Skip to main content
NCBI home page
Infection and Immunity logoLink to Infection and Immunity
. 1994 Mar;62(3):880–886. doi: 10.1128/iai.62.3.880-886.1994

Isolation and characterization of a gene encoding a Chlamydia pneumoniae 76-kilodalton protein containing a species-specific epitope.

M Perez Melgosa 1, C C Kuo 1, L A Campbell 1
PMCID: PMC186197  PMID: 7509320

Abstract

Chlamydia pneumoniae is a human respiratory pathogen. Unlike the other two Chlamydia species, no species-specific antigen has been defined for C. pneumoniae. An immunoreactive clone containing a 0.8-kb fragment was isolated from a C. pneumoniae (AR-39) genomic library by using anti-C. pneumoniae rabbit immune serum. By Southern hybridization analysis of chromosomal digests of the different Chlamydia spp., the 0.8-kb fragment was shown to react specifically with C. pneumoniae. Subcloning of this fragment into the pGEX-1 lambda T expression vector resulted in the expression of a 62-kDa fusion protein. This fusion protein as well as the cleaved C. pneumoniae peptide were recognized by anti-C. pneumoniae rabbit immune serum, while the glutathione S-transferase moiety was not recognized. The fusion protein was used to produce monospecific rabbit antiserum. This antiserum was shown to react with a 76-kDa protein in all C. pneumoniae isolates tested, specifically recognize C. pneumoniae inclusions in tissue culture, and neutralize infectivity of C. pneumoniae in cell culture. No reactivity was observed with Chlamydia trachomatis or Chlamydia psittaci. To isolate the entire coding sequence of the 76-kDa protein, two partially overlapping fragments of C. pneumoniae DNA, a 3.2-kb HindIII fragment and a 1.2-kb PvuII fragment, were isolated, cloned, and sequenced. No significant sequence similarity was found with any previously reported nucleotide or amino acid sequence of the other Chlamydia species. This C. pneumoniae protein containing a species-specific epitope could play a role in pathogenesis and may be useful as a diagnostic tool.

Full text

PDF
880

Images in this article

882Image
on p.882
882Image
on p.882
882Image
on p.882
883Image
on p.883

Selected References

These references are in PubMed. This may not be the complete list of references from this article.

