The aim of the proposed research is to elucidate the electronic structures of metallo-protein active centers using primarily Mossbauer, EPR, and ENDOR spectroscopy. Of particular interest are the heme proteins that interact with O2 or H2O2 and their various intermediates. The ultimate goal is to understand the structural and dynamic features that control reactivity and function. The following systems will be studied: (i) Work in progress on O2 activation and product formation in the bacterial monoxygenase cytochrome P450cam will continue. Earlier data suggest that an electron can be transferred to the ternary complex of cytochrome P450cam with substrate and O2 by low-temperature X-ray irradiation, and that new EPR- active heme species are formed on stepwise annealing to higher temperatures. The nature of these intermediates will be explored and the products of the reaction will be analyzed. Low-temperature techniques will also be used to search for intermediates in the product-forming steps of the native system and with modified substrates and proteins. (ii) Prof. Sligar's group has produced genetically modified myoglobins, which will be characterized by Mossbauer, EPR and ENDOR spectroscopy in a collaborative effort to correlate amino acid substitutions with changes in function, structure and electronic state. (iii) A Mossbauer study of lignin and manganese peroxidase from the white rot fungus Phanerochaete chrysosporium and of their reaction products with H2O2 and substrate will be started in collaboration with Prof. Gold, Oregon Graduate Institute. (iv) Two terminal oxidases, cytochrome o and cytochrome d from E. coli, will be investigated by EPR, ENDOR and Mossbauer spectroscopy in collaboration with Prof. Gennis. (v) Work on hemerythrin and structurally related binuclear iron proteins and model compounds will continue.
