The effects of NADPH and 3-hydroxy-3-methylglutaryl-CoA on the thiol/disulfide redox behavior of rat liver microsomal 3-hydroxy-3-methylglutaryl-CoA reductase.
Microsomal 3-hydroxy-3-methylglutaryl-CoA reductase isolated from the livers of rats fed a diet containing cholestyramine (HMGR-C) is oxidized to a protein-SS-protein disulfide via a thermodynamically favorable thiol/disulfide exchange in glutathione redox buffers which approach the normal in vivo redox poise. In the presence of either substrate (NADPH or 3-hydroxy-3-methylglutaryl-CoA), the equilibrium thiol/disulfide redox behavior of HMGR-C is substantially different than that observed in the absence of substrates or in the presence of both substrates. NADPH present during redox equilibrium in a glutathione redox buffer decreases the equilibrium constant for formation of the protein-SS-protein disulfide (Kox,i) from 0.55 +/- 0.07 M to 0.18 +/- 0.02 M and increases the Kox,m for formation of an inactive protein-SS-glutathione mixed disulfide from less than 1 to 6 +/- 1. The presence of 3-hydroxy-3-methylglutaryl-CoA during redox equilibrium has a similar effect, decreasing the Kox,i for protein-SS-protein disulfide formation to 0.10 +/- 0.02 M and increasing the Kox,m for protein-SS-glutathione mixed disulfide formation to 3.8 +/- 0.9. A three-state model is developed which describes the simultaneous accumulation of protein-SS-protein and protein-SS-glutathione mixed disulfides at redox equilibrium with glutathione redox buffers. Because of the different redox behavior of the free and substrate-liganded forms of the enzyme, addition of 3-hydroxy-3-methylglutaryl-CoA or NADPH to HMGR-C at redox equilibrium results in increased reduction and activation of the enzyme.
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Verna and Marrs McLean Department of Biochemistry, Baylor College of Medicine, Houston, Texas 77030.
Gilbert
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D000214:Acyl Coenzyme A / Q000494:pharmacology*; D000818:Animals; D004229:Dithiothreitol / Q000494:pharmacology*; D005978:Glutathione / Q000378:metabolism / Q000494:pharmacology*; D006903:Hydroxymethylglutaryl CoA Reductases / Q000378:metabolism*; D007700:Kinetics; D008297:Male; D008433:Mathematics; D008862:Microsomes, Liver / Q000201:enzymology*; D008962:Models, Theoretical; D009249:NADP / Q000494:pharmacology*; D010084:Oxidation-Reduction; D051381:Rats; D011919:Rats, Inbred Strains
D000214:Acyl Coenzyme A; C008047:3-hydroxy-3-methylglutaryl-coenzyme A; D009249:NADP; D006903:Hydroxymethylglutaryl CoA Reductases; D005978:Glutathione; D004229:Dithiothreitol