  1. Alexander A. G., Kenny G. E. Characterization of membrane and cytoplasmic antigens of Mycoplasma arginini by two-dimensional (crossed) immunoelectrophoresis. Infect Immun. 1977 Jan;15(1):313–321. doi: 10.1128/iai.15.1.313-321.1977. [DOI] [PMC free article] [PubMed] [Google Scholar]
  2. Campbell J. F., Barnes R. C., Kozarsky P. E., Spika J. S. Culture-confirmed pneumonia due to Chlamydia pneumoniae. J Infect Dis. 1991 Aug;164(2):411–413. doi: 10.1093/infdis/164.2.411. [DOI] [PubMed] [Google Scholar]
  3. Campbell L. A., Kuo C. C., Grayston J. T. Characterization of the new Chlamydia agent, TWAR, as a unique organism by restriction endonuclease analysis and DNA-DNA hybridization. J Clin Microbiol. 1987 Oct;25(10):1911–1916. doi: 10.1128/jcm.25.10.1911-1916.1987. [DOI] [PMC free article] [PubMed] [Google Scholar]
  4. Campbell L. A., Kuo C. C., Wang S. P., Grayston J. T. Serological response to Chlamydia pneumoniae infection. J Clin Microbiol. 1990 Jun;28(6):1261–1264. doi: 10.1128/jcm.28.6.1261-1264.1990. [DOI] [PMC free article] [PubMed] [Google Scholar]
  5. Campbell L. A., Perez Melgosa M., Hamilton D. J., Kuo C. C., Grayston J. T. Detection of Chlamydia pneumoniae by polymerase chain reaction. J Clin Microbiol. 1992 Feb;30(2):434–439. doi: 10.1128/jcm.30.2.434-439.1992. [DOI] [PMC free article] [PubMed] [Google Scholar]
  6. Ekman M. R., Grayston J. T., Visakorpi R., Kleemola M., Kuo C. C., Saikku P. An epidemic of infections due to Chlamydia pneumoniae in military conscripts. Clin Infect Dis. 1993 Sep;17(3):420–425. doi: 10.1093/clinids/17.3.420. [DOI] [PubMed] [Google Scholar]
  7. GORDON F. B., QUAN A. L. OCCURENCE OF GLYCOGEN IN INCLUSIONS OF THE PSITTACOSIS-LYMPHOGRANULOMA VENEREUM-TRACHOMA AGENTS. J Infect Dis. 1965 Apr;115:186–196. doi: 10.1093/infdis/115.2.186. [DOI] [PubMed] [Google Scholar]
  8. Grayston J. T., Campbell L. A., Kuo C. C., Mordhorst C. H., Saikku P., Thom D. H., Wang S. P. A new respiratory tract pathogen: Chlamydia pneumoniae strain TWAR. J Infect Dis. 1990 Apr;161(4):618–625. doi: 10.1093/infdis/161.4.618. [DOI] [PubMed] [Google Scholar]
  9. Hahn D. L., Dodge R. W., Golubjatnikov R. Association of Chlamydia pneumoniae (strain TWAR) infection with wheezing, asthmatic bronchitis, and adult-onset asthma. JAMA. 1991 Jul 10;266(2):225–230. [PubMed] [Google Scholar]
  10. Hammerschlag M. R., Hyman C. L., Roblin P. M. In vitro activities of five quinolones against Chlamydia pneumoniae. Antimicrob Agents Chemother. 1992 Mar;36(3):682–683. doi: 10.1128/aac.36.3.682. [DOI] [PMC free article] [PubMed] [Google Scholar]
  11. Howard L., Orenstein N. S., King N. W. Purification on renografin density gradients of Chlamydia trachomatis grown in the yolk sac of eggs. Appl Microbiol. 1974 Jan;27(1):102–106. doi: 10.1128/am.27.1.102-106.1974. [DOI] [PMC free article] [PubMed] [Google Scholar]
  12. Kikuta L. C., Puolakkainen M., Kuo C. C., Campbell L. A. Isolation and sequence analysis of the Chlamydia pneumoniae GroE operon. Infect Immun. 1991 Dec;59(12):4665–4669. doi: 10.1128/iai.59.12.4665-4669.1991. [DOI] [PMC free article] [PubMed] [Google Scholar]
  13. Kornak J. M., Kuo C. C., Campbell L. A. Sequence analysis of the gene encoding the Chlamydia pneumoniae DnaK protein homolog. Infect Immun. 1991 Feb;59(2):721–725. doi: 10.1128/iai.59.2.721-725.1991. [DOI] [PMC free article] [PubMed] [Google Scholar]
  14. Kuo C. C., Chen H. H., Wang S. P., Grayston J. T. Identification of a new group of Chlamydia psittaci strains called TWAR. J Clin Microbiol. 1986 Dec;24(6):1034–1037. doi: 10.1128/jcm.24.6.1034-1037.1986. [DOI] [PMC free article] [PubMed] [Google Scholar]
