Hydrogen Tunneling in Enzyme Reactions

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Numerous examples exist in the literature documenting the importance of electron tunneling in enzymatic processes. Although much larger than the electron, protium possesses a de Brogile wave length which is similar to the expected dimension of the reaction coordinate in simple H transfer reactions. This property has led to the recognition that the behavior of protium is poised between classical and quantum mechanical behavior. Published studies from this laboratory have demonstrated that quantum effects contribute significantly to several enzyme catalyzed H transfer reactions at room temperature. Ongoing experiments suggest that this phenomenon will be a general one for enzyme catalyzed reactions. A major difficulty in the unambiguous demonstration of tunneling has been the absence of suitable experimental probes. A series of protocols have been developed toward this end: these include the comparative study of H/T and D/T isotope effects and their temperature dependences. Investigations during the next funding period will include the following areas: (1) A number of new enzyme catalyzed H transfer reactions will be examined for tunneling, in order to establish a data base for this phenomenon. In particular, the PI plans to focus on the flavin containing enzymes, monoamine oxidase and glucose oxidase: (2) Studies of the influence on tunneling of specific protein substrate interactions will be pursued through the use of site-specific mutagenesis; and (3) Computational work will be continued, with the goal of reproducing experimental data and providing suitable models for the mechanism of tunneling in enzyme reactions. %%% Although electron tunneling in proteins has been extensively studied, little attention has been paid to nuclear (hydrogen) tunneling. Dr. Klinman's elegant work has shown that nuclear tunneling contributes significantly to certain enzymatic reaction rates under biologically relevant conditions. This project builds and expands upon these groundbreaking observations.