Efficient mapping of protein antigenic determinants.
A recombinant DNA expression strategy has been used to deduce the amino acid sequences of six different antigenic determinants in a single protein of Mycobacterium leprae, the etiologic agent of leprosy. The gene encoding the M. leprae 65-kDa antigen was sequenced and a lambda gt11 gene sublibrary was constructed with fragments of the gene. Recombinant DNA clones producing specific antigenic determinants were isolated by screening with monoclonal antibodies, and the sequences of their insert DNAs were determined with a rapid primer-extension method. The amino acid sequence of each determinant was deduced from the minimum overlap of insert DNAs from multiple antibody-positive DNA clones. Amino acid sequences for six different epitopes were elucidated. A peptide containing sequences for one of these epitopes was synthesized and shown to bind the appropriate monoclonal antibody; this antigenic determinant is unique to M. leprae. The approach described here can be used to rapidly elucidate protein epitopes that are recognized by antibodies or T cells.
D000595:Amino Acid Sequence; D000911:Antibodies, Monoclonal / Q000276:immunology; D000942:Antigens, Bacterial / Q000032:analysis* / Q000235:genetics; D001483:Base Sequence; D004269:DNA, Bacterial / Q000032:analysis; D000939:Epitopes / Q000032:analysis*; D006801:Humans; D007918:Leprosy / Q000276:immunology; D009166:Mycobacterium leprae / Q000276:immunology*
D000911:Antibodies, Monoclonal; D000942:Antigens, Bacterial; D004269:DNA, Bacterial; D000939:Epitopes