MOLECULAR-CLONING, EXPRESSION, AND IMMUNOGENICITY OF MTB12, A NOVEL LOW-MOLECULAR-WEIGHT ANTIGEN SECRETED BY MYCOBACTERIUM-TUBERCULOSIS

Proteins secreted into the culture medium by Mycobacterium tuberculosi s are thought to play an important role in the development of protecti ve immune responses. In this report,we describe the molecular cloning off a novel, low-molecular-weight antigen (MTB12) secreted by M. tuber culosis. Sequence analysis of the MTB12 gene indicates that the protei n is initially synthesized as a 16.6-kDA precursor protein containing a 48-amino-acid hydrophobic leader sequence. The mature, fully process ed form of MTB12 protein found in culture filtrates has a molecular ma ss of 12.5 kDa. MTB12 protein constitutes a major component of time M. tuberculosis culture supernatant and appears to be mt least as abunda nt as several other well-characterized culture filtrate proteins, incl uding members of the 85B complex. MTB12 is encoded by a single-copy ge ne which is present in both virulent and avirulent strains of the M. t uberculosis complex, the BCG strain of M. bovis, and M. leprae. Recomb inant MTB12 containing an N-terminal six-histidine tag was expressed i n Escherichia coli and purified by affinity chromatography. Recombinan t MTB12 protein elicited in vitro proliferative responses fram the per ipheral blood mononuclear cells of a number of purified protein deriva tive-positive (PPD+) human donors hut not from PPD- donors.