Mitochondrial NAD(P)-malic enzyme from herring skeletal muscle - Purification and some kinetic and regulatory properties

Mitochondrial NAD(P)-dependent malic enzyme [EC 1.1.1.39, L-malate: NAD+ oxidoreductase (decarboxylating)] was purified from herring skeletal muscle to a specific activity of 8.2 μmol/min/mg. The purification procedure involved chromatography on DEAE-cellulose, Red Agarose and a Sephacryl S-300 with a final recovery of 38% of enzyme activity. This enzyme catalyzes the oxidative decarboxylation of malate in the presence of either NAD or NADP in the presence of Mn2+. Some kinetic characteristics of this enzyme were determined. The pH optimum of activity is 7.0. ATP was shown to be a competitive inhibitor with malate. The inhibition by ATP displayed hyperbolic competitive kinetics with a Ki (ATP) of 0.28 mM in the presence of NAD and 0.75 mM in the presence of NADP. Fumarate reversed ATP inhibition. In vivo, regulation of NAD(P)-dependent malic enzyme might respond to changing levels of mitochondrial ATP and fumarate with the enzyme undergoing kinetic activation by an increase in the concentration of mitochondrial fumarate which could reverse enzyme inhibition by ATP