The present proposal outlines a group of projects requiring a state-of-the-art electron paramagnetic resonance spectrometer in examining a number of biological problems. Applications of the instrument being requested include: (1) structural and mechanistic studies of the molybdenum centers of xanthine oxidase and sulfite oxidase, (2) studies of the effects of chemical modification of plastocyanin on the structure and function of its copper center, (3) determination of the oxidation-reduction potentials of genetically engineered forms of the flavodoxin from Desulfovibrio vulgaris, (4) a spin-label study of substrate binding to xanthine oxidase aimed at establishing the Kd for binding of xanthine to the oxidized form of the enzyme and the number of substrate binding sites present in the active site, (5) use of laser-induced excitation of carbonmonoxy cytochrome c oxidase and nitrosylhemoglobin to examine changes in the EPR signals of these centers, and (6) an examination of spin-stabilized free radicals generated by cultured heart cells under ischemic conditions to determine the extent to which such radicals play a role in ischemia-related tissue damage. Together, these projects constitute a major application of electron paramagnetic resonance spectroscopy that would greatly enhance the basic research capability of an extended group of biochemists at Ohio State University
