The overall goal of these proposed studies is to evaluate the factors which may affect the redox potentials of the b cytochromes, including the nature of the axial ligands, axial ligand plane orientation, the effect of hydrogen-bonding or deprotonation of imidazole ligands, the nature of the dielectric medium surrounding the heme, and the effect of buried charged groups near the heme. To accomplish this goal, both model compounds and proteins will be investigated. Model hemes will be utilized to probe the effect of axial ligand plane orientation on spectroscopic and redox properties, including proposed correlations of epr, Mossbauer and solid state structural properties, preparation and electrochemical study of tow series of porphyrin isomers designed to hold axial ligands in parallel or perpendicular plans, and CPMAS nmr studies of frozen solutions of iron porphyrins. Electronic and steric effects of axial ligands and anions on the structural, spectroscopic, and redox properties of metalloporphyrins will be elucidated, utilizing 59Co and 57Fe nmr spectroscopy, investigations of the properties of Fe(III) and Fe(II) complexes with phosphines, axial ligand exchange kinetics, preparation and investigation of 5-corrdiante Ni(II) porphyrins, and investigation of the effect of deportonation of coordinated imidazoles on the reactivity of the dioxygen complexes of Co(II) and Fe(II) porphyrins. In addition, a new series of pyrrole- substituted synthetic porphyrin isomers will be prepared in order to probe the effect of pyrrole substitution pattern on spectroscopic and redox properties, and the dependence of the Fe(III)/(II) redox potential on solvent polarity will be determined. Native and site-directed mutagenetically modified cytochromes b5 will be utilized to investigate the effect of heme and axial ligand modifications on orbital energies and redox properties. Specific studies will involve nmr investigations of the effect of heme propionate esterification on cation binding and charge neutralization in oxidized and reduced cytochrome b5, nmr, epr and electrochemical investigations of genetically modified cytochromes b5 in which a missing histidine ligand has been replaced by an exogenous imidazole, and ultrasonic measurement of the adiabatic compressibility of ferri-, ferro- and apocytochrome b5.
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