
Tyr Lys Ala Pro Trp Ile Ile Ala Met Gly Ile Asn Leu Arg Ala Lys
280 285 290
agg ata ctt gat gaa get gaa aag tac gga att ccc att atg cga aac 1027
Arg Ile Leu Asp Glu Ala Glu Lys Tyr Gly Ile Pro Ile Met Arg Asn
295 300 305
gta cct tta gca cat cag ctt ttg gat gaa ggg aag gaa tta aaa ttt 1075
Val Pro Leu Ala His Gln Leu Leu Asp Glu Gly Lys Glu Leu Lys Phe
310 315 320 325
att cca gaa tct act tac gaa get att gga gaa att cta ctc tat atc 1123
Ile Pro Glu Ser Thr Tyr Glu Ala Ile Gly Glu Ile Leu Leu Tyr Ile
330 335 340
act tca ctg aat gcg caa aat cct aat aat aaa aat act aac caa cct 1171
Thr Ser Leu Asn Ala Gln Asn Pro Asn Asn Lys Asn Thr Asn Gln Pro
345 350 355
gat cat tta taatgaataa gctactcaat ttcgtcagca gaacacttgg 1220
Asp His Leu
360
tggcgatacc gccttaaaca tgatcaataa gtccagcgac ttaatccttg ctctttggat 1280
gatgggcgtt gtcttaatga 1300
20
CA 02341637 2001-02-19
WO 00/111$3 PCT/IB99/01449
<210> 6
<211> 360
<212> PRT
<213> Chlamydia sp.
<400> 6
Met Gly Glu Lys Thr Glu Lys Ala Thr Pro Lys Arg Leu Arg Asp Ala
1 5 10 15
Arg Lys Lys Gly Gln Val Ala Lys Ser Gln Asp Phe Pro Ser Ala Val
20 25 30
Thr Phe Ile Val Ser Met Phe Thr Ala Phe Ser Leu Ser Thr Phe Phe
35 40 45
Phe Lys His Leu Gly Gly Phe Leu Val Ser Met Leu Ser Gln Ala Pro
50 55 60
Thr Arg His Asp Pro Val Ile Thr Leu Phe Tyr Leu Lys Asn Cys Leu
6S 70 75 80
Met Leu Ile Leu Thr Ala Ser Leu Pro Leu Leu Gly Ala Val Ala Val
85 90 95
Val Gly Val Ile Val Gly Phe Leu Ile Val Gly Pro Thr Phe Ser Thr
100 105 110
Glu Val Phe Lys Pro Asp Ile Lys Lys Phe Asn Pro Ile Glu Asn Ile
115 120 125
Lys Gln Lys Phe Lys Ile Lys Thr Leu Ile Glu Leu Ile Lys Ser Ile
130 135 140
Leu Lys Ile Phe Gly Ala Ala Leu Ile Leu Tyr Ile Thr Leu Lys Ser
145 150 155 160
Lys Val Ser Leu Ile Ile Glu Thr Ala Gly Val Ser Pro Ile Ile Thr
165 170 175
Ala Gln Ile Phe Lys Glu Ile Phe Tyr Lys Ala Val Thr Ser Ile Gly
180 185 190
Ile Phe Phe Leu Ile Val Ala Ile Leu Asp Leu Val Tyr Gln Arg His
195 200 205
Asn Phe Ala Lys Glu Leu Lys Met Glu Lys Phe Glu Val Lys Gln Glu
210 215 220
21
CA 02341637 2001-02-19
WO 00/11183 PCT/IB99/01449
Phe Lys Asp Thr Glu Gly Asn Pro Glu Ile Lys Gly Arg Arg Arg Gln
225 230 235 240
Ile Ala Gln Glu Ile Ala Tyr Glu Asp Ser Ser Ser Gln Val Lys His
245 250 255
Ala Ser Thr Val Val Ser Asn Pro Lys Asp Ile Ala Val Ala Ile Gly
260 265 270
Tyr Met Pro Glu Lys Tyr Lys Ala Pro Trp Ile Ile Ala Met Gly Ile
275 280 285
Asn Leu Arg Ala Lys Arg Ile Leu Asp Glu Ala Glu Lys Tyr Gly Ile
290 295 300
Pro Ile Met Arg Asn Val Pro Leu Ala His Gln Leu Leu Asp Glu Gly
305 310 315 320
Lys Glu Leu Lys Phe Ile Pro Glu Ser Thr Tyr Glu Ala Ile Gly Glu
325 330 335
Ile Leu Leu Tyr Ile Thr Ser Leu Asn Ala Gln Asn Pro Asn Asn Lys
340 395 350
Asn Thr Asn Gln Pro Asp His Leu
355 360
<210> 7
<211> 2350
<212> DNA
<213> Chlamydia sp.
<220>
<221> CDS
<222> (101)..(2230)
<400> 7
aatctactta cgaagctatt ggagaaattc tactctatat cacttcactg aatgcgcaaa 60
atcctaataa taaaaatact aaccaacctg atcatttata atg aat aag cta ctc 115
Met Asn Lys Leu Leu
1 S
aat ttc gtc agc aga aca ctt ggt ggc gat acc gcc tta aac atg atc 163
Asn Phe Val Ser Arg Thr Leu Gly Gly Asp Thr Ala Leu Asn Met Ile
10 15 20
22
CA 02341637 2001-02-19
WO 00/11183 PCT/IB99/01449
aat aag tcc agc gac tta atc ctt get ctt tgg atg atg ggc gtt gtc 211
Asn Lys Ser Ser Asp Leu Ile Leu Ala Leu Trp Met Met Gly Val Val
25 30 35
tta atg atc att att cct ttg cct ccg cct atc gtt gac ttg atg atc 259
Leu Met Ile Ile Ile Pro Leu Pro Pro Pro Ile Val Asp Leu Met Ile
40 45 50
acc atc aac tta tcg atc tct gta ttc tta ttg atg gtg get ctt tat 307
Thr Ile Asn Leu Ser Ile Ser Val Phe Leu Leu Met Val Ala Leu Tyr
55 60 65
att cca agt get ttg cag ctg tct gtt ttt ccc tcg ttg ctc ctc atc 355
Ile Pro Ser Ala Leu Gln Leu Ser Val Phe Pro Ser Leu Leu Leu Ile
70 75 80 85
act acg atg ttc cgc ttg ggg att aat att tcc tct tct cga cag att 403
Thr Thr Met Phe Arg Leu Gly Ile Asn Ile Ser Ser Ser Arg Gln Ile
90 95 100
ctc ctt aaa gcg tat gcg agt cat gtc att cag gcc ttc gga gac ttc 451
Leu Leu Lys Ala Tyr Ala Ser His Val Ile Gln Ala Phe Gly~Asp Phe
105 110 115
gtg gtt gga ggg aac tat gtg gtc ggg ttc att atc ttc ctc att att 499
Val Val Gly Gly Asn Tyr Val Val Gly Phe Ile Ile Phe Leu Ile Ile
120 125 130
aca atc att cag ttt atc gta gta act aag ggt gcc gag cgt gtt gcc 547
Thr Ile Ile Gln Phe Ile Val Val Thr Lys Gly Ala Glu Arg Val Ala
135 140 145
gaa gtt get gcc cga ttc cga ttg gat gcg atg cca ggt aaa cag atg 595
Glu Val Ala Ala Arg Phe Arg Leu Asp Ala Met Pro Gly Lys Gln Met
150 155 160 165
gcg att gat gcg gac tta cga get ggt atg att gat gcc aca caa get 643
Ala Ile Asp Ala Asp Leu Arg Ala Gly Met Ile Asp Ala Thr Gln Ala
170 175 180
cgt gat aaa agg get caa atc caa aag gaa agt gaa ctc tac gga gcc 691
Arg Asp Lys Arg Ala Gln Ile Gln Lys Glu Ser Glu Leu Tyr Gly Ala
185 190 195
atg gac ggt gcc atg aag ttc atc aaa gga gac gtt atc get ggt atc 739
Met Asp Gly Ala Met Lys Phe Ile Lys Gly Asp Val Ile Ala Gly Ile
200 205 210
23
CA 02341637 2001-02-19
WO 00/11183 PCT/IB99/01449
gtt atc tct ttg att aac att gtt ggc ggt ttg acg att ggg gtg get 787
Val Ile Ser Leu Ile Asn Ile Val Gly Gly Leu Thr Ile Gly Val Ala
215 220 225
atg cac ggc atg gac ctc get caa gca get cac gtc tac act ctt ctc 835
Met His Gly Met Asp Leu Ala Gln Ala Ala His Val Tyr Thr Leu Leu
230 235 240 245
tcc att gga gat ggt tta gtc tct caa att cct tct ctt ttg att gcg 883
Ser Ile Gly Asp Gly Leu Val Ser Gln Ile Pro Ser Leu Leu Ile Ala
250 255 260
ttg aca gcg ggt att gtc acg act cgt gta tcg agt gac aaa aat acg 931
Leu Thr Ala Gly Ile Val Thr Thr Arg Val Ser Ser Asp Lys Asn Thr
265 270 275
aac ttg ggt aaa gag att tct act cag ctc gtt aaa gaa cca cga gca 979
Asn Leu Gly Lys Glu Ile Ser Thr Gln Leu Vai Lys Glu Pro Arg Ala
280 285 290
cta ctc ctt gca ggt get gca act tta ggg gtt ggt ttc ttc aag ggc 1027
Leu Leu Leu Ala Gly Ala Ala Thr Leu Gly Val Gly Phe Phe Lys Gly
295 300 305
ttc cct cta tgg tcc ttc tcc att tta gca tta att ttc gtt gcc tta 1075
Phe Pro Leu Trp Ser Phe Ser Ile Leu Ala Leu Ile Phe Val Ala Leu
310 315 320 325
ggg att ctc cta ctg act aag aaa tca gcg gca gga aaa aaa ggt ggt 1123
Gly Ile Leu Leu Leu Thr Lys Lys Ser Ala Ala Gly Lys Lys Gly Gly
330 335 340
ggc tca gga get tca aca acc gta ggg get get ggt gat ggc get get 1171
Gly Ser Gly Ala Ser Thr Thr Val Gly Ala Ala Gly Asp Gly Ala Ala
345 350 355
act gtt ggg gat aat ccc gat gac tat tct cta act ctt ccc gta att 1219
Thr Val Gly Asp Asn Pro Asp Asp Tyr Ser Leu Thr Leu Pro Val Ile
360 365 370
cta gaa ctt gga aaa gat ctc tct aag ctt atc caa cac aag aca aaa 1267
Leu Glu Leu Gly Lys Asp Leu Ser Lys Leu Ile Gln His Lys Thr Lys
375 380 385
tca gga caa agc ttt gtt gat gat atg att cct aaa atg cgg caa get 1315
Ser Gly Gln Ser Phe Val Asp Asp Met Ile Pro Lys Met Arg Gln Ala
390 395 400 405
24
CA 02341637 2001-02-19
WO 00/11183 PCT/IB99/01449
