Kinetic Properties of NADP+-Dependent Decarboxylating Malate Dehydrogenase from Corn Leaves

Abstract

One isoform of NADP+-dependent decarboxylating malate dehydrogenase (EC 1.1.1.40) was found in the leaf mesophyll of corn. The enzyme was purified in four stages: homogenization, fractionation with ammonium sulfate, gel filtration on Sephadex G-25, and ion exchange on Sephacel. The specific activity of the purified, electrophoretically homogeneous preparation was 92 E/mg of protein, while its yield was 15%, and the purification degree was 109. The relative electrophoretic mobility of the decarboxylating malate dehydrogenase was 0.1. The optimal pH for enzyme functioning during the catalysis of forward and reverse reactions was 8.0. The Michaelis constants were determined for the direct and reverse reactions: 5.5 mM for malate and 1.3 mM for pyruvate. Moreover, the affinity for coenzymes varied significantly. Thus, KM was equal to 4.5 mM for NADP+ and 0.9 mM for NADP.