Murine and human B cell epitope mapping of the Mycobacterium tuberculosis 10-kD heat shock protein using overlapping peptides.
The human immune response to the 10-kD M. tuberculosis protein was studied by a competition ELISA using monoclonal antibody (MoAb) SA-12. Twenty-five per cent of the sera from 20 patients with tuberculosis and none from 21 control subjects inhibited binding of SA-12 to the 10-kD antigen. To characterize the antigenic parts of the 10-kD antigen, overlapping decapeptides according to the amino acid sequence of the 10-kD protein were synthesized. In total, 91 sequential decapeptides, with an overlap of nine amino acids, were tested in ELISA with MoAb SA-12, human and murine sera (PEP scan). SA-12 recognized the amino acid sequence WDEDGEK (amino acid 50-56). All human sera, from patients with tuberculosis as well as from control subjects, gave almost identical undulating patterns of reactivity with the decapeptides. No relationship was found between the ability of the patients' sera to inhibit binding of MoAb SA-12 and the binding of these sera to the decapeptides comprising the epitope recognized by SA-12 in the PEP scan. Apparently, antibodies in patients' sera against the 10-kD protein are predominantly directed against discontinuous epitopes and, consequently, the continuous epitopes as presented in the PEP scan are not suitable to discriminate between patients with tuberculosis and control subjects. In the PEP scan, sera from BALB/c mice, both non-immunized and immunized with either live M. tuberculosis or the 10-kD protein gave similar patterns of reactivity, albeit different from the patterns obtained with the human sera. However, after immunization of the mice, clearly increased levels of antibodies to primary structures of the 10-kD protein were observed.
Verbon
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Laboratory of Tropical Hygiene, Royal Tropical Institute, Amsterdam, The Netherlands.
Hartskeerl
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RA
Kolk
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AH
D000595:Amino Acid Sequence; D000818:Animals; D000911:Antibodies, Monoclonal / Q000276:immunology; D000942:Antigens, Bacterial / Q000737:chemistry / Q000276:immunology*; D001402:B-Lymphocytes / Q000276:immunology*; D001426:Bacterial Proteins / Q000737:chemistry / Q000276:immunology; D000939:Epitopes; D006360:Heat-Shock Proteins / Q000737:chemistry / Q000276:immunology*; D006801:Humans; D051379:Mice; D008969:Molecular Sequence Data; D008970:Molecular Weight; D009169:Mycobacterium tuberculosis / Q000276:immunology*; D010455:Peptides / Q000737:chemistry / Q000276:immunology; D012995:Solubility; D014397:Tuberculosis, Pulmonary / Q000276:immunology
D000911:Antibodies, Monoclonal; D000942:Antigens, Bacterial; D001426:Bacterial Proteins; D000939:Epitopes; D006360:Heat-Shock Proteins; D010455:Peptides