Rat hepatic microsomal acetoacetyl-CoA reductase. A beta-ketoacyl-CoA reductase distinct from the long chain beta-ketoacyl-CoA reductase component of the microsomal fatty acid chain elongation system.
The present study provides evidence for a new rat liver microsomal enzyme, a short chain beta-ketoacyl (acetoacetyl)-CoA reductase, which is separate from the long chain beta-ketoacyl-CoA reductase component of the microsomal fatty acid chain elongation system. This microsomal reductase converts acetoacetyl-CoA to beta-hydroxybutyryl-CoA at a rate of 70 nmol/min/mg of protein; the enzyme has a specific requirement for NADH and appears to obtain electrons directly from the reduced pyridine nucleotide without the intervention of cytochrome b5 and its flavoprotein reductase. The apparent Km of the enzyme of the acetoacetyl-CoA was 21 microM and for the cofactor, 18 microM. The pH optimum was broad, ranging from 6.5 to 8.0. The product formed is the D-isomer of beta-hydroxybutyryl-CoA. High carbohydrate fat-free diet resulted in a small but significant (35%) increase in microsomal acetoacetyl-CoA reductase activity. The cytosol also contains this enzyme activity, measuring approximately 57% of that found in the microsomes. The mitochondrial activity which is 20-25% higher than the microsomal activity appears to be due to L-beta-hydroxyacyl-CoA dehydrogenase which converts acetoacetyl-CoA to L-beta-hydroxybutyryl-CoA. The microsomal acetoacetyl-CoA reductase activity was extracted from the microsomal membrane by 0.4 M KCl, resulting in an 8- to 10-fold purification; in addition, the long chain fatty acid elongation system was unaffected by this extraction procedure. Employing beta- hydroxyhexanoyl -CoA as a substrate, evidence is also provided for a separate dehydratase which acts on short chain substrates. Lastly, the liver microsomes had no detectable acetoacetyl-CoA synthetase or acetyl-CoA acetyltransferase activities. Hence, the possible involvement of the rat hepatic microsomal short chain beta-ketoacyl-CoA reductase, short chain beta-hydroxyacyl-CoA dehydratase, and the previously reported short chain trans-2-enoyl-CoA reductase in the hepatic utilization of acetoacetyl-CoA and in the synthesis of butyryl-CoA for hepatic lipogenesis is discussed.
D000090:Acetoacetates / Q000378:metabolism; D000105:Acetyl Coenzyme A / Q000031:analogs & derivatives / Q000378:metabolism; D000214:Acyl Coenzyme A; D000429:Alcohol Oxidoreductases / Q000032:analysis*; D000818:Animals; D004040:Dietary Carbohydrates / Q000008:administration & dosage; D064429:Fatty Acid Synthases / Q000032:analysis*; D007700:Kinetics; D008297:Male; D008316:Malonyl Coenzyme A / Q000378:metabolism; D008862:Microsomes, Liver / Q000201:enzymology*; D009243:NAD / Q000378:metabolism; D051381:Rats; D011919:Rats, Inbred Strains
D000090:Acetoacetates; D000214:Acyl Coenzyme A; D004040:Dietary Carbohydrates; D009243:NAD; C010667:acetoacetyl CoA; C016635:acetoacetic acid; D008316:Malonyl Coenzyme A; D000105:Acetyl Coenzyme A; D000429:Alcohol Oxidoreductases; C023744:acetoacetyl-CoA reductase; D064429:Fatty Acid Synthases