ctc tat cag gat atc gga atc cga tac cct ggc att cat gtt cgc aca 1363
Leu Tyr Gln Asp Ile Gly Ile Arg Tyr Pro Gly Ile His Val Arg Thr
410 415 420
gat tcc cct tct tta gaa gga tac gat tat atg att ctg ctt aat gaa 1411
Asp Ser Pro Ser Leu Glu Gly Tyr Asp Tyr Met Ile Leu Leu Asn Glu
425 430 435
gtc cct tat gtg cga gga aaa att cct ccg cac cat gtg tta acc aat 1459
Val Pro Tyr Val Arg Gly Lys Ile Pro Pro His His Val Leu Thr Asn
440 445 450
gag gtg gag gac aat ctc agc cgt tat aat cta cct ttc att acc tat 1507
Glu Val Glu Asp Asn Leu Ser Arg Tyr Asn Leu Pro Phe Ile Thr Tyr
455 460 465
aag aat get gcg ggt ctt cct tct get tgg gtt agt gaa gat gca aaa 1555
Lys Asn Ala Ala Gly Leu Pro Ser Ala Trp Val Ser Glu Asp Ala Lys
470 475 480 4$5
get att cta gag aag gca gca att aaa tat tgg acg ccg ctc gaa gtg 1603
Ala Ile Leu Glu Lys Ala Ala Ile Lys Tyr Trp Thr Pro Leu Glu Val
490 495 500
atc att ctc cat ctt tcg tac ttt ttc cat aaa agc tct caa gag ttt 1651
Ile Ile Leu His Leu Ser Tyr Phe Phe His Lys Ser Ser Gln Glu Phe
505 510 515
ttg gga att caa gag gta cgt tct atg atc gaa ttt atg gaa cgt tca 1699
Leu Gly Ile Gln Glu Val Arg Ser Met Ile Glu Phe Met Glu Arg Ser
520 525 530
ttc ccg gac tta gtg aag gaa gtc aca agg ctt att cca ttg caa aag 1747
Phe Pro Asp Leu Val Lys Glu Val Thr Arg Leu Ile Pro Leu Gln Lys
535 540 545
ctt acg gaa atc ttt aag aga ttg gtt caa gag caa atc tca att aaa 1795
Leu Thr Glu Ile Phe Lys Arg Leu Val Gln Glu Gln Ile Ser Ile Lys
550 555 560 565
gac cta cgt aca atc tta gaa tct ctg agc gag tgg gcg caa act gag 1843
Asp Leu Arg Thr Ile Leu Glu Ser Leu Ser Glu Trp Ala Gln Thr Glu
570 575 580
aaa gat aca gtt ttg ctt aca gaa tat gta cgg tct tct tta aag ctt 1891
Lys Asp Thr Val Leu Leu Thr Glu Tyr Val Arg Ser Ser Leu Lys Leu
585 590 595
25
CA 02341637 2001-02-19
WO 00/11183 PCT/IB99/01449
tat atc agc ttc aag ttc tct caa gga caa tca gca att tct gtt tat 1939
Tyr Ile Ser Phe Lys Phe Ser Gln Gly Gln Ser Ala Ile Ser Val Tyr
600 605 610
ctc tta gat cca gaa att gaa gag atg att cgt gga gca att aaa cag 1987
Leu Leu Asp Pro Glu Ile Glu Glu Met Ile Arg Gly Ala Ile Lys Gln
615 620 625
aca tcg gca ggt tct tac ctt get cta gat cct gat tct gtg aac cta 2035
Thr Ser Ala Gly Ser Tyr Leu Ala Leu Asp Pro Asp Ser Val Asn Leu
630 635 640 645
att tta aaa tct atg agg aat acg atc acg cca aca cct gca gga ggc 2083
Ile Leu Lys Ser Met Arg Asn Thr Ile Thr Pro Thr Pro Ala Gly Gly
650 655 660
caa cca cca gta tta ttg aca gca att gat gta aga aga tat gta cga 2131
Gln Pro Pro Val Leu Leu Thr Ala Ile Asp Val Arg Arg Tyr Val Arg
665 670 675
aaa tta ata gaa aca gaa ttc cet gac att get gtg att tct tat caa 2179
Lys Leu Ile Glu Thr Glu Phe Pro Asp Ile Ala Val Ile Ser Tyr Gln
680 685 690
gaa atc cta cca gaa atc cgc atc cag cct tta gga aga att cag att 2227
Glu Ile Leu Pro Glu Ile Arg Ile Gln Pro Leu Gly Arg Ile Gln Ile
695 700 705
ttc taattgatac gttgtcgctc ataggaggca tatggcagca tcaggaggca 2280
Phe
710
caggtggttt aggaggcact cagggtgtca accttgcagc tgtagaagct gcagctgcaa 2340
aagcagatgc 2350
<210> 8
<211> 710
<212> PRT
<213> Chlamydia sp.
<400> 8
Met Asn Lys Leu Leu Asn Phe Val Ser Arg Thr Leu Gly Gly Asp Thr
1 5 10 15
Ala Leu Asn Met Ile Asn Lys Ser Ser Asp Leu Ile Leu Ala Leu Trp
20 25 30
26
CA 02341637 2001-02-19
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Met Met Gly Val Val Leu Met Ile Ile Ile Pro Leu Pro Pro Pro Ile
35 40 45
Val Asp Leu Met Ile Thr Ile Asn Leu Ser Ile Ser Val Phe Leu Leu
50 55 60
Met Val Ala-Leu Tyr Ile Pro Ser Ala Leu Gln Leu Ser Val Phe Pro
65 70 75 80
Ser Leu Leu Leu Ile Thr Thr Met Phe Arg Leu Gly Ile Asn Ile Ser
85 90 95
Ser Ser Arg Gln Ile Leu Leu Lys Ala Tyr Ala Ser His Val Ile Gln
100 I05 110
Ala Phe Gly Asp Phe Val Val Gly Gly Asn Tyr Val Val Gly Phe Ile
115 120 125
Ile Phe Leu Ile Ile Thr Ile Ile Gln Phe Ile Val Val Thr Lys Gly
130 135 140
Ala Glu Arg Val Ala Glu Val Ala Ala Arg Phe Arg Leu Asp Ala Met
145 150 155 160
Pro Gly Lys Gln Met Ala Ile Asp Ala Asp Leu Arg Ala Gly Met Ile
165 170 175
Asp Ala Thr Gln Ala Arg Asp Lys Arg Ala Gln Ile Gln Lys Glu Ser
180 185 190
Glu Leu Tyr Gly Ala Met Asp Gly Ala Met Lys Phe Ile Lys Gly Asp
195 200 205
Val Ile Ala Gly Ile Val Ile Ser Leu Ile Asn Ile Val Gly Gly Leu
210 215 220
Thr Ile Gly Val Ala Met His Gly Met Asp Leu Ala Gln Ala Ala His
225 230 235 240
Val Tyr Thr Leu Leu Ser Ile Gly Asp Gly Leu Val Ser Gln Ile Pro
245 250 255
Ser Leu Leu Ile Ala Leu Thr Ala Gly Ile Val Thr Thr Arg Val Ser
260 265 270
Ser Asp Lys Asn Thr Asn Leu Gly Lys Glu Ile Ser Thr Gln Leu Val
275 280 285
27
CA 02341637 2001-02-19
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Lys Glu Pro Arg Ala Leu Leu Leu Ala Gly Ala Ala Thr Leu Gly Val
290 295 300
Gly Phe Phe Lys Gly Phe Pro Leu Trp Ser Phe Ser Ile Leu Ala Leu
305 310 315 320
Ile Phe Val Ala Leu Gly Ile Len'Leu~Leu~Thr Lys Lps Ser Ala Ala
325 330 335
Gly Lys Lys Gly Gly Gly Ser Gly Ala Ser Thr Thr Val Gly Ala Ala
340 345 350
Gly Asp Gly Ala Ala Thr Val Gly Asp Asn Pro Asp Asp Tyr Ser Leu
35S 360 365
Thr Leu Pro Val Ile Leu Glu Leu Gly Lys Asp Leu Ser Lys Leu Ile
370 375 380
Gln His Lys Thr Lys Ser Gly Gln Ser Phe Val Asp Asp Met Ile Pro
385 390 395 400
Lys Met Arg Gln Ala Leu Tyr Gln Asp Ile Gly Ile Arg Tyr Pro Gly
405 410 415
Ile His Val Arg Thr Asp Ser Pro Ser Leu Glu Gly Tyr Asp Tyr Met
920 425 430
Ile Leu Leu Asn Glu Val Pro Tyr Val Arg Gly Lys Ile Pro Pro His
435 440 445
His Val Leu Thr Asn Glu Val Glu Asp Asn Leu Ser Arg Tyr Asn Leu
450 455 460
Pro Phe Ile Thr Tyr Lys Asn Ala Ala Gly Leu Pro Ser Ala Trp Val
465 470 475 480
Ser Glu Asp Ala Lys Ala Ile Leu Glu Lys Ala Ala Ile Lys Tyr Trp
485 490 495
Thr Pro Leu Glu Val Ile Ile Leu His Leu Ser Tyr Phe Phe His Lys
500 505 510
Ser Ser Gln Glu Phe Leu Gly Ile Gln Glu Val Arg Ser Met Ile Glu
515 520 525
Phe Met Glu Arg Ser Phe Pro Asp Leu Val Lys Glu Val Thr Arg Leu
530 535 540
28
CA 02341637 2001-02-19
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Ile Pro Leu Gln Lys Leu Thr Glu Ile Phe Lys Arg Leu Val Gln Glu
545 550 555 560
Gln Tle Ser Ile Lys Asp Leu Arg Thr Ile Leu Glu Ser Leu Ser Glu
565 570 575
Trp Ala Gln Thr Glu Lys Asp Thr Val Leu Leu Thr Glu Tyr Val Arg
580 585 590
Ser Ser Leu Lys Leu Tyr Ile Ser Phe Lys Phe Ser Gln Gly Gln Ser
595 600 605
Ala Ile Ser Val Tyr Leu Leu Asp Pro Glu Ile Glu Glu Met Ile Arg
610 615 620
Gly Ala Ile Lys Gln Thr Ser Ala Gly Ser Tyr Leu Ala Leu Asp Pro
625 630 635 640
Asp Ser Val Asn Leu Ile Leu Lys Ser Met Arg Asn Thr Ile Thr Pro
645 650 655
Thr Pro Ala Gly Gly Gln Pro Pro Val Leu Leu Thr Ala Ile Asp Val
660 665 670
Arg Arg Tyr Val Arg Lys Leu Ile Glu Thr Glu Phe Pro Asp Ile Ala
675 680 685
Val Ile Ser Tyr Gln Glu Ile Leu Pro Glu Ile Arg Ile Gln Pro Leu
690 695 700
Gly Arg Ile Gln Ile Phe
705 710
<210> 9
<211> 1400
<212> DNA
<213> Chlamydia sp.
