The re-emergence of tuberculosis as a public health problem has been complicated by the lack of effective chemotherapeutic agents and the development of drug-resistant strains. The cell wall of the pathogen Mycobacterium tuberculosis, is known to be the target of some of the most effective anti-mycobacterial drugs including ethambutol which is known to inhibit the biosynthesis of the arabinan of cell wall arabinogalactan (AG) and the associated lipoarabinomannan (LAM). A diverse range of biological studies over the past few years has collectively provided compelling evidence implicating LAM as a key surface molecule in host-pathogen interactions. The availability of truncated, mutated LAM variants as a consequence of drug resistance and genetic manipulation provides invaluable model compounds for both structural and functional studies aiming at defining the relevance of LAM in pathogenesis. Specifically, the fine details of the arabinan assembly and its point(s) of attachment to the phosphatidylinositol mannan core will be characterized and structural niceties positively correlating with particular biological attributes of clinical isolates will be identified. Structural basis of microheterogeneity in LAM will be defined and chemically and/or enzymatically modified arabinan and mannan will be derived from LAM for structural/biological studies. As a major spin off, the recent availability of the consequential cell wall mutants due to genetic manipulation of the embCAB proteins, and analyses of the gene products now allows us rationally to dissect the pathway to the formation of the arabinan of LAM/AG. In the same vein, studies on LAM mutants in M. tuberculosis and our concerted efforts on generating LAM depleted M. tuberculosis will contribute directly into addressing the role of LAM in survival/infectivity of the organism. Thus, the unifying theme of this Research Proposal encompasses structural analysis and manipulation of LAM, supplemented by genetic probes to alter its structure and mutate LAM in M. smegmatis and M. tuberculosis all in relation to biology and biosynthesis.
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