Purification and Properties of Succinyl‐CoA:3‐Oxo‐Acid CoA‐Transferase from Rat Brain

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Abstract
Rat brain succinyl‐CoA:3‐oxo‐acid CoA‐transferase (3‐Oxo‐acid CoA‐transferase, EC 2.8.3.5), the first committed enzyme in the oxidation of ketone bodies in mitochondria, was purified to apparent homogeneity as judged by polyacrylamide gel electrophoresis. The enzyme has an apparent molecular weight of 90,000 as determined by (3‐150 Sephadex chromatography, and an apparent subunit molecular weight of 53,000 as determined by sodium dodecyl sulfate polyacrylamide gel electrophoresis. The specific activity of the purified enzyme was approximately 161 μmol/min/mg of protein. Initial velocity studies of the forward reaction (acetoacetate → acetoacetyl‐CoA) are consistent with a “ping pong” mechanism. Substrate inhibition appears above approximately 1 mM acetoacetate. Apparent Km, values were 70 μM for acetoacetate and 156 μM for succinyl‐CoA (the forward reaction), and 59 μM for acetoacetyl‐CoA and 25 mM for succinate (the reverse reaction). These values are markedly different from those reported for this enzyme from pig heart.