<220>
<221> CDS
<222> (101)..(1297)
<400> 9
gctgtgattt cttatcaaga aatcctacca gaaatccgca tccagccttt aggaagaatt 60
cagattttct aattgatacg ttgtcgctca taggaggcat atg gca gca tca gga 115
29
CA 02341637 2001-02-19
WO 00/11183 PCT/IB99/01449
Met Ala Ala Ser Gly
1 5
ggc aca ggt ggt tta gga ggc act cag ggt gtc aac ctt gca get gta 163_
Gly Thr Gly Gly Leu Gly Gly Thr Gln Gly Val Asn Leu A1a Ala Val
10 15 20
gaa get gca get gca aaa gca gat gca gca gaa gtt gta gcc agc caa 211
Glu Ala Ala Ala Ala Lys Ala Asp Ala Ala Glu Val Val Ala Ser Gln
25 30 35
gaa ggt tct gag atg aac atg att caa caa tct cag gac ctg aca aat 259
Glu Gly Ser Glu Met Asn Met Ile Gln Gln Ser Gln Asp Leu Thr Asn
40 45 50
ccc gca gca gca aca cgc acg aaa aaa aag gaa gag aag ttt caa act 307
Pro Ala Ala Ala Thr Arg Thr Lys Lys Lys Glu Glu Lys Phe Gln Thr
55 60 65
cta gaa tct cgg aaa aaa gga gaa get gga aag get gag aaa aaa tct 355
Leu Glu Ser Arg Lys Lys Gly Glu Ala Gly Lys Ala Glu Lys Lys Ser
70 75 80 85
gaa tct aca gaa gag aag cct gac aca gat ctt get gat aag tat get 403
Glu Ser Thr Glu Glu Lys Pro Asp Thr Asp Leu Ala Asp Lys Tyr Ala
90 95 100
tct ggg aat tct gaa atc tct ggt caa gaa ctt cgc ggc ctg cgt gat 451
Ser Gly Asn Ser Glu Ile Ser Gly Gln Glu Leu Arg Gly Leu Arg Asp
105 110 115
gca ata gga gac gat get tct cca gaa gac att ctt get ctt gta caa 499
Ala Ile Gly Asp Asp Ala Ser Pro Glu Asp Ile Leu Ala Leu Val Gln
120 125 130
gag aaa att aaa gac cca get ctg caa tcc aca get ttg gac tac ctg 547
Glu Lys Ile Lys Asp Pro Ala Leu Gln Ser Thr Ala Leu Asp Tyr Leu
135 140 145
gtt caa acg act cca ccc tcc caa ggt aaa tta aaa gaa gcg ctt atc 595
Val Gln Thr Thr Pro Pro Ser Gln Gly Lys Leu Lys Glu Ala Leu Ile
150 155 160 165
caa gca agg aat act cat acg gag caa ttc gga cga act get att ggt 643
Gln Ala Arg Asn Thr His Thr Glu Gln Phe Gly Arg Thr Ala Ile Gly
170 175 180
gcg aaa aac atc tta ttt gcc tct caa gaa tat gca gac caa ctg aat 691
30
CA 02341637 2001-02-19
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Ala Lys Asn Ile Leu Phe Ala Ser Gln Glu Tyr Ala Asp Gln Leu Asn
185 190 195
gtt tct cct tca ggg ctt cgc tct ttg tac tta gaa gtg act gga gac 739.
Val Ser Pro Ser Gly Leu Arg Ser Leu Tyr Leu Glu Val Thr Gly Asp
200 205 210
aca cat acc tgt gat cag cta ctt tct atg ctt caa gac cgc tat acc 787
Thr His Thr Cys Asp Gln Leu Leu Ser Met Leu Gln Asp Arg Tyr Thr
215 220 225
tac caa gat atg get att gtc agc tcc ttt cta atg aaa gga atg gea 835
Tyr Gln Asp Met Ala Ile Val Ser $er Phe Leu Met Lys Gly Met Ala
230 235 240 245
aca gaa tta aaa agg cag ggt ccc tac gta ccc agt gcg caa cta caa 883
Thr Glu Leu Lys Arg Gln Gly Pro Tyr Val Pro Ser Ala Gln Leu Gln
250 255 260
gtt ctc atg aca gaa act cgt aac ctg caa gca gtt ctt acc tcg tac 931
Val Leu Met Thr Glu Thr Arg Asn Leu Gln Ala Val Leu Thr Ser Tyr
265 270 275
gat tac ttt gaa agt cgc gtt cct att tta ctc gat agc tta aaa get 979
Asp Tyr Phe Glu Ser Arg Val Pro Ile Leu Leu Asp Ser Leu Lys Ala
280 285 290
gag gga atc caa act ect tct gat cta aac ttt gtg aag gta get gag 1027
Glu Gly Ile Gln Thr Pro Ser Asp Leu Asn Phe Val Lys Val Ala Glu
295 300 305
tcc tac cat aaa atc att aac gat aag ttc cca aca gca tct aaa gta 1075
Ser Tyr His Lys Ile Ile Asn Asp Lys Phe Pro Thr Ala Ser Lys Val
310 315 320 325
gaa cga gaa gtc cgc aat ctc ata gga gac gat gtt gat tct gtg acc 1123
Glu Arg Glu Val Arg Asn Leu Ile Gly Asp Asp Val Asp Ser Val Thr
330 335 340
ggt gtc ttg aac tta ttc ttt tet get tta cgt caa acg tcg tca egc 1171
Gly Val Leu Asn Leu Phe Phe Ser Ala Leu Arg Gln Thr Ser Ser Arg
345 350 355
ctt ttc tct tca gca gac aaa cgt cag caa tta gga get atg att get 1219
Leu Phe Ser Ser Ala Asp Lys Arg Gln Gln Leu Gly Ala Met Ile Ala
360 365 370
aat get tta gat get gta aat ata aac aat gaa gat tat cce aaa gca 1267
31
CA 02341637 2001-02-19
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Asn Ala Leu Asp Ala Val Asn Ile Asn Asn Glu Asp Tyr Pro Lys Ala
375 380 385
tca gac ttc cct aaa ccc tat cct tgg tca tgattaaaaa aggattgcca 1317
Ser Asp Phe Pro Lys Pro Tyr Pro Trp Ser
390 395
tgcaaaacca atacgagcaa ttactagaat ccttagcacc cctattaaat acgacacttg 1377
ctccagataa aaataactct tgt 1400
<210> 10
<211> 399
<212> PRT
<213> Chlamydia sp.
<400> 10
Met Ala Ala Ser Gly Gly Thr Gly Gly Leu Gly Gly Thr Gln Gly Val
1 5 10 15
Asn Leu Ala Ala Val Glu Ala Ala Ala Ala Lys Ala Asp Ala Ala Glu
20 25 30
Val Val Ala Ser Gln Glu Gly Ser Glu Met Asn Met Ile Gln Gln Ser
35 40 45
Gln Asp Leu Thr Asn Pro Ala Ala Ala Thr Arg Thr Lys Lys Lys Glu
50 55 60
Glu Lys Phe Gln Thr Leu Glu Ser Arg Lys Lys Gly Glu Ala Gly Lys
65 70 75 80
Ala Glu Lys Lys Ser Glu Ser Thr Glu Glu Lys Pro Asp Thr Asp Leu
85 90 95
Ala Asp Lys Tyr Ala Ser Gly Asn Ser Glu Ile Ser Gly Gln Glu Leu
100 105 110
Arg Gly Leu Arg Asp Ala Ile Gly Asp Asp Ala Ser Pro Glu Asp Ile
115 120 125
Leu Ala Leu Val Gln Glu Lys Ile Lys Asp Pro Ala Leu Gln Ser Thr
130 135 140
Ala Leu Asp Tyr Leu Val Gln Thr Thr Pro Pro Ser Gln Gly Lys Leu
145 150 155 160
32
CA 02341637 2001-02-19
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Lys Glu Ala Leu Ile Gln Ala Arg Asn Thr His Thr Glu Gln Phe Gly
165 170 175
Arg Thr Ala Ile Gly Ala Lys Asn Ile Leu Phe Ala Ser Gln Glu Tyr
180 185 190
Ala Asp Gln Leu Asn Val Ser Pro Ser Gly Leu Arg Ser Leu Tyr Leu
195 200 205
Glu Val Thr Gly Asp Thr His Thr Cys Asp Gln Leu Leu Ser Met Leu
210 215 220
Gln Asp Arg Tyr Thr Tyr Gln Asp Met Ala Ile Val Ser Ser Phe Leu
225 230 235 240
Met Lys Gly Met Ala Thr Glu Leu Lys Arg Gln Gly Pro Tyr Val Pro
245 250 255
Ser Ala Gln Leu Gln Val Leu Met Thr Glu Thr Arg Asn Leu Gln Ala
260 265 270
Val Leu Thr Ser Tyr Asp Tyr Phe Glu Ser Arg Val Pro Ile Leu Leu
275 280 285
Asp Ser Leu Lys Ala Glu Gly Ile Gln Thr Pro Ser Asp Leu Asn Phe
290 295 300
Val Lys Val Ala Glu Ser Tyr His Lys Ile Ile Asn Asp Lys Phe Pro
305 310 315 320
Thr Ala Ser Lys Val Glu Arg Glu Val Arg Asn Leu Ile Gly Asp Asp
325 330 335
Val Asp Ser Val Thr Gly Val Leu Asn Leu Phe Phe Ser Ala Leu Arg
340 395 350
Gln Thr Ser Ser Arg Leu Phe Ser Ser Ala Asp Lys Arg Gln Gln Leu
355 360 365
Gly Ala Met Ile Ala Asn Ala Leu Asp Ala Val Asn Ile Asn Asn Glu
370 375 380
Asp Tyr Pro Lys Ala Ser Asp Phe Pro Lys Pro Tyr Pro Trp Ser
385 390 395
<210> 11
<211> 650
33
CA 02341637 2001-02-19
WO 00/11183 PCT/IB99/01449
<212> DNA
<213> Chlamydia sp.
