Influence of glutathione on the catalytic activity of reconstituted cytochrome P450 3A4.
The role of NADPH reductase and cytochrome b5 on glutathione (GSH)-induced stimulation of P450 3A4 activity was investigated. GSH increased the Vmax of testosterone 6 beta-hydroxylation without changing the K(m) for testosterone whereas it decreased the K(m) for NADPH-P450 reductase. Addition of cytochrome b5 inhibited testosterone 6 beta-hydroxylation in the reconstituted system, depleting GSH, while it dramatically enhanced the rate of testosterone 6 beta-hydroxylation in the presence of GSH. Cumene hydroperoxide-mediated P450 3A4 activity, which is independent of NADPH-P450 reductase and cytochrome b5, was not affected by GSH. High concentration of GSH above 4 mM was inhibitory in the reconstituted systems. These results suggest that GSH increases the apparent affinity between P450 3A4 and NADPH-P450 reductase, and between P450 3A4 and cytochrome b5, but has no effect on the affinity between P450 3A4 and testosterone.
Kim
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BR
Department of Biochemistry, School of Medicine, Won Kwang University, Iksan, Korea.
Kim
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DH
D001555:Benzene Derivatives / Q000378:metabolism; D002474:Cell-Free System; D051544:Cytochrome P-450 CYP3A; D003577:Cytochrome P-450 Enzyme System / Q000187:drug effects / Q000378:metabolism*; D015786:Cytochromes b5 / Q000187:drug effects / Q000378:metabolism*; D005978:Glutathione / Q000494:pharmacology*; D006900:Hydroxylation; D007700:Kinetics; D006899:Mixed Function Oxygenases / Q000187:drug effects / Q000378:metabolism*; D009247:NADH, NADPH Oxidoreductases / Q000187:drug effects / Q000378:metabolism*; D009251:NADPH-Ferrihemoprotein Reductase; D010084:Oxidation-Reduction / Q000187:drug effects; D013739:Testosterone / Q000378:metabolism*
D001555:Benzene Derivatives; D013739:Testosterone; D015786:Cytochromes b5; D003577:Cytochrome P-450 Enzyme System; D006899:Mixed Function Oxygenases; C104464:CYP3A protein, human; D051544:Cytochrome P-450 CYP3A; D009247:NADH, NADPH Oxidoreductases; D009251:NADPH-Ferrihemoprotein Reductase; D005978:Glutathione; C007164:cumene hydroperoxide