ACID PROTEASE STRUCTURE, DYNAMICS AND MECHANISM

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The aim of this research is to understand in detail how inhibitors (and substrates) interact with the active sites of acid proteases, including pepsin, cathespin D and renin. The work has importance for a full understanding of the mechanism of action of these enzymes, and for design of specific inhibitors of renin for control of hypertension. The method of study involves mainly 1H and 13C NMR studies of pepstatin and pepstatin analogs binding to porcine pepsin. Specifically deuterated analogs will be used, as well as 13C labeled pepstatin derivatives. The objective is to provide a detailed molecular description of the interaction of strong inhibitors with amino acid groups in the active site of acid proteases. Much of the research will be directed to take advantage of the discovery that ketone analogs of pepstatin, isosteric with substrates, and enriched with 13C in the carbonyl, provide direct 13C NMR evidence for formation of tetrahedral intermediates upon binding to pepsin. Studies will also be done by 1H NMR on the solution conformation of oligosaccharides found on some aspartyl protease enzymes.