<220>
<221> CDS
<222> (101)..(541)
<400> 11
gctaatgctt tagatgctgt aaatataaac aatgaagatt atcccaaagc atcagacttc 60
cctaaaccct atccttggtc atgattaaaa aaggattgcc atg caa aac caa tac 115
Met Gln Asn Gln Tyr
1 5
gag caa tta cta gaa tcc tta gca ccc cta tta aat acg aca ctt get 163
Glu Gln Leu Leu Glu Ser Leu Ala Pro Leu Leu Asn Thr Thr Leu Ala
10 15 20
cca gat aaa aat aac tct tgt tta atc cgt ttc agc gat acc cat gtc 211
Pro Asp Lys Asn Asn Ser Cys Leu Ile Arg Phe Ser Asp Thr His Val
25 30 35
cct gtg caa ata gaa gaa gat gga aat tcc gga gat ctt gca gta tcg 259
Pro Val Gln Ile Glu Glu Asp Gly Asn Ser Gly Asp Leu Ala Val Ser
40 45 50
aca cta cta ggt act ctt cct gaa aac gta ttt cgc gag cgt att ttc 307
Thr Leu Leu Gly Thr Leu Pro Glu Asn Val Phe Arg Glu Arg Ile Phe
55 60 65
aaa get get ctc tct gta aat ggc tcg ttc caa tcc agc atc aag gga 355
Lys Ala Ala Leu Ser Val Asn Gly Ser Phe Gln Ser Ser Ile Lys Gly
70 75 80 85
att cta ggc tac ggt gag gtc act caa cag ctc tat ctt tca gat atc 403
Ile Leu Gly Tyr Gly Glu Val Thr Gln Gln Leu Tyr Leu Ser Asp Ile
90 95 100
ctg agt atg aac tac cta aat gga gaa aag tta ttc gag tat ctc aag 451
Leu Ser Met Asn Tyr Leu Asn Gly Glu Lys Leu Phe Glu Tyr Leu Lys
105 110 115
ctc ttt tct ttg cat get aag att tgg atg gaa tcc cta aga aca ggg 499
Leu Phe Ser Leu His Ala Lys Ile Trp Met Glu Ser Leu Arg Thr Gly
120 125 130
aat ctt cct gac ctt cat gtt ttg gga atc tac tac gtc gcg 541
Asn Leu Pro Asp Leu His Val Leu Gly Ile Tyr Tyr Val Ala
34
CA 02341637 2001-02-19
WO 00/11183 PCT/IB99/01449
135 140 145
tgaatgtttt aaaatacaca aaacactcac cctcagcaca tgcttggaaa cttataggaa 601
cctctcctaa acacgggatt tatctcccac tattttcaat acacacaaa 650
<210> 12
<211> 147
<212> PRT
<213> Chlamydia sp.
<400> 12
Met Gln Asn Gln Tyr Glu Gln Leu Leu Glu Ser Leu Ala Pro Leu Leu
1 5 10 15
Asn Thr Thr Leu Ala Pro Asp Lys Asn Asn Ser Cys Leu Ile Arg Phe
2D 25 30
Ser Asp Thr His Val Pro Val Gln Ile Glu Glu Asp Gly Asn Ser Gly
35 40 45
Asp Leu Ala Val Ser Thr Leu Leu Gly Thr Leu Pro Glu Asn Val Phe
50 55 60
Arg Glu Arg Ile Phe Lys Ala Ala Leu Ser Val Asn Gly Ser Phe Gln
65 ,70 75 80
Ser Ser Ile Lys Gly Ile Leu Gly Tyr Gly Glu Val Thr Gln Gln Leu
85 90 95
Tyr Leu Ser Asp Ile Leu Ser Met Asn Tyr Leu Asn Gly Glu Lys Leu
100 105 110
Phe Glu Tyr Leu Lys Leu Phe Ser Leu His Ala Lys Ile Trp Met Glu
115 120 125
Ser Leu Arg Thr Gly Asn Leu Pro Asp Leu His Val Leu Gly Ile Tyr
130 135 140
Tyr Val Ala
145
<210>13
<211>3050
<212>DNA
<213>Chlamydia
sp.
35
CA 02341637 2001-02-19
WO 00/11183 PCT/IB99/01449
<220>
<221> CDS
<222> (101) . . (2935)
<400> 13
aaatgcttct agtctagggg aacttttttc tatttctacg aagggagtgt cttgtctaga 60
actccatagg gagatcgcac gctaaatatg agaatataat atg aag act tca gtt 115
Met Lys Thr Ser Val
1 5
tct atg ttg ttg gcc ctg ctt tgc tcg ggg get agc tct att gta ctc 163
Ser Met Leu Leu Ala Leu Leu Cys Ser Gly Ala Ser Ser Ile Val Leu
10 15 20
cat gcc gca acc act cca cta aat cct gaa gat ggg ttt att ggg gag 211
His Ala Ala Thr Thr Pro Leu Asn Pro Glu Asp Gly Phe Ile Gly Glu
25 30 35
ggc aat aca aat act ttt tct ccg aaa tct aca acg gat get gca gga 259
Gly Asn Thr Asn Thr Phe Ser Pro Lys Ser Thr Thr Asp Ala Ala Gly
40 45 SO
act acc tac tct ctc aca gga gag gtt ctg tat ata gat ccg ggg aaa 307
Thr Thr Tyr Ser Leu Thr Gly Glu Val Leu Tyr Ile Asp Pro Gly Lys
55 60 65
ggt ggt tca att aca gga act tgc ttt gta gaa act get ggc gat ctt 355
Gly Gly Ser Ile Thr Gly Thr Cys Phe Val Glu Thr Ala Gly Asp Leu
70 75 80 85
aca ttt tta ggt aat gga aat acc cta aag ttc ctg tcg gta gat gca 403
Thr Phe Leu Gly Asn Gly Asn Thr Leu Lys Phe Leu Ser Val Asp Ala
90 95 100
ggt get aat atc gcg gtt get cat gta caa gga agt aag aat tta agc 451
Gly Ala Asn Ile Ala Val Ala His Val Gln Gly Ser Lys Asn Leu Ser
105 110 115
ttc aca gat ttc ctt tct ctg gtg atc aca gaa tct cca aaa tcc get 499
Phe Thr Asp Phe Leu Ser Leu Val Ile Thr Glu Ser Pro Lys Ser Ala
120 12S 130
gtt act aca gga aaa ggt agc cta gtc agt tta ggt gca gtc caa ctg 547
Val Thr Thr Gly Lys Gly Ser Leu Val Ser Leu Gly Ala Val Gln Leu
135 140 145
36
CA 02341637 2001-02-19
WO 00/11183 PCT/IB99/01449
caa gat ata aac act cta gtt ctt aca agc aat gcc tct gtc gaa gat 595
Gln Asp Ile Asn Thr Leu Val Leu Thr Ser Asn Ala Ser Val Glu Asp
150 155 160 165
ggt ggc gtg att aaa gga aac tcc tgc ttg att cag gga atc aaa aat 643
Gly Gly Val Ile Lys Gly Asn Ser Cys Leu Ile Gln Gly Ile Lys Asn
170 175 180
agt gcg att ttt gga caa aat aca tct tcg aaa aaa gga ggg gcg atc 691
Sex Ala Ile Phe Gly Gln Asn Thr Ser Ser Lys Lys Gly Gly Ala Ile
185 190 195
tcc acg act caa gga ctt acc ata gag aat aac tta ggg acg cta aag 739
Ser Thr Thr Gln Gly Leu Thr Ile Glu Asn Asn Leu Gly Thr Leu Lys
200 205 210
ttc aat gaa aac aaa gca gtg acc tca gga ggc gcc tta gat tta gga 787
Phe Asn Glu Asn Lys Ala Val Thr Ser Gly Gly Ala Leu Asp Leu Gly
215 220 225
gcc gcg tct aca ttc act gcg aac cat gag ttg ata ttt tca caa aat 835
Ala Ala Ser Thr Phe Thr Ala Asn His Glu Leu Ile Phe Ser Gln Asn
230 235 240 245
aag act tct ggg aat get gca aat ggc gga gcc ata aat tgc tca ggg 883
Lys Thr Ser Gly Asn Ala Ala Asn Gly Gly Ala Ile Asn Cys Ser Gly
250 255 260
gac ctt aca ttt act gat aac act tct ttg tta ctt caa gaa aat agc 931
Asp Leu Thr Phe Thr Asp Asn Thr Ser Leu Leu Leu Gln Glu Asn Ser
265 270 275
aca atg cag gat ggt gga get ttg tgt agc aca gga acc ata agc att 979
Thr Met Gln Asp Gly Gly Ala Leu Cys Ser Thr Gly Thr Ile Ser Ile
280 285 290
acc ggt agt gat tct atc aat gtg ata gga aat act tca gga caa aaa 1027
Thr Gly Ser Asp Ser Ile Asn Val Ile Gly Asn Thr Ser Gly Gln Lys
2g5 300 305
gga gga gcg att tct gca get tct ctc aag att ttg gga ggg cag gga 1075
Gly Gly Ala Ile Ser Ala Ala Ser Leu Lys Ile Leu Gly Gly Gln Gly
310 315 320 325
ggc get ctc ttt tct aat aac gta gtg act cat gcc acc cct cta gga 1123
Gly Ala Leu Phe Ser Asn Asn Val Val Thr His Ala Thr Pro Leu Gly
330 335 340
37
CA 02341637 2001-02-19
