Submitochondrial localization and partial purification of the succinylCoA: 3-hydroxy-3-methylglutarate coenzyme A transferase from rat liver.
The presence and the localization of the enzyme catalyzing the transfer of a coenzyme A molecule from succinyl-CoA to 3-hydroxy-3-methylglutarate has been established in rat liver mitochondria. The enzyme was found mainly in the mitochondrial matrix but some activity was also found in the inner membrane fraction. The enzyme has been purified about 100-fold from sonically-disrupted mitochondria by high-speed centrifugation, DEAE-cellulose chromatography, (NH4)2SO4 precipitation and Sephadex G-100 filtration. The enzymatic activity was recovered in the final step as a single peak. The coenzyme A transferase appears to have a molecular weight of 42 000, the highest activity at pH 8.5 and an energy of activation of 13 kcal/mol. Mercaptoethanol increases the activity and improves its stability. The enzyme is different from the succinylCoA: 3-oxoacids coenzyme A transferase and is active also on malonylCoA. The apparent Km values obtained for succinylCoA, malnylCoA and 3-hydroxy-3-methylglutarate were 2.2 . 10(-4) M, 3.7 . 10(-4) M and 1.7 . 10(-3) M, respectively. Acetoacetate, which is the final product of the mitochondrial metabolism of hydroxy-methylglutarylCoA, showed an inhibitory effect on the enzyme activity with a Ki of 0.5 mM. The physiological role of the enzyme is discussed.
D000090:Acetoacetates; D000818:Animals; D002848:Chromatography, DEAE-Cellulose; D002850:Chromatography, Gel; D015256:Coenzyme A-Transferases; D005260:Female; D006863:Hydrogen-Ion Concentration; D007651:Keto Acids / Q000494:pharmacology; D007700:Kinetics; D008297:Male; D008316:Malonyl Coenzyme A / Q000378:metabolism; D008623:Mercaptoethanol / Q000494:pharmacology; D008930:Mitochondria, Liver / Q000201:enzymology*; D008970:Molecular Weight; D051381:Rats; D011919:Rats, Inbred Strains; D013466:Sulfurtransferases / Q000302:isolation & purification*; D014461:Ultracentrifugation
D000090:Acetoacetates; D007651:Keto Acids; C016635:acetoacetic acid; D008316:Malonyl Coenzyme A; D008623:Mercaptoethanol; D013466:Sulfurtransferases; D015256:Coenzyme A-Transferases; C032820:succinyl-CoA-3-hydroxy-3-methylglutarate CoA transferase