Isolation and regulatory properties of mitochondrial malic enzyme from rat skeletal muscle.

Mitochondrial malic enzyme (L-malate:NADP+ oxidoreductase (oxalo-acetate-decarboxylating), EC 1.1.1.40) has been isolated from rat skeletal muscle by (NH4)2SO4 fractionation, chromatography on DEAE-cellulose and Ultrogel AcA 34. Specific activity of the purified enzyme was 25 micromol/min per mg of protein which corresponds to about 840-folf purification. The enzyme was shown to carboxylate pyruvate in the presence of high concentrations of KHCO3 and pyruvate at about 15% of the rate of the forward reaction. The Km values determined at pH 7.2 for malate, NADP and Mn2+ were 0.33 mM, 6.8 microM and 7.1 microM, respectively. The Km values for pyruvate, NADPH and KHCO3 were 8.3 mM, 19.6 microM, and 24.4 mM, respectively. Purified enzyme showed allosteric properties at low concentration of malate and this characteristic can be modified by succinate and fumarate which do not affect the maximum velocity of the reaction. The pH optimum for decarboxylation reaction was between 7.2 and 8.4. Possible metabolic role of mitochondrial malic enzyme in skeletal muscle is discussed.