  15. Kuo C. C., Shor A., Campbell L. A., Fukushi H., Patton D. L., Grayston J. T. Demonstration of Chlamydia pneumoniae in atherosclerotic lesions of coronary arteries. J Infect Dis. 1993 Apr;167(4):841–849. doi: 10.1093/infdis/167.4.841. [DOI] [PubMed] [Google Scholar]
  16. Ladany S., Black C. M., Farshy C. E., Ossewaarde J. M., Barnes R. C. Enzyme immunoassay to determine exposure to Chlamydia pneumoniae (strain TWAR). J Clin Microbiol. 1989 Dec;27(12):2778–2783. doi: 10.1128/jcm.27.12.2778-2783.1989. [DOI] [PMC free article] [PubMed] [Google Scholar]
  17. Laemmli U. K. Cleavage of structural proteins during the assembly of the head of bacteriophage T4. Nature. 1970 Aug 15;227(5259):680–685. doi: 10.1038/227680a0. [DOI] [PubMed] [Google Scholar]
  18. Lucero M. E., Kuo C. C. Neutralization of Chlamydia trachomatis cell culture infection by serovar-specific monoclonal antibodies. Infect Immun. 1985 Nov;50(2):595–597. doi: 10.1128/iai.50.2.595-597.1985. [DOI] [PMC free article] [PubMed] [Google Scholar]
  19. Perez Melgosa M., Kuo C. C., Campbell L. A. Sequence analysis of the major outer membrane protein gene of Chlamydia pneumoniae. Infect Immun. 1991 Jun;59(6):2195–2199. doi: 10.1128/iai.59.6.2195-2199.1991. [DOI] [PMC free article] [PubMed] [Google Scholar]
  20. Puolakkainen M., Kuo C. C., Shor A., Wang S. P., Grayston J. T., Campbell L. A. Serological response to Chlamydia pneumoniae in adults with coronary arterial fatty streaks and fibrolipid plaques. J Clin Microbiol. 1993 Aug;31(8):2212–2214. doi: 10.1128/jcm.31.8.2212-2214.1993. [DOI] [PMC free article] [PubMed] [Google Scholar]
  21. Saikku P., Leinonen M., Mattila K., Ekman M. R., Nieminen M. S., Mäkelä P. H., Huttunen J. K., Valtonen V. Serological evidence of an association of a novel Chlamydia, TWAR, with chronic coronary heart disease and acute myocardial infarction. Lancet. 1988 Oct 29;2(8618):983–986. doi: 10.1016/s0140-6736(88)90741-6. [DOI] [PubMed] [Google Scholar]
  22. Sanger F., Nicklen S., Coulson A. R. DNA sequencing with chain-terminating inhibitors. Proc Natl Acad Sci U S A. 1977 Dec;74(12):5463–5467. doi: 10.1073/pnas.74.12.5463. [DOI] [PMC free article] [PubMed] [Google Scholar]
  23. Smith P. K., Krohn R. I., Hermanson G. T., Mallia A. K., Gartner F. H., Provenzano M. D., Fujimoto E. K., Goeke N. M., Olson B. J., Klenk D. C. Measurement of protein using bicinchoninic acid. Anal Biochem. 1985 Oct;150(1):76–85. doi: 10.1016/0003-2697(85)90442-7. [DOI] [PubMed] [Google Scholar]
  24. Stephens R. S., Wagar E. A., Schoolnik G. K. High-resolution mapping of serovar-specific and common antigenic determinants of the major outer membrane protein of Chlamydia trachomatis. J Exp Med. 1988 Mar 1;167(3):817–831. doi: 10.1084/jem.167.3.817. [DOI] [PMC free article] [PubMed] [Google Scholar]
  25. Thom D. H., Grayston J. T., Siscovick D. S., Wang S. P., Weiss N. S., Daling J. R. Association of prior infection with Chlamydia pneumoniae and angiographically demonstrated coronary artery disease. JAMA. 1992 Jul 1;268(1):68–72. [PubMed] [Google Scholar]
  26. Towbin H., Staehelin T., Gordon J. Electrophoretic transfer of proteins from polyacrylamide gels to nitrocellulose sheets: procedure and some applications. Proc Natl Acad Sci U S A. 1979 Sep;76(9):4350–4354. doi: 10.1073/pnas.76.9.4350. [DOI] [PMC free article] [PubMed] [Google Scholar]
  27. Yuan Y., Zhang Y. X., Watkins N. G., Caldwell H. D. Nucleotide and deduced amino acid sequences for the four variable domains of the major outer membrane proteins of the 15 Chlamydia trachomatis serovars. Infect Immun. 1989 Apr;57(4):1040–1049. doi: 10.1128/iai.57.4.1040-1049.1989. [DOI] [PMC free article] [PubMed] [Google Scholar]

Articles from Infection and Immunity are provided here courtesy of American Society for Microbiology (ASM)

RESOURCES