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Isolation and characterization of a gene encoding a Chlamydia pneumoniae 76-kilodalton protein containing a
M Perez Melgosa1, C C Kuo, L A Campbell
1Department of Pathobiology, University of Washington, Seattle 98195.
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.
Insights
Researchers identified a unique 76-kDa protein in Chlamydia pneumoniae, a respiratory pathogen. This species-specific antigen offers potential for developing new diagnostic tools for C. pneumoniae infections.
Area of Science:
- Microbiology
- Immunology
- Molecular Biology
Background:
- Chlamydia pneumoniae is a significant human respiratory pathogen.
- Unlike other Chlamydia species, a species-specific antigen for C. pneumoniae has not been previously defined.
- The lack of a specific antigen hinders accurate diagnosis and understanding of C. pneumoniae pathogenesis.
Purpose of the Study:
- To identify and characterize a species-specific antigen for Chlamydia pneumoniae.
- To investigate the potential diagnostic and pathogenetic roles of this antigen.
- To isolate and sequence the gene encoding the identified antigen.
Main Methods:
- Screening a C. pneumoniae genomic library with anti-C. pneumoniae serum.
- Southern hybridization to confirm species specificity.
- Expression of a fusion protein and generation of specific antiserum.
- DNA sequencing of relevant gene fragments.
Main Results:
- A 0.8-kb DNA fragment specifically reactive with C. pneumoniae was isolated.
- This fragment expressed a 62-kDa fusion protein recognized by specific antibodies.
- Antiserum against the fusion protein recognized a 76-kDa protein in all C. pneumoniae isolates, neutralized infectivity, and identified C. pneumoniae inclusions.
- The gene sequence showed no similarity to other Chlamydia species.
Conclusions:
- A novel, species-specific 76-kDa protein antigen of Chlamydia pneumoniae has been identified.
- This protein possesses a species-specific epitope and shows potential for diagnostic applications.
- The identified antigen may play a role in C. pneumoniae pathogenesis.