HIGH OXIDATION STATE HEMOPROTEINS AND HALIDE ACTIVATION

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This application is concerned with further elucidating the chemistry of halide-activating peroxidase enzymes. Studies will focus on defining the active site molecular structure, the electronic structure, and the reactivity properties of lactoperoxidase and chloroperoxidase. Emphasis will be placed on nuclear magnetic resonance (NMR) spectrocopy of the hemoproteins, as well as synthesis and reactions of appropriate iron porphyrin model compounds. The importance of this work is found in the observation that major classes of mammalian peroxidases exert their biosynthetic or biological defense functions through activation of halide or pseudohalide ions. Particular attention will be given to elucidating the chemistry of lactoperoxidase-thiocynate interactions, as this system functions to protect exocrine fluids from bacterial invasion. Likewise, the chloride-activating bactericidal functions of leukocyte myeloperoxidase will be modeled by the microbial chloroperoxidase enzyme. Specific objectives follow: (1) electronic structures of lactoperoxidase and chloroperoxidase in various redox states will be better defined and compared with those of other hemoproteins through high-field NMR measurements; (2) the active site molecular structure will be probed through carbon-13 and nitrogen-15 NMR spectroscopy, of ligands (CO, NC-, SCN, and isonitriles) bound at the iron center, (3) non-heme anion binding sites will be probed through use of multinuclear NMR methods; and (4) attempts will be made to detect the direct products and discern the mechanisms of halide and pseudohalide activation in the peroxidases and in model heme compounds.