STRUCTURES INFLUENCE ON REACTIVITY IN METALLOENZYMES

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DESCRIPTION: Research described in this proposal will be aimed at: Understanding how structure influences function and/or the characteristic properties of metalloenzyme active sites. Understanding how protein constraint can influence both the reactivity, and geometric/electronic structure of metalloenzymes. More specifically: Studies will continue to focus on the design and synthesis of analogues for the iron and cobalt sites of nitrile hydratases. Synthetic models, possessing a variety of structures, will be synthesized (using a versatile, one-pot synthesis), structurally characterized (using X-ra crystallography), and then screened for key biophysical properties such as (1) a S=1/2 spin-state (epr, magnetism (both solution and solid state)), and (2) a intense visible band near 700 nm. Reactions between our synthetic models and enzyme substrates (RCN),inhibitor (N3-), and inactivator (NO), will then be monitored both spectrophotometricall and by EPR. By systematically altering the structure of our models, and probing the influence that these changes have on the electronic, and reactivity, properties, we will determine if there is any correlation between structure an reactivity, and determine if spin-state and other electronic properties have a important influence on reactivity. Ligand constraints will be incorporated into our models, as a model for protei constraints, so that we can probe the influence that protein constraints can have on structure and reactivity. For example, by using a ligand with a fixed "backbone" length, angles can be constrained, and, in some cases, opened, so a to create a substrate binding site (entatic state). Similar constraints may be responsible for the reactivity of metalloenzymes. Studies aimed at understanding structure/reactivity relationships in other cysteine-ligated metalloenzymes, such as hydrogenase, and liver alcohol dehydrogenase, will also be pursued.