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ggt gcc att ttt atc aac aca gga gga tcc ttg cag ctc ttc act caa 1171
Gly Ala Ile Phe Ile Asn Thr Gly Gly Ser Leu Gln Leu Phe Thr Gln
345 350 355
gga ggg gat atc gta ttc gag ggg aat cag gtc act aca aca get cca 1219
Gly Gly Asp Ile Val Phe Glu Gly Asn Gln Val Thr Thr Thr Ala Pro
360 365 370
aat get acc act aag aga aat gta att cac ctc gag agc acc gcg aag 1267
Asn Ala Thr Thr Lys Arg Asn Val Ile His Leu Glu Ser Thr Ala Lys
375 380 385
tgg aeg gga ctt get gca agt caa ggt aac get atc tat ttc tat gat 1315
Trp Thr Gly Leu Ala Ala Ser Gln Gly Asn Ala Ile Tyr Phe Tyr Asp
390 395 400 405
ccc att acc acc aac gat acg gga gca agc gat aac tta cgt atc aat 1363
Pro Ile Thr Thr Asn Asp Thr Gly Ala Ser Asp Asn Leu Arg Ile Asn
410 415 420
gag gtc agt gca aat caa aag ctc tcg gga tct ata gta ttt tct gga 1411
Glu Val Ser Ala Asn Gln Lys Leu Ser Gly Ser Ile Val Phe Ser Gly
425 430 435
gag aga ttg tcg aca gca gaa get ata get gaa aat ctt act tcg agg 1459
Glu Arg Leu Ser Thr Ala Glu Ala Ile Ala Glu Asn Leu Thr Ser Arg
440 445 450
atc aac cag cct gtc act tta gta gag ggg agc tta gta ctt aaa cag 1507
Ile Asn Gln Pro Val Thr Leu Val Glu Gly Ser Leu Val Leu Lys Gln
455 460 465
gga gtg acc ttg atc aca caa gga ttc tcg cag gag cca gaa tcc acg 1555
Gly Val Thr Leu Ile Thr Gln Gly Phe Ser Gln Glu Pro Glu Ser Thr
470 475 480 485
ctt ctt ttg gat ctg ggg acc tca tta aaa get tct aea gaa gat att 1603
Leu Leu Leu Asp Leu Gly Thr Ser Leu Lys Ala Ser Thr Glu Asp Ile
490 495 500
gtc atc aca aat tta tct ata aat gcc gat acc att tac gga aag aat 1651
Va1 Ile Thr Asn Leu Ser Ile Asn Ala Asp Thr Ile Tyr Gly Lys Asn
505 510 515
cct atc aat att gta get tca gca gcg aat aag aac att acc cta aca 1699
Pro Ile Asn Ile Val Ala Ser Ala Ala Asn Lys Asn Ile Thr Leu Thr
520 525 530
38
CA 02341637 2001-02-19
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gga acc tta gca ctt gta aat gca gat gga get ttc tat gag aac cat 1747
Gly Thr Leu Ala Leu Val Asn Ala Asp Gly Ala Phe Tyr Glu Asn His
535 540 545
acc ttg caa gac tct caa gac tat agc ttt gta aag tta tct cca gga 1795
Thr Leu Gln Asp Ser Gln Asp Tyr Ser Phe Val Lys Leu Ser Pro Gly
550 555 560 565
gcg gga ggg act ata att act caa gat get tct cag aag cct ctt gaa 1843
Ala Gly Gly Thr Ile Ile Thr Gln Asp Ala Ser Gln Lys Pro Leu Glu
570 575 580
gta get cct tct aga cca cat tat ggc tat caa gga cat tgg aat gtg 1891
Val Ala Pro Ser Arg Pro His Tyr Gly Tyr Gln Gly His Trp Asn Val
585 590 595
caa gtc atc cca gga acg gga act caa ccg agc cag gca aat tta gaa 1939
Gln Val Ile Pro Gly Thr Gly Thr Gln Pro Ser Gln Ala Asn Leu Glu
600 605 610
tgg gtg cgg aca gga tac ctt ccg aat ccc gaa cgg caa gga tct tta 1987
Trp Val Arg Thr Gly Tyr Leu Pro Asn Pro Glu Arg Gln Gly Ser Leu
615 620 625
gtt ccc aat agc ctg tgg ggt tct ttt gtt gat cag cgt get atc caa 2035
Val Pro Asn Ser Leu Trp Gly Ser Phe Val Asp Gln Arg Ala Ile Gln
630 635 640 645
gaa atc atg gta aat agt agc caa atc tta tgt cag gaa cgg gga gtc 2083
Glu Ile Met Val Asn Ser Ser Gln Ile Leu Cys Gln Glu Arg Gly Val
650 655 660
tgg gga get gga att get aat ttc cta cat aga gat aaa att aat gag 2131
Trp Gly Ala Gly Ile Ala Asn Phe Leu His Arg Asp Lys Ile Asn Glu
665 670 675
cac ggc tat cgc cat agc ggt gtc ggt tat ctt gtg gga gtt ggc act 2179
His Gly Tyr Arg His Ser Gly Val Gly Tyr Leu Val Gly Val Gly Thr
680 685 690
cat get ttt tct gat get acg ata aat gcg get ttt tgc cag ctc ttc 2227
His Ala Phe Ser Asp Ala Thr Ile Asn Ala Ala Phe Cys Gln Leu Phe
695 700 705
agt aga gat aaa gac tac gta gta tcc aaa aat cat gga act agc tac 2275
Ser Arg Asp Lys Asp Tyr Val Val Ser Lys Asn His Gly Thr Ser Tyr
71p 715 720 725
39
CA 02341637 2001-02-19
WO 00/11183 PCT/IB99/01449
tca ggg gtc gta ttt ctt gag gat acc cta gag ttt aga agt cca cag 2323
Ser Gly Val Val Phe Leu Glu Asp Thr Leu Glu Phe Arg Ser Pro Gln
730 735 740
gga ttc tat act gat agc tcc tca gaa get tgc tgt aac caa gtc gtc 2371
Gly Phe Tyr Thr Asp Ser Ser Ser Glu Ala Cys Cys Asn Gln Val Val
745 750 755
act ata gat atg cag ttg tct tac agc cat aga aat aat gat atg aaa 2419
Thr Ile Asp Met Gln Leu Ser Tyr Ser His Arg Asn Asn Asp Met Lys
760 765 770
acc aaa tac acg aca tat cca gaa get cag gga tct tgg gca aat gat 2467
Thr Lys Tyr Thr Thr Tyr Pro Glu Ala Gln Gly Ser Trp Ala Asn Asp
775 780 785
gtt ttt ggt ctt gag ttt gga gcg act aca tac tac tac cct~aac agt 2515
Val Phe Gly Leu Glu Phe Gly Ala Thr Thr Tyr Tyr Tyr Pro Asn Ser
790 795 800 805
act ttt tta ttt gat tac tac tct ccg ttt ctc agg ctg cag tgc acc 2563
Thr Phe Leu Phe Asp Tyr Tyr Ser Pro Phe Leu Arg Leu Gln Cys Thr
810 815 820
tat get cac cag gaa gac ttc aaa gag aca gga ggt gag gtt cgt cac 2611
Tyr Ala His Gln Glu Asp Phe Lys Glu Thr Gly Gly Glu Val Arg His
825 830 835
ttt act agc gga gat ctt ttc aat tta gca gtt cct att ggc gtg aag 2659
Phe Thr Ser Gly Asp Leu Phe Asn Leu Ala Val Pro Ile Gly Val Lys
840 845 850
ttt gag aga ttt tca gac tgt aaa agg gga tct tat gaa ctt acc ctt 2707
Phe Glu Arg Phe Ser Asp Cys Lys Arg Gly Ser Tyr Glu Leu Thr Leu
855 860 865
get tat gtt cct gat gtg att cgc aaa gat ccc aag agc acg gca aca 2755
Ala Tyr Vai Pro Asp Val Ile Arg Lys Asp Pro Lys Ser Thr Ala Thr
870 875 880 885
ttg get agt gga get acg tgg agc acc cac gga aac aat ctc tcc aga 2803
Leu Ala Ser Gly Ala Thr Trp Ser Thr His Gly Asn Asn Leu Ser Arg
890 895 900
caa gga tta caa ctg cgt tta ggg aac cac tgt ctc ata aat cct gga 2851
Gln Gly Leu Gln Leu Arg Leu Gly Asn His Cys Leu Ile Asn Pro Gly
905 910 915
40
CA 02341637 2001-02-19
WO 00/11183 PCT/IB99/01449
att gag gtg ttc agt cac gga get att gaa ttg cgg gga tcc tct cgt 2899
Ile Glu Val Phe Ser His Gly Ala Ile Glu Leu Arg Gly Ser Ser Arg
920 925 930
aat tat aac atc aat ctc ggg ggt aaa tac cga ttt taatagggaa 2945
Asn Tyr Asn Ile Asn Leu Gly Gly Lys Tyr Arg Phe
935 940 945
ctgagagttc cttattgtag agagtattta caaggatttt gagacgaaag agatttcttg 3005
ttgtaggtat ctctccacag ggtctagttc aatttctata gttca 3050
<210> 14
<211> 945
<212> PRT
<213> Chlamydia sp.
