来自在甲醇上生长的假丝酵母属N-16的两种苹果酸脱氢酶的纯化及性质

Purification and properties of two malate dehydrogenases from Candida sp. N-16 grown on methanol.

作者信息

Yoshikawa J, Seki K, Shinoyama H, Fujii T

机构信息

Department of Bioresources Science, Graduate School of Science and Technology, Chiba University, Matsudo-shi, Japan.

出版信息

Biosci Biotechnol Biochem. 2001 Jul;65(7):1659-62. doi: 10.1271/bbb.65.1659.

DOI:10.1271/bbb.65.1659

PMID:11515554

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 Vmax(oxaloacetate)/Vmax(malate) and Kcats (555 s(-1) for oxaloacetate, 481 s(-1) for NADH) of MDH-M2 were higher than the ratio (1.37) of Vmax and Kcats (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.

摘要

在一株利用甲醇的酵母——假丝酵母N-16中发现了两种苹果酸脱氢酶（MDH-M1和MDH-M2）。MDH-M2由甲醇诱导产生。这些酶被纯化成为电泳和等电聚焦均一的蛋白质。MDH-M1和MDH-M2的分子量估计分别约为78,000（同型二聚体）和160,000（同型四聚体）。这两种酶的几个动力学特性存在显著差异。MDH-M2的Vmax（草酰乙酸）/Vmax（苹果酸）值（2.07）和催化常数（草酰乙酸为555 s(-1)，NADH为481 s(-1)）分别高于MDH-M1的该比值（1.37）和催化常数（草酰乙酸为241 s(-1)，NADH为271 s(-1)）。MDH-M2的活性受到高浓度NAD+的抑制，而MDH-M1的活性则受到草酰乙酸的抑制。