Catalysis by Syrian hamster 3-hydroxy-3-methylglutaryl-coenzyme A reductase. Proposed roles of histidine 865, glutamate 558, and aspartate 766.

We employed the overexpressed catalytic domains of wild-type Syrian hamster 3-hydroxy-3-methylglutaryl-coenzyme A reductase (HMG-CoA reductase, EC 1.1.1.34) and of mutant enzymes E558Q, H865Q, and D766N to investigate the roles of Glu558, His865, and Asp786 in catalysis. Five reactions were studied: reductive deacylation of HMG-CoA or of mevaldyl-CoA to mevalonate, reduction of mevaldehyde to mevalonate, oxidation of mevaldyl-CoA to HMG-CoA, and oxidative acylation of mevaldehyde to HMG-CoA. While only the wild-type enzyme catalyzed all five reactions, mutant enzymes E558Q and H865Q catalyzed other reactions at significant rates. Mutant enzyme D766N, although apparently structurally similar to the wild-type enzyme, was inactive for all five reactions. While an ineffective catalyst for the overall reaction, mutant enzyme H865Q catalyzed the reduction of mevaldehyde to mevalonate at about the wild-type rate. Coenzyme A, which stimulated mevaldehyde reduction by the wild-type enzyme, inhibited mutant enzyme H865Q, apparently due to an impaired ability to protonate the coenzyme A thioanion, CoAS-. Based on these data, we infer that Glu558, His865, and Asp766 function in catalysis at the indicated stage of the overall reaction, [formula: see text] and we propose a revised mechanism for catalysis by mammalian HMG-CoA reductases.