<400> 14
Met Lys Thr Ser Val Ser Met Leu Leu Ala Leu Leu Cys Ser Gly Ala
1 5 10 15
Ser Ser Ile Val Leu His Ala Ala Thr Thr Pro Leu Asn Pro Glu Asp
20 25 30
Gly Phe Ile Gly Glu Gly Asn Thr Asn Thr Phe Ser Pro Lys Ser Thr
35 90 45
Thr Asp Ala Ala Gly Thr Thr Tyr Ser Leu Thr Gly Glu Val Leu Tyr
50 55 60
Ile Asp Pro Gly Lys Gly Gly Ser Ile Thr Gly Thr Cys Phe Val Glu
65 70 75 80
Thr Ala Gly Asp Leu Thr Phe Leu Gly Asn Gly Asn Thr Leu Lys Phe
85 90 95
Leu Ser Val Asp Ala Gly Ala Asn Ile Ala Val Ala His Val Gln Gly
100 105 110
Ser Lys Asn Leu Ser Phe Thr Asp Phe Leu Ser Leu Val Ile Thr Glu
115 120 125
Ser Pro Lys Ser Ala Val Thr Thr Gly Lys Gly Ser Leu Val Ser Leu
130 135 140
Gly Ala Val Gln Leu Gln Asp Ile Asn Thr Leu Val Leu Thr Ser Asn
145 150 155 160
41
CA 02341637 2001-02-19
WO 00/11183 PCT/IB99/01449
Ala Ser Val Glu Asp Gly Gly Val Ile Lys Gly Asn Ser Cys Leu Ile
165 170 175
Gln Gly Ile Lys Asn Ser Ala Ile Phe Gly Gln Asn Thr Ser Ser Lys
180 185 190
Lys Gly Gly Ala Ile Ser Thr Thr Gln Gly Leu Thr Ile Glu Asn Asn
195 200 205
Leu Gly Thr Leu Lys Phe Asn Glu Asn Lys Ala Val Thr Ser Gly Gly
210 215 220
Ala Leu Asp Leu Gly Ala Ala Ser Thr Phe Thr Ala Asn His Glu Leu
225 230 235 240
Ile Phe Ser Gln Asn Lys Thr Ser Gly Asn Ala Ala Asn Gly Gly Ala
245 250 255
Ile Asn Cys Ser Gly Asp Leu Thr Phe Thr Asp Asn Thr Ser Leu Leu
260 265 270
Leu Gln Glu Asn Ser Thr Met Gln Asp Gly Gly Ala Leu Cys Ser Thr
275 280 285
Gly Thr Ile Ser Ile Thr Gly Ser Asp Ser Ile Asn Val Ile Gly Asn
290 295 300
Thr Ser Gly Gln Lys Gly Gly Ala Ile Ser Ala Ala Ser Leu Lys Ile
305 310 315 320
Leu Gly Gly Gln Gly Gly Ala Leu Phe Ser Asn Asn Val Val Thr His
325 330 335
Ala Thr Pro Leu Gly Gly Ala Ile Phe Ile Asn Thr Gly Gly Ser Leu
340 345 350
Gln Leu Phe Thr Gln Gly Gly Asp Ile Val Phe Glu Gly Asn Gln Val
355 360 365
Thr Thr Thr Ala Pro Asn Ala Thr Thr Lys Arg Asn Val Ile His Leu
370 375 380
Glu Ser Thr Ala Lys Trp Thr Gly Leu Ala Ala Ser Gln Gly Asn Ala
385 390 395 400
Ile Tyr Phe Tyr Asp Pro Ile Thr Thr Asn Asp Thr Gly Ala Ser Asp
405 410 415
42
CA 02341637 2001-02-19
WO 00/11183 PCT/IB99/01449
Asn Leu Arg Ile Asn Glu Val Ser Ala Asn Gln Lys Leu Ser Gly Ser
420 425 430
Ile Val Phe Ser Gly Glu Arg Leu Ser Thr Ala Glu Ala Ile Ala Glu
435 440 445
Asn Leu Thr Ser Arg Ile Asn Gln Pro Val Thr Leu Val Glu Gly Ser
450 455 460
Leu Val Leu Lys Gln Gly Val Thr Leu Ile Thr Gln Gly Phe Ser Gln
465 470 475 480
Glu Pro Glu Ser Thr Leu Leu Leu Asp Leu Gly Thr Ser Leu Lys Ala
485 490 495
Ser Thr Glu Asp Ile Val Ile Thr Asn Leu Ser Ile Asn Ala Asp Thr
500 505 510
Ile Tyr Gly Lys Asn Pro Ile Asn Ile Val Ala Ser Ala Ala Asn Lys
515 520 525
Asn Ile Thr Leu Thr Gly Thr Leu Ala Leu Val Asn Ala Asp Gly Ala
530 535 540
Phe Tyr Glu Asn His Thr Leu Gln Asp Ser Gln Asp Tyr Ser Phe Val
545 550 555 560
Lys Leu Ser Pro Gly Ala Gly Gly Thr Ile Ile Thr Gln Asp Ala Ser
565 570 575
Gln Lys Pro Leu Glu Val Ala Pro Ser Arg Pro His Tyr Gly Tyr Gln
580 S85 590
Gly His Trp Asn Val Gln Val Ile Pro Gly Thr Gly Thr Gln Pro Ser
595 600 605
Gln Ala Asn Leu Glu Trp Val Arg Thr Gly Tyr Leu Pro Asn Pro Glu
610 615 620
Arg Gln Gly Ser Leu Val Pro Asn Ser Leu Trp Gly Ser Phe Val Asp
625 630 635 640
Gln Arg Ala Ile Gln Glu Ile Met Val Asn Ser Ser Gln Ile Leu Cys
645 650 655
Gln Glu Arg Gly Val Trp Gly Ala Gly Ile Ala Asn Phe Leu His Arg
660 665 670
43
CA 02341637 2001-02-19
WO 00/11183 PCT/IB99/01449
Asp Lys Ile Asn Glu His Gly Tyr Arg His Ser Gly Val Gly Tyr Leu
675 680 685
Val Gly Val Gly Thr His Ala Phe Ser Asp Ala Thr Ile Asn Ala Ala
690 695 700
Phe Cys Gln Leu Phe Ser Arg Asp Lys Asp Tyr Val Val Ser Lys Asn
705 710 715 720
His Gly Thr Ser Tyr Ser Gly Val Val Phe Leu Glu Asp Thr Leu Glu
725 730 735
Phe Arg Ser Pro Gln Gly Phe Tyr Thr Asp Ser Ser Ser Glu Ala Cys
740 745 750
Cys Asn Gln Val Val Thr Ile Asp Met Gln Leu Ser Tyr Ser His Arg
755 760 765
Asn Asn Asp Met Lys Thr Lys Tyr Thr Thr Tyr Pro Glu Ala Gln Gly
770 775 780
Ser Trp Ala Asn Asp Val Phe Gly Leu Glu Phe Gly Ala Thr Thr Tyr
785 790 795 800
Tyr Tyr Pro Asn Ser Thr Phe Leu Phe Asp Tyr Tyr Ser Pro Phe Leu
805 810 815
Arg Leu Gln Cys Thr Tyr Ala His Gln Glu Asp Phe Lys Glu Thr Gly
g2p 825 830
Gly Glu Val Arg His Phe Thr Ser Gly Asp Leu Phe Asn Leu Ala Val
835 840 845
Pro Ile Gly Val Lys Phe Glu Arg Phe Ser Asp Cys Lys Arg Gly Ser
850 855 860
Tyr Glu Leu Thr Leu Ala Tyr Val Pro Asp Val Ile Arg Lys Asp Pro
865 870 875 880
Lys Ser Thr Ala Thr Leu Ala Ser Gly Ala Thr Trp Ser Thr His Gly
885 890 895
Asn Asn Leu Ser Arg Gln Gly Leu Gln Leu Arg Leu Gly Asn His Cys
900 905 910
Leu Ile Asn Pro Gly Ile Glu Val Phe Ser His Gly Ala Ile Glu Leu
915 920 925
44
~:,Y", . , ., .a
CA 02341637 2001-02-19
WO 00/11183 PCT/IB99/01449
Arg Gly Ser Ser Arg Asn Tyr Asn Ile Asn Leu Gly Gly Lys Tyr Arg
930 935 940
Phe
945
<210> 15
<211> 2957
<212> DNA
<213> Chlamydia sp.
