CHARACTERIZATION OF A LIPOPROTEIN IN MYCOBACTERIUM-BOVIS (BCG) WITH SEQUENCE SIMILARITY TO THE SECRETED PROTEIN MPB70

The monoclonal antibody, mAb3C4, raised against sonicated Mycobacteriu m bovis (Mb) BCG (Tokyo strain 172) cells recognises a 23-kDa protein in the cell wall. The gene encoding this protein was cloned and sequen ced and found to be 100% homologous to mpb83 and mpt83 and the putativ e protein to have a 76% sequence similarity to the secreted, Mb-specif ic protein, MPB70. MPB83 contains the amino acid (aa) sequence LAGC, w hich corresponds to the consensus sequence for bacterial lipoprotein m odification and processing. MPB83 associated with the detergent phase when separated with Triton X-114 confirming that it is a lipoprotein. When the putative site of acylation, the Cys in the sequence LAGC, was substituted with Ser, the mutated MPB83 associated with the aqueous p hase. The cloned gene was used to determine the distribution of mpb83 in various Mycobacterium species. The gene was present in the M. tuber culosis (Mt) complex organisms, as well as in M. kansasii. In addition , Southern blot analysis of Mb and Mt DNA indicated that the mpb83 and mpb70 genes are located close to each other on the genome. Western bl ot analysis of cell lysates of various Mycobacterium species indicated that only Mt H37Rv and H37Ra produced proteins which reacted with mAb 3C4. Furthermore, only two out of six of the Mb field isolates produce d detectable antigen, indicating that expression of the mpb83 gene is variable within the Mt complex organisms.