Antigens secreted by live Mycobacterium tuberculosis represent attractive candidates for inclusion in novel vaccine preparations. While promising results have been reported using """"""""native"""""""" proteins isolated from M. tuberculosis cultures, recombinant DNA approaches for generation of bulk quantities of these antigens have so far proved disappointing. We propose that problems in expressing secreted antigens in recombinant systems are related to the requirement for post-translational modification of the proteins, and furthermore speculate that modification events may have important effects on immunogenicity. To test this hypothesis, and to identify procedures for production of secreted antigens in a form suitable for vaccine preparation, we propose to express genes encoding the 38kD and l9kD secreted antigens of M. tuberculosis in a series of mycobacterial expression systems (BCG, M. smegmatis, M. vaccae) . Recombinant proteins will be purified and compared with the corresponding native proteins from M. tuberculosis by detailed chemical and immunological analyses. We will determine (a) the precise nature of post-translational modifications affecting the native antigens, (b) whether or not species-specific variations occur in post-translational modification pathways, and (c) whether or not post-translational modification affects the immunogenicity of these proteins. In addition to identifying optimal procedures for production of recombinant mycobacterial antigens, the results of this study will provide important new insights into the basic biology and immunogenicity of M. tuberculosis.
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