<220>
<221> CDS
<222> (101)..(2854)
<400> 15
ttgaaaaaaa gagagttacg gcagctgtaa agtttttaat attgctccct ttgttccatt 60
tatgtagcgt tcagactttg cactaaaacg agggtgtcat atg aga tcg tct ttt 115
Met Arg Ser Ser Phe
1 5
tcc ttg tta tta ata tct tca tct cta gcc ttt cct ctc tta atg agt 163
Ser Leu Leu Leu Ile Ser Ser Ser Leu Ala Phe Pro Leu Leu Met Ser
10 15 20
gtt tct gca gat get gcc gat ctc aca tta ggg agt cgt gac agt tat 211
Val Ser Ala Asp Ala Ala Asp Leu Thr Leu Gly Ser Arg Asp Ser Tyr
25 30 35
aat ggt gat aca agc acc aca gaa ttt act cct aaa gcg gca act tct 259
Asn Gly Asp Thr Ser Thr Thr Glu Phe Thr Pro Lys Ala Ala Thr Ser
40 45 50
gat get agt ggc acg acc tat att ctc gat ggg gat gtc tcg ata agc 307
Asp Ala Ser Gly Thr Thr Tyr Ile Leu Asp Gly Asp Val Ser Ile Ser
55 60 65
caa gca ggg aaa caa acg agc tta acc aca agt tgt ttt tct aac act 355
Gln Ala Gly Lys Gln Thr Ser Leu Thr Thr Ser Cys Phe Ser Asn Thr
70 75 80 85
gca gga aat ctt acc ttc tta ggg aac gga ttt tct ctt cat ttt gac 403
Ala Gly Asn Leu Thr Phe Leu Gly Asn Gly Phe Ser Leu His Phe Asp
90 95 100
45
CA 02341637 2001-02-19
WO 00/11183 PCT/IB99/01449
aat att att tcg tct act gtt gca ggt gtt gtt gtt agc aat aca gca 451
Asn Ile Ile Ser Ser Thr Val Ala Gly Val Val Val Ser Asn Thr Ala
105 110 115
get tct ggg att acg aaa ttc tca gga ttt tca act ctt cgg atg ctt 499
Ala Ser Gly Ile Thr Lys Phe Ser Gly Phe Ser Thr Leu~rg Met Leu
120 125 130
gca get cct agg acc aca ggt aaa gga gcc att aaa att acc gat ggt 547
Ala Ala Pro Arg Thr Thr Gly Lys Gly Ala Ile Lys Ile Thr Asp Gly
135 140 145
ctg gtg ttt gag agt ata ggg aat ctt gat ctt aat gaa aat gcc tct 595
Leu Val Phe Glu Ser Ile Gly Asn Leu Asp Leu Asn Glu Asn Ala Ser
150 155 160 165
agt gaa aat ggg gga gcc atc aat acg aag act ttg tct ttg act ggg 693
Ser Glu Asn Gly Gly Ala Ile Asn Thr Lys Thr Leu Ser Leu Thr Gly
170 175 180
agt acg cgg ttt gta gcg ttc ctt ggc aat agc tcg tcg caa caa ggg 691
Ser Thr Arg Phe Val Ala Phe Leu Gly Asn Ser Ser Ser Gln Gln Gly
185 190 195
gga gcg atc tat get tct ggt gac tct gtg att tct gag aat gca gga 739
Gly Ala Ile Tyr Ala Ser Gly Asp Ser Val Ile Ser Glu Asn Ala Gly
200 205 210
atc ttg agc ttc gga aac aac agt gcg aca aca tca gga ggc gcg atc 787
Ile Leu Ser Phe Gly Asn Asn Ser Ala Thr Thr Ser Gly Gly Ala Ile
215 220 225
tct get gaa ggg aac ctt gtg atc tcc aat aac caa aat atc ttt ttc 835
Ser Ala Glu Gly Asn Leu Val Ile Ser Asn Asn Gln Asn Ile Phe Phe
230 235 240 245
gat ggc tgc aaa gca act aca aat ggc gga get att gat tgt aac aaa 883
Asp Gly Cys Lys Ala Thr Thr Asn Gly Gly Ala Ile Asp Cys Asn Lys
250 255 260
gca ggg gcg aac cca gac cct atc ttg act ctt tca gga aat gag agc 931
Ala Gly Ala Asn Pro Asp Pro Ile Leu Thr Leu Ser Gly Asn Glu Ser
265 270 275
ctg cat ttt ctg aat aac aca gca gga aat agt gga ggt gcg att tat 979
Leu His Phe Leu Asn Asn Thr Ala Gly Asn Ser Gly Gly Ala Ile Tyr
280 285 290
46
CA 02341637 2001-02-19
WO 00/11183 PCT/IB99/01449
acc aaa aaa ttg gtg tta tcc tca gga cga gga gga gtg tta ttt tct 1027
Thr Lys Lys Leu Val Leu Ser Ser Gly Arg Gly Gly Val Leu Phe Ser
295 300 305
aac aac aaa get gcg aat get act cct aaa gga ggg gca att gcg att 1075
Asn Asn Lys Ala Ala Asn Ala Thr Pro Lys Gly Gly Ala Ile Ala Ile
310 315 320 325
cta gat tct gga gag att agc att tct gca gat ctc ggc aat atc att 1123
Leu Asp Ser Gly Glu Ile Ser Ile Ser Ala Asp Leu Gly Asn Ile Ile
330 335 340
ttc gag ggc aat act acg agc act aca gga agt cct gcg agt gtg acc 1171
Phe Glu Gly Asn Thr Thr Ser Thr Thr Gly Ser Pro Ala Ser Val Thr
345 350 355
aga aat get ata gat ctt gca tcg aat gca aaa ttt tta aat ctc cga 1219
Arg Asn Ala Ile Asp Leu Ala Ser Asn Ala Lys Phe Leu Asn Leu Arg
360 365 370
gcg act cgg gga aat aaa gtt att ttc tat gat cct atc acg agc tca 1267
Ala Thr Arg Gly Asn Lys Val Ile Phe Tyr Asp Pro Ile Thr Ser Ser
375 380 385
gga get act gat aag ctc tct ttg aat aaa get gac gca gga tct gga 1315
Gly Ala Thr Asp Lys Leu Ser Leu Asn Lys Ala Asp Ala Gly Ser Gly
390 395 400 405
aat acc tat gaa ggc tac atc gtt ttc tct gga gag aaa ctc tca gaa 1363
Asn Thr Tyr Glu Gly Tyr Ile Val Phe Ser Gly Glu Lys Leu Ser Glu
410 415 420
gag gaa ctt aag aaa cct gac aat ctg aag tct aca ttt aca cag get 1411
Glu Glu Leu Lys Lys Pro Asp Asn Leu Lys Ser Thr Phe Thr Gln Ala
425 430 435
gta gag ctt get gca ggt gcc tta gta ttg aaa gat gga gtg act gta 1459
Val Glu Leu Ala Ala Gly Ala Leu Val Leu Lys Asp Gly Val Thr Val
440 445 450
gtt gca aat act ata acg cag gtc gag gga tcg aaa gtc gtt atg gat 1507
Val Ala Asn Thr Ile Thr Gln Val Glu Gly Ser Lys Val Val Met Asp
455 460 465
gga ggg act act ttt gag gca agc get gag ggg gtc act ctc aat ggc 1555
Gly Gly Thr Thr Phe Glu Ala Ser Ala Glu Gly Val Thr Leu Asn Gly
470 475 480 485
47
CA 02341637 2001-02-19
WO 00/11183 PCT/IB99/01449
cta gcc att aat ata gat tcc tta gat ggg aca aat aaa get atc att 1603
Leu Ala Ile Asn Ile Asp Ser Leu Asp Gly Thr Asn Lys Ala Ile Ile
490 495 500
aag gcg acg gca gca agt aag gat gtt gcc tta tca ggg cct atc atg 1651
Lys Ala Thr Ala Ala Ser Lys Asp Val Ala Leu Ser Gly Pro Ile Met
505 510 515
ctt gta gat get cag ggg aac tat tat gag cat cat aat ctc agt caa 1699
Leu Val Asp Ala Gln Gly Asn Tyr Tyr Glu His His Asn Leu Ser Gln
520 525 530
cag cag gtc ttt cct tta ata gag ctt tct gca caa gga acg atg act 1747
Gln Gln Val Phe Pro Leu Ile Glu Leu Ser Ala Gln Gly Thr Met Thr
535 540 545
act aca gat atc ccc gat acc cca att cta aat act acg aat cac tat 1795
Thr Thr Asp Ile Pro Asp Thr Pro Ile Leu Asn Thr Thr Asn His Tyr
550 555 560 565
ggg tat caa ggg aac tgg aat att gtt tgg gtc gac gat gca act gca 1843
Gly Tyr Gln Gly Asn Trp Asn Ile Val Trp Val Asp Asp Ala Thr Ala
570 575 580
aaa aca aaa aat get acc tta act tgg act aaa aca gga tac aag ccg 1891
Lys Thr Lys Asn Ala Thr Leu Thr Trp Thr Lys Thr Gly Tyr Lys Pro
585 590 595
aat cca gaa cgt cag gga cct ttg gtt cct aat agc ctg tgg ggt tct 1939
Asn Pro Glu Arg Gln Gly Pro Leu Val Pro Asn Ser Leu Trp Gly Ser
600 605 610
ttt gtc gat gtc cgc tcc att cag agc ctc atg gac cgg agc aca agt 1987
Phe Val Asp Val Arg Ser Ile Gln Ser Leu Met Asp Arg Ser Thr Ser
615 620 625
tcg tta tct tcg tca aca aat ttg tgg gta tca gga atc gcg gac ttt 2035
Ser Leu Ser Ser Ser Thr Asn Leu Trp Val Ser Gly Ile Ala Asp Phe
630 635 640 645
ttg cat gaa gat cag aaa gga aac caa cgt agt tat cgt cat tct agc 2083
Leu His Glu Asp Gln Lys Gly Asn Gln Arg Ser Tyr Arg His Ser Ser
650 655 660
gcg ggt tat gca tta gga gga gga ttc ttc acg get tct gaa aat ttc 2131
Ala Gly Tyr Ala Leu Gly Gly Gly Phe Phe Thr Ala Ser Glu Asn Phe
665 670 675
48
CA 02341637 2001-02-19
WO 00/111$3 PCT/IB99/01449
ttt aat ttt get ttt tgt cag ctt ttt ggc tac gac aag gac cat ctt 2179
Phe Asn Phe Ala Phe Cys Gln Leu Phe Gly Tyr Asp Lys Asp His Leu
680 685 690
gtg get aag aac cat acc cat gta tat gca ggg gca atg agt tac cga 2227
Val Ala Lys Asn His Thr His Val Tyr Ala Gly Ala Met Ser Tyr Arg
695 700 705
cac ctc gga gag tct aag acc ctc get aag att ttg tca gga aat tct 2275
His Leu Gly Glu Ser Lys Thr Leu Ala Lys Ile Leu Ser Gly Asn Ser
710 715 720 725
gac tcc cta cct ttt gtc ttc aat get cgg ttt get tat ggc cat acc 2323
Asp Ser Leu Pro Phe Val Phe Asn Ala Arg Phe Ala Tyr Gly His Thr
730 735 740
gac aat aac atg acc aca aag tac act ggc tat tct cct gtt aag gga 2371
Asp Asn Asn Met Thr Thr Lys Tyr Thr Gly Tyr Ser Pro Val Lys Gly
745 750 755
agc tgg gga aat gat gcc ttc ggt ata gaa tgt gga gga get atc ccg 2419
Ser Trp Gly Asn Asp Ala Phe Gly Ile Glu Cys Gly Gly Ala Ile Pro
760 765 770
gta gtt get tca gga cgt cgg tct tgg gtg gat acc cac acg cca ttt 2467
Val Val Ala Ser Gly Arg Arg Ser Trp Val Asp Thr His Thr Pro Phe
775 780 785
cta aac cta gag atg atc tat gca cat cag aat gac ttt aag gaa aac 2515
Leu Asn Leu Glu Met Ile Tyr Ala His Gln Asn Asp Phe Lys Glu Asn
790 795 B00 805
ggc aca gaa ggc cgt tct ttc caa agt gaa gac ctc ttc aat cta gcg 2563
Gly Thr Glu Gly Arg Ser Phe Gln Ser Glu Asp Leu Phe Asn Leu Ala
810 815 820
gtt cct gta ggg ata aaa ttt gag aaa ttc tcc gat aag tct acg tat 2611
Val Pro Val Gly Ile Lys Phe Glu Lys Phe Ser Asp Lys Ser Thr Tyr
825 830 835
gat ctc tcc ata get tac gtt ccc gat gtg att cgt aat gat cca ggc 2659
Asp Leu Ser Ile Ala Tyr Val Pro Asp Val Ile Arg Asn Asp Pro Gly
840 845 850
tgc acg aca act ctt atg gtt tct ggg gat tct tgg tcg aca tgt ggt 2707
Cys Thr Thr Thr Leu Met Val Ser Gly Asp Ser Trp Ser Thr Cys Gly
855 860 865
49
CA 02341637 2001-02-19
WO 00/11183 PCT/IB99/01449
aca agc ttg tct aga caa get ctt ctt gta cgt get gga aat cat cat 2755
Thr Ser Leu Ser Arg Gln Ala Leu Leu Val Arg Ala Gly Asn His His
870 875 880 885
gcc ttt get tca aac ttt gaa gtt ttc agt cag ttt gaa gtc gag ttg 2803
Ala Phe Ala Ser Asn Phe Glu Val Phe Ser Gln Phe Glu Val Glu Leu
890 895 900
cga ggt tct tct cgt agc tat get atc gat ctt gga gga aga ttc gga 2851
Arg Gly Ser Ser Arg Ser Tyr Ala Ile Asp Leu Gly Gly Arg Phe Gly
905 910 915
ttt taatcctaag ttttccaacg agatagcatc agggtaagcc agggctctat 2904
Phe
gtaagagatt tcatagagcc ctctctttgt cttgcttttt gtattttatt ttt 2957
<210> 16
<211> 918
<212> PRT
<213> Chlamydia sp.
