Purification and Properties of Two Malate Dehydrogenases from Candida sp. N-16 Grown on Methanol

Abstract

Two malate dehydrogenases (MDH-M1 and MDH-M2) were found in a methanol-using yeast, Candida sp. N-16. MDH-M2 was induced with methanol. These enzymes were purified as electrophoretically and isoelectrophoretically homogeneous proteins. The molecular weights of MDH-M1 and MDH-M2 were estimated to be about 78,000 (homodimer) and 160,000 (homotetramer). Several kinetic properties were significantly different between the two enzymes. The value (2.07) of V_{max(oxaloacetate)}/V_max(malate) and K_cats (555 s^-1 for oxaloacetate, 481 s^-1 for NADH) of MDH-M2 were higher than the ratio (1.37) of V_max and K_cats(241 s^-1 for oxaloacetate, 271 s^-1 for NADH) of MDH-M1, respectively. The activity of MDH-M2 was inhibited by a high concentration of NAD⁺ and the activity of MDH-M1 by oxaloacetate.