Mortality in AIDS patients is due to overwhelming opportunistic infections caused by bacteria, fungi, protozoa, and viruses. The long term goal of this project is to develop lead compounds useful for the treatment of bacterial opportunistic infections in AIDS by developing strong inhibitors of alanine racemase, an essential bacterial enzyme involved in cell wall biosynthesis. These compounds will be engineered in a collaborative structure based drug design program based at the University of Houston. Three common opportunistic pathogens will be targeted, Mycobacterium tuberculosis, Mycobacterium avium, and Streptococcus pneumoniae, but several other pathogens will be studied as well. To date, overexpressing clones of alanine racemase from E. coli, Mycobacterium tuberculosis, Pseudomonas aeruginosa, Staphylococcus aureus, and Streptococcus pneumoniae have been used to produce purified protein. These enzymes are being characterized kinetically. Crystallization of all of these racemases will be attempted, and alanine racemase from E. coli, Pseudomonas aeruginosa and Mycobacterium tuberculosis have already been crystallized. The crystals from the E. coli enzyme are of sufficient quality to allow for a three-dimensional structure determination. These structures will serve as target molecules for a comprehensive structure based drug design effort. Promising lead compounds will be synthesized and tested in vitro. Their strength and specificity will be improved through structural determinations of inhibitor-enzyme complexes followed by molecular dynamics simulations. Studies of mutant racemases will be used to help anticipate common pathways for developing resistance to these compounds. While the treatment of opportunistic infections in AIDS patients will benefit most from this research, development of highly effective alanine racemase inhibitors may broadly impact the treatment of other infectious diseases, including those caused by multi-drug resistant bacteria.
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