Characterization of the 35-kilodalton Treponema pallidum subsp. pallidum recombinant lipoprotein TmpC and antibody response to lipidated and nonlipidated T. pallidum antigens.
The gene encoding the 35-kDa immunogenic Treponema pallidium subsp. pallidum (T. pallidum) membrane protein C, TmpC, was cloned, sequenced, and expressed in Escherichia coli. The deduced amino acid sequence carries on N-terminal signal sequence with a four-amino-acid motif, which is characteristic for bacterial lipoproteins. Metabolic labeling with radioactive palmitic acid of E. coli expressing TmpC revealed incorporation of the fatty acid into the antigen. The antigen was overproduced, purified to near homogeneity and used in an enzyme-linked immunosorbent assay (ELISA) to evaluate its potential for the serodiagnosis of syphilis. Although all sera from untreated secondary syphilis patients were reactive in this TmpC ELISA, only a minority of the serum samples from untreated patients in the primary or early latent stage of the disease contained significant anti-TmpC antibodies. To study the influence of the lipid moiety on the antigenic properties of the TmpC, TmpA, and TpD lipoproteins, plasmids encoding nonlipidated forms of these antigens were constructed. In addition, a plasmid expressing a lipidated form of the otherwise non-lipid-modified antigen TmpB was constructed. Immunization and absorption experiments with these lipidated and nonlipidated antigens showed that antibodies against the lipid moiety of lipoproteins could not be detected on immunoblots, neither in sera from infected rabbits nor in sera from animals immunized with the lipoproteins. In addition, we were unable to demonstrate cross-reactivity between antibodies against the T. pallidum lipoproteins and those reactive to the Venereal Diseases Research Laboratories test, suggesting that antibodies reactive to the Venereal Diseases Research Laboratories test are unrelated to antilipoprotein antibodies.
Schouls
L M
LM
Unit Molecular Microbiology, National Institute of Public Health and Environmental Protection, Bilthoven, The Netherlands.
van der Heide
H G
HG
van Embden
J D
JD
D000595:Amino Acid Sequence; D000907:Antibodies, Bacterial / Q000032:analysis*; D000942:Antigens, Bacterial / Q000276:immunology*; D001483:Base Sequence; D006801:Humans; D008055:Lipids / Q000276:immunology*; D008074:Lipoproteins / Q000096:biosynthesis / Q000235:genetics* / Q000276:immunology; D008565:Membrane Proteins / Q000235:genetics*; D008969:Molecular Sequence Data; D011499:Protein Processing, Post-Translational; D011994:Recombinant Proteins / Q000096:biosynthesis; D013587:Syphilis / Q000276:immunology; D014210:Treponema pallidum / Q000276:immunology*
D000907:Antibodies, Bacterial; D000942:Antigens, Bacterial; D008055:Lipids; D008074:Lipoproteins; D008565:Membrane Proteins; D011994:Recombinant Proteins