title: Biosynthesis and characterization of (S)-and (R)-3-hydroxy-3-methylglutaryl coenzyme A.
abstract: (S)-3-Hydroxy-3-methylglutaryl coenzyme A (HMG-CoA), the physiologic substrate of HMG-CoA reductase and of HMG-CoA lyase, is available commercially only as (R,S)-HMG-CoA, a mixture of diastereomers. To provide (S)-HMG-CoA for our continuing investigation of HMG-CoA reductase, we used homogeneous, overexpressed Pseudomonas mevalonii HMG-CoA reductase (EC 1.1.1.88) and an NAD(+)-regenerating system to convert (R)-mevalonate to (S)-HMG-CoA with an overall yield in excess of 50%. We also used P. mevalonii HMG-CoA lyase (EC 4.1.3.4) to prepare (R)-HMG-CoA from (R,S)-HMG-CoA. Each diastereomer was then isolated by ion-exchange chromatography. Large-scale preparations provide for economical production of (S)-HMG-CoA, particularly when recovered coenzyme A is recycled. (S)-HMG-CoA was evaluated as a substrate, and (R)-HMG-CoA as an inhibitor, for the P. mevalonii enzymes HMG-CoA reductase and HMG-CoA lyase, and for Syrian hamster HMG-CoA reductase (EC 1.1.1.34). For both HMG-CoA reductases, (R)-HMG-CoA inhibited competitively with respect to (S)-HMG-CoA. The ratio Ki/Km was 0.7 +/- 0.1 and 0.6 +/- 0.2 for the bacterial and hamster enzymes, respectively. By contrast, (R)-HMG-CoA did not inhibit P. mevalonii HMG-CoA lyase.
authors.affiliation: Department of Biochemistry, Purdue University, West Lafayette, Indiana 47907-1153.
authors.forename: K M
authors.initials: KM
authors.lastname: Bischoff
authors.forename: V W
authors.initials: VW
authors.lastname: Rodwell
mesh_terms: D000214:Acyl Coenzyme A / Q000096:biosynthesis* / Q000302:isolation & purification; D000818:Animals; D002852:Chromatography, Ion Exchange; D003124:Colorimetry; D006224:Cricetinae; D006903:Hydroxymethylglutaryl CoA Reductases / Q000378:metabolism*; D007700:Kinetics; D008647:Mesocricetus; D007652:Oxo-Acid-Lyases / Q000378:metabolism*; D011549:Pseudomonas / Q000201:enzymology; D011994:Recombinant Proteins / Q000378:metabolism; D013056:Spectrophotometry, Ultraviolet; D013237:Stereoisomerism
chemical_list: D000214:Acyl Coenzyme A; D011994:Recombinant Proteins; C008047:3-hydroxy-3-methylglutaryl-coenzyme A; D006903:Hydroxymethylglutaryl CoA Reductases; D007652:Oxo-Acid-Lyases; C021410:3-hydroxy-3-methylglutaryl-coenzyme A lyase