The goal of the proposed research is to develop and apply theoretical simulation techniques to study the nature and energetics of protein- substrate and protein-inhibitor interactions, the mechanisms of uncatalyzed solution phase reactions, and enzymatically catalyzed reactions. Theoretical insights into these phenomenon will have a major impact on human health. The increase in our understanding of existing inhibitors and the design of new enzyme inhibitors will have a major impact on improving the human condition. An increase in our knowledge of protein structure and function will have a major impact on biotechnology, because it will allow for the ab initio design of new proteins with increased or more selective catalytic properties. To address these issues we have decided to focus on one protein and subject to intense theoretical scrutiny. Our model will be the zinc metalloenzyme human carbonic anhydrase II (HCAII), which is of great biomedical importance because of its role in physiological processes involving CO2, and because inhibitors of this enzyme are drugs used in the treatment of glaucoma. Our first goal will be to develop the necessary molecular mechanical potential function parameters for the catalytically necessary zinc ion in this enzyme, followed by the development of a coupled quantum mechanical/molecular mechanical simulation package, which will be used to study both the uncatalyzed and catalyzed reactions of CO2. In order to identify what structural features are important for enzymatic catalysis it is first useful to understand the uncatalyzed gas-phase and solution phase reactions. Thus, we will initiate theoretical studies to examine the uncatalyzed solution and gas-phase reaction of CO2 with hydroxide, which will allow for insights into how HCAII catalyzes this reaction. The remainder of the proposed studies include investigations on the solution and gas-phase reaction of CO2 with water, which is an important reaction in the buffering of blood, studies on the catalytic mechanism of HCAII, studies on HCAII-substrate interactions, and a through scrutiny of known and potentially new HCAII inhibitors.
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