Cloning and characterization of a Chlamydia trachomatis L3 DNA fragment that codes for an antigenic region of the major outer membrane protein and specifically hybridizes to the C- and C-related-complex serovars

Carlson, Elaine J.; Peterson, Ellena M.; Maza, Luis M. de la

doi:10.1128/iai.57.2.487-494.1989

Cited by 11 publications

(7 citation statements)

References 33 publications

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“…Immunoassays. Western blots (immunoblots) were performed as described previously (5), with the exception that a tricine-sodium dodecyl sulfate-polyacrylamide gel electrophoresis system was used (20). The dot blot assay utilizing untreated and heat-treated (56°C, 30 min) elementary bodies was performed as previously described by Zhang et al (30).…”

Section: Methodsmentioning

confidence: 99%

Characterization of a neutralizing monoclonal antibody directed at variable domain I of the major outer membrane protein of Chlamydia trachomatis C-complex serovars

Cheng

Maza

et al. 1993

Infect Immun

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A monoclonal antibody (MAb), C10, that neutralized in vitro the infectivity of serovars C, I, J, and L3 (members of the C and C-related complexes) of Chlamydia trachomatis was identified. Of the 15 major serovars and the mouse pneumonitis strain of C. trachomatis, Chlamydia psittaci, and Chlamydia pneumoniae, which were used as nontreated and heat-treated (56°C, 30 min) antigens in a dot blot assay, only serovars C, I, J, and L3 were recognized with both the native and treated antigens. Western blot (immunoblot) results showed that MAb C10 recognized the major outer membrane protein of these four serovars. Overlapping hexameric peptides corresponding to variable domains (VDs) I, II, III, and IV of the major outer membrane protein of C. trachomatis serovar C were synthesized, and peptide screening showed that MAb C10 mapped to the VD I amino acid sequence VAGLQNDPT. Results of an in vitro neutralization assay correlated with those of the indirect immunofluorescence assay, Western blot, and dot blot assay in that only serovars C, I, J, and L3 were neutralized by MAb C10. In vitro competitive neutralization experiments, using a peptide representing VD I of serovar C to compete with C. trachomatis serovar C for MAb C10 binding, revealed that both serological and neutralizing activities of MAb C10 were inhibited by the VD I peptide. In an in vivo toxicity/infectivity assay using serovar L3 pretreated with MAb C10, there was 100o survival of mice infected with a lethal dose at 48 h. In contrast, the control group, consisting of mice injected with the same dose of 13 pretreated with a MAb that does not recognize L3, had no survivors during a 48-h observation period. In summary, since the surface-exposed contiguous epitope recognized by MAb C10 binds neutralizing antibodies that are subspecies specific for the C and C-related complexes, it should be considered for inclusion in the development of a chlamydial vaccine.

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Section: Methodsmentioning

confidence: 99%

Characterization of a neutralizing monoclonal antibody directed at variable domain I of the major outer membrane protein of Chlamydia trachomatis C-complex serovars

Cheng

Maza

et al. 1993

Infect Immun

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“…A C. trachomatis serovar E DNA library digested with EcoRI (New England BioLabs, Beverly, Mass.) was constructed in Xgtll and screened as previously described (6). An MAb E4-reactive clone was purified, and DNA from this clone was digested with EcoRI and separated on a 1% low-melting-point agarose gel.…”

Section: Methodsmentioning

confidence: 99%

“…The DNA coding for the MOMP of C. trachomatis has been sequenced, and the antigenic determinants of the protein have been mapped with several monoclonal antibodies (MAbs) (1,6,8,9,31). This protein is composed of four variable domains (VDs), which are interspersed between conserved regions (30).…”

mentioning

confidence: 99%

“…This protein is composed of four variable domains (VDs), which are interspersed between conserved regions (30). The epitopes for MAbs that are serovar, subspecies, and species specific have been mapped to these VDs (1,6,8,9,31). In general, it is believed that antibodies directed to serovar-and subspecies-specific epitopes are protective, while those recognizing speciesspecific epitopes are not (15,39).…”