<400> 16
Met Arg Ser Ser Phe Ser Leu Leu Leu Ile Ser Ser Ser Leu Ala Phe
1 5 10 15
Pro Leu Leu Met Ser Val Ser Ala Asp Ala Ala Asp Leu Thr Leu Gly
20 25 30
Ser Arg Asp Ser Tyr Asn Gly Asp Thr Ser Thr Thr Glu Phe Thr Pro
35 40 45
Lys Ala Ala Thr Ser Asp Ala Ser Gly Thr Thr Tyr Ile Leu Asp Gly
50 55 60
Asp Val Ser Ile Ser Gln Ala Gly Lys Gln Thr Ser Leu Thr Thr Ser
65 70 75 80
Cys Phe Ser Asn Thr Ala Gly Asn Leu Thr Phe Leu Gly Asn Gly Phe
85 90 95
Ser Leu His Phe Asp Asn Ile Ile Ser Ser Thr Val Ala Gly Val Val
100 105 110
Val Ser Asn Thr Ala Ala Ser Gly Ile Thr Lys Phe Ser Gly Phe Ser
115 120 125
50
CA 02341637 2001-02-19
WO 00/11183 PCT/IB99/01449
Thr Leu Arg Met Leu Ala Ala Pro Arg Thr Thr Gly Lys Gly Ala Ile
130 135 140
Lys Ile Thr Asp Gly Leu Val Phe Glu Ser Ile Gly Asn Leu Asp Leu
145 150 155 160
Asn Glu Asn Ala Ser Ser Glu Asn Gly Gly Ala Ile Asn Thr Lys Thr
165 170 175
Leu Ser Leu Thr Gly Ser Thr Arg Phe Val Ala Phe Leu Gly Asn Ser
180 185 190
Ser Ser Gln Gln Gly Gly Ala Ile Tyr Ala Ser Gly Asp Ser Val Ile
195 200 205
Ser Glu Asn Ala Gly Ile Leu Ser Phe Gly Asn Asn Ser Ala Thr Thr
210 215 220
Ser Gly Gly Ala Ile Ser Ala Glu Gly Asn Leu Val Ile Ser Asn Asn
225 230 235 240
Gln Asn Ile Phe Phe Asp Gly Cys Lys Ala Thr Thr Asn Gly Gly Ala
245 250 255
Ile Asp Cys Asn Lys Ala Gly Ala Asn Pro Asp Pro Ile Leu Thr Leu
260 265 270
Ser Gly Asn Glu Ser Leu His Phe Leu Asn Asn Thr Ala Gly Asn Ser
275 280 285
Gly Gly Ala Ile Tyr Thr Lys Lys Leu Val Leu Ser Ser Gly Arg Gly
290 295 300
Gly Val Leu Phe Ser Asn Asn Lys Ala Ala Asn Ala Thr Pro Lys Gly
305 310 315 320
Gly Ala Ile Ala Ile Leu Asp Ser Gly Glu Ile Ser Ile Ser Ala Asp
325 330 335
Leu Gly Asn Ile Ile Phe Glu Gly Asn Thr Thr Ser Thr Thr Gly Ser
340 345 350
Pro Ala Ser Val Thr Arg Asn Ala Ile Asp Leu Ala Ser Asn Ala Lys
355 360 365
Phe Leu Asn Leu Arg Ala Thr Arg Gly Asn Lys Val Ile Phe Tyr Asp
370 375 380
S1
CA 02341637 2001-02-19
WO 00/11183 PCT/IB99/01449
Pro Ile Thr Ser Ser Gly Ala Thr Asp Lys Leu Ser Leu Asn Lys Ala
385 390 395 400
Asp Ala Gly Ser Gly Asn Thr Tyr Glu Gly Tyr Ile Val Phe Ser Gly
405 410 415
Glu Lys Leu Ser Glu Glu Glu Leu Lys Lys Pro Asp Asn Leu Lys Ser
420 425 430
Thr Phe Thr Gln Ala Val Glu Leu Ala Ala Gly Ala Leu Val Leu Lys
435 440 445
Asp Gly Val Thr Val Val Ala Asn Thr Ile Thr Gln Val Glu Gly Ser
450 955 460
Lys Val Val Met Asp Gly Gly Thr Thr Phe Glu Ala Ser Ala Glu Gly
465 470 475 480
Val Thr Leu Asn Gly Leu Ala Ile Asn Ile Asp Ser Leu Asp Gly Thr
485 490 495
Asn Lys Ala Ile Ile Lys Ala Thr Ala Ala Ser Lys Asp Val Ala Leu
500 505 510
Ser Gly Pro Ile Met Leu Val Asp Ala Gln Gly Asn Tyr Tyr Glu His
515 520 525
His Asn Leu Ser Gln Gln Gln Val Phe Pro Leu Ile Glu Leu Ser Ala
530 535 540
Gln Gly Thr Met Thr Thr Thr Asp Ile Pro Asp Thr Pro Ile Leu Asn
545 550 555 560
Thr Thr Asn His Tyr Gly Tyr Gln Gly Asn Trp Asn Ile Val Trp Val
565 570 575
Asp Asp Ala Thr Ala Lys Thr Lys Asn Ala Thr Leu Thr Trp Thr Lys
580 585 590
Thr Gly Tyr Lys Pro Asn Pro Glu Arg Gln Gly Pro Leu Val Pro Asn
595 600 605
Ser Leu Trp Gly Ser Phe Val Asp Val Arg Ser Ile Gln Ser Leu Met
610 615 620
Asp Arg Ser Thr Ser Ser Leu Ser Ser Ser Thr Asn Leu Trp Val Ser
625 630 635 640
52
CA 02341637 2001-02-19
WO 00/11183 PCT/IB99/01449
Gly Ile Ala Asp Phe Leu His Glu Asp Gln Lys Gly Asn Gln Arg Ser
645 650 655
Tyr Arg His Ser Ser Ala Gly Tyr Ala Leu Gly Gly Gly Phe Phe Thr
66D 665 670
Ala Ser Glu Asn Phe Phe Asn Phe Ala Phe Cys Gln Leu Phe Gly Tyr
675 680 685
Asp Lys Asp His Leu Val Ala Lys Asn His Thr His Val Tyr Ala Gly
690 695 700
Ala Met Ser Tyr Arg His Leu Gly Glu Ser Lys Thr Leu Ala Lys Ile
705 7I0 715 720
Leu Ser Gly Asn Ser Asp Ser Leu Pro Phe Val Phe Asn Ala Arg Phe
725 730 735
Ala Tyr Gly His Thr Asp Asn Asn Met Thr Thr Lys Tyr Thr Gly Tyr
740 745 750
Ser Pro Val Lys Gly Ser Trp Gly Asn Asp Ala Phe Gly Ile Glu Cys
755 760 765
Gly Gly Ala Ile Pro Val Val Ala Ser Gly Arg Arg Ser Trp Val Asp
770 775 780
Thr His Thr Pro Phe Leu Asn Leu Glu Met Ile Tyr Ala His Gln Asn
785 790 795 800
Asp Phe Lys Glu Asn Gly Thr Glu Gly Arg Ser Phe Gln Ser Glu Asp
805 810 815
Leu Phe Asn Leu Ala Val Pro Val Gly Ile Lys Phe Glu Lys Phe Ser
820 825 830
Asp Lys Ser Thr Tyr Asp Leu Ser Ile Ala Tyr Val Pro Asp Val Ile
835 840 845
Arg Asn Asp Pro Gly Cys Thr Thr Thr Leu Met Val Ser Gly Asp Ser
850 855 860
Trp Ser Thr Cys Gly Thr Ser Leu Ser Arg Gln Ala Leu Leu Val Arg
865 870 875 880
Ala Gly Asn His His Ala Phe Ala Ser Asn Phe Glu Val Phe Ser Gln
885 890 895
53
CA 02341637 2001-02-19
WO 00/11183 PCT/IB99/01449
Phe Glu Val Glu Leu Arg Gly Ser Ser Arg Ser Tyr Ala Ile Asp Leu
900 905 910
Gly Gly Arg Phe Gly Phe
915
54
