Some properties of mitochondrial NAD+ linked malic enzyme and malate dehydrogenase from the flight muscles of Leptinotarsa decemlineata

Abstract Malic enzyme and malate dehydrogenase were purified to homogeneity from the flight muscles of the Colorado potato beetle, Leptinotarsa decemlineata , by gel filtration on Sephadex G-200 and affinity chromatography on 5-AMP Sepharose. Malic enzyme utilizes NAD + preferentially and is activated by low concentrations of manganese ions. The molecular weights of the native enzyme and its subunits were 220,000 and 49,000 as determined by gel filtration and SDS polyacrylamide gel electrophoresis respectively. In the direction of pyruvate formation, the enzyme showed apparent K m values of 0.09 mM for malate and 0.10 mM for NAD + and was inhibited by oxaloacetate, succinate, pyruvate, ATP, ADP and AMP. Under alkaline pH condition, purified malate dehydrogenase utilises malate and oxaloacetate at approximately the same rate. The enzyme showed a molecular weight of about 61,000 composing two subunits of 30,000. In the direction of oxaloacetate formation, the enzyme showed apparent K m values of 1.5 mM for malate and 0.22 mM for NAD + and was inhibited by succinate, pyruvate and the adenine nucleotides. The physiological significance of the biochemical properties of both enzymes is discussed.