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confidence: 99%

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Functional and structural mapping of Chlamydia trachomatis species-specific major outer membrane protein epitopes by use of neutralizing monoclonal antibodies

et al. 1991

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Three monoclonal antibodies (MAbs), E4, Ll-4, and L1-24, to the major outer membrane protein (MOMP) of Chiamydia trachomatis were identified that neutralized in vitro the infectivity of members of the Band C-related complex as well as the mouse pneumonitis strain. MAbs L1-4, Ll-24, and E4 gave a strong signal in an indirect immunofluorescence assay and/or Western immunoblot with all serovars of the lymphogranuloma venereum and trachoma biovars and a weak signal with the mouse biovar. In addition, C. psittaci and C. pneumoniae were also weakly recognized by MAbs Ll-4 and Ll-24. As determined by the technique of overlapping peptides, all three MAbs showed reactivity to variable domain (VD) IV of MOMP. While all three MAbs had different recognition patterns, all strongly bound to the peptides TLNPTI and LNPTIA within the species-conserved region of VD IV. MAb E4 also recognized the peptide SATAIF in the subspecies region of VD IV. Peptides corresponding to VY IV of MOMP were synthesized and used in competitive inhibition experiments to determine the functional location of the epitope recognized by these three MAbs. Both the serological and neutralizing activities of MAb E4 were inhibited by the peptides ATAIFDTTTLNPTIAG and FDTTTLNPTIAG; however, none of the peptides made to the VD IV region blocked the neutralizing activity of MAbs L1-4 and Ll-24. Therefore, the neutralizable domain of the epitope recognized by MAb E4 is contiguous and may be an important candidate for inclusion in a subunit vaccine.

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“…The 40-kDa major outer membrane protein accounts for approximately 60% of the weight of the outer membrane and is acidic, with a pl ranging from 5.3 to 5.5 (4). This protein is a strong immunogen for humans and other mammals, and its molecular and antigenic structure has been extensively characterized (3,7,22). The 60-kDa protein appears as a doublet in the lymphogranuloma venereum (LGV) biovar, whereas only a single band has been detected in the trachoma biovar (4).…”

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Sequence diversity of the 60-kilodalton protein and of a putative 15-kilodalton protein between the trachoma and lymphogranuloma venereum biovars of Chlamydia trachomatis

et al. 1991

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DNA from Chiamydia trachomatis serovars L3, C, and E corresponding to the open reading frames of the 60-kDa protein and of a putative 15-kDa protein was sequenced. The open reading frames coding for the 60-kDa protein had 1,641 bp in the three serovars. Compared with the L3 serovar, there were 9 and 11 amino acid changes in the C and E serovars, respectively. The open reading frames corresponding to the putative 15-kDa protein had 450, 456, and 453 bp for the L3, C, and E serovars, respectively. When compared with the L3 serovar, the C and E serovars had 14 and 16 amino acid differences, respectively.

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Cloning and characterization of a Chlamydia trachomatis L3 DNA fragment that codes for an antigenic region of the major outer membrane protein and specifically hybridizes to the C- and C-related-complex serovars

Cited by 11 publications

References 33 publications

Characterization of a neutralizing monoclonal antibody directed at variable domain I of the major outer membrane protein of Chlamydia trachomatis C-complex serovars

Characterization of a neutralizing monoclonal antibody directed at variable domain I of the major outer membrane protein of Chlamydia trachomatis C-complex serovars

Functional and structural mapping of Chlamydia trachomatis species-specific major outer membrane protein epitopes by use of neutralizing monoclonal antibodies

Sequence diversity of the 60-kilodalton protein and of a putative 15-kilodalton protein between the trachoma and lymphogranuloma venereum biovars of Chlamydia trachomatis

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