Differential Effect of Copper (II) on the Cytochrome P450 Enzymes and NADPH−Cytochrome P450 Reductase: Inhibition of Cytochrome P450-Catalyzed Reactions by Copper (II) Ion

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Abstract
Inhibitory effects of Cu2+ on the cytochrome P450 (P450)-catalyzed reactions of liver microsomes and reconstituted systems containing purified P450 and NADPH−P450 reductase (NPR) were seen. However, Zn2+, Mg2+, Mn2+, Ca2+, and Co2+ had no apparent effects on the activities of microsomal P450s. Cu2+ inhibited the reactions catalyzed by purified P450s 1A2 and 3A4 with IC50 values of 5.7 and 8.4 μM, respectively. Cu2+ also inhibited reduction of cytochrome c by NPR (IC50 value of 5.8 μM). Copper caused a decrease in semiquinone levels of NPR, although it did not disturb the rate of formation of semiquinone. P450 reactions supported by an oxygen surrogate, tert-butyl hydroperoxide, instead of NPR and NADPH, were inhibited by the presence of Cu2+. The results indicate that Cu2+ inhibits the P450-catalyzed reactions by affecting both P450s and NPR. It was also found that the inhibition of catalytic activities of P450s by Cu2+ involves overall conformational changes of P450s and NPR, investigated by CD and intrinsic fluorescence spectroscopy. These results suggest that the inhibitory effect of Cu2+ on the P450-catalyzed reactions may come from the inability of an efficient electron transfer from NPR to P450 and also the dysfunctional conformation of NPR and P450.