A multi-faceted research project is directly aimed at computational studies of enzymatic processes in aqueous solution. The theoretical approach centers on molecular dynamics simulations of enzymatic systems using combined quantum mechanical and molecular mechanical (QM/MM) methods. To achieve greater accuracy, we propose to further develop a mixed molecular orbital-valence bond (MOVB) method for simulation of enzyme reactions and sampling of the reaction pathway. In addition, we plan to implement a semi-empirical density functional theory for combined QM/MM calculations, which will significantly expand the scope of QM/MM applications to enzymatic systems, including metailoenzymes. A major thrust is to provide a deeper understanding of the remarkable catalytic power of enzymes. Our approach is to seek general catalytic principles, by examining individual enzymatic systems that share common features, but have different biological functions. In particular, the hydrolytic cysteine protease, human cathepsin K, and alanine and glutamate racemases, will be investigated in detail to understand substrate binding, reaction mechanism, and free energy profiles. Inhibitors of cathepsin K can reduce bone resorption, providing a promising therapeutic target for the treatment of osteoporosis and rheumatoid arthritis, while amino acid racemases are essential in the synthesis of the peptidoglycan layer of bacteria cell walls, rendering them attractive targets for inhibitors. The proposed computational study will provide insight into the mechanism of acid/base catalysis of these two important classes of enzymes. In addition, the dynamic conformational changes in thymidylate synthase (TS), that take place throughout the many-steps of the enzymatic reaction, will be studied. TS catalyzes the de novo synthesis of dTMP nucleotide for DNA synthesis, which has been extensively investigated experimentally. The proposed study will provide a deeper understanding of the roles of protein dynamic conformation change in the function of thymidylate synthase, and the results will be of general importance in enzyme catalysis. ? ?
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