Submitochondrial localization and partial purification of the succinylCoA: 3-hydroxy-3-methylglutarate coenzyme A transferase from rat liver

Abstract

The presence and the localization of the enzyme catalyzing the transfer of a coenzyme A molecule from succinylCoA 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 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 $\text{K}{}_{\mathrm{<mtext>m</mtext>}}$ values obtained for succinylCoA, malnyCoA 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 $\text{K}{}_{\mathrm{i}}$ of 0.5 mM. The physiological role of the enzyme is discussed.