The P-450-dependent mixed-function oxidases play a critical role in the metabolic activation and detoxication of many chemical carcinogens. The primary purpose of the proposed research is to gain a better understanding of the mechanisms by which the cytochrome P-450-dependent mixed-function oxidases catalyze carcinogen metabolism. In order to achieve this objective, the specific aims of the research proposal are: 1) To ivestigate inter- and intramolecular isotope effects for the peroxidase- and cytochrome P-450-catalyzed metabolism of N-methylcarbazole to gain a better understanding of the mechanism(s) of N- demetheylation reactions: 2) To determine the source of the oxygen incorporated into products during metabolism of N- methylcarbazole by purified isozymes of cytochrome P-450 and peroxidases to obtain information regarding the mechanism of oxygen insertion; 3) To investigate the possible involvement of free radicals and iminium ions as intermediates in the metabolism of N-methylcarbazole; 4) To characterize the substrate-binding sites of purified isozymes of cytochrome P-450 using photoaffinity-labeling reagents and identify polypeptide sequences critical for substrate binding; 5) To investigate the mechanism- based inactivation of purified isozymes of cytochrome P-450 during the metabolism of N-methylcarbazole, to identify the amino acid(s) and peptide(s) at the active site of the enzyme which are modified during inactivation, and to determine the mechanism(s) by which inactivation occurs; and 6) To investigate the mechanism-based inactivaton of P-450 during 1- aminobenzotriazole metabolism. The P-450-dependent mixed- function oxidases are present in almost all human tissues and play a critical role in the metabolism of many chemical carcinogens. These studies should provide a better understanding of the mechanism(s) by which these enzyme systems activate chemical carcinogens. The determination of the mechanism(s) by which these enzymes undergo irreversible """"""""suicide"""""""" inactivation during the metabolism of certain substrates could be valuable for developing approaches for selectively modulating the catalytic activity of these enzymes. The results of these studies could provide information for developing methods to decrease the risk of developing cancer.

National Institute of Health (NIH)
National Cancer Institute (NCI)
Method to Extend Research in Time (MERIT) Award (R37)
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Chemical Pathology Study Section (CPA)
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Wayne State University
Schools of Medicine
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