The cytochrome P450s metabolize a wide variety of xenobiotic and endogenous compounds. Biochemical, biophysical, and computational approaches were applied to elucidate the structure-function relationships which govern the interactions of P450s with substrates, inhibitors, membrane lipids, and microsomal proteins. Since these interactions modulate P450 activity, elucidation of their molecular mechanism will aid in (1) clarifying the mechanism of P450-mediated drug and carcinogen metabolism; (2) defining the role of individual P450s in the metabolism of endogenous and environmental chemicals; and (3) development of specific P450 inhibitors.Kinetics of CO binding to P450 were measured, using the flash photolysis method. The Maximum Entropy Method was used to obtain kinetic distribution profiles that correspond to conformational landscapes. The results show that the structures of P450s 1A1 and 3A4 are best viewed as distributions of conformers. This approach revealed a substrate-dependency of these distributions. In contrast, monoexponential kinetics were observed for P450 2E1, which indicates a more homogeneous conformation.Homology modeling techniques were used to generate a molecular model of human P450 1A2. On the basis of active groups in the predicted substrate binding site, we computationally searched a small molecule database for complementary molecules. Several P450 inhibitors were identified by this approach. In addition, mutation of residues in this site modified the catalytic activity. These results thus provide support for our P450 homology model.
