Yanagawa H, Egami F
J Biol Chem. 1976 Jun 25;251(12):3637-44.
Asparagusate dehydrogenases I and II and lipoyl dehydrogenase have been obtained in homogeneous state from asparagus mitochondria. They are flavin enzymes with 1 mol of FAD/mol of protein. Asparagusate dehydrogenases I and II and lipoyl dehydrogenase have s20,w of 6.22 S, 6.39 S, and 5.91 S, respectively, and molecular weights of 111,000, 110,000, and 95,000 (sedimentation equilibrium) or 112,000, 112,000, and 92,000 (gel filtration). They are slightly acidic proteins with isoelectric points of 6.75, 5.75, and 6.80. Both asparagusate dehydrogenases catalyzed the reaction Asg(SH)2 + NAD+ equilibrium AsgS2 + NADH + H+ and exhibit lipoyl dehydrogenase and diaphorase activities. Lipoyl dehydrogenase is specific for lipoate and has no asparagusate dehydrogenase activity. NADP cannot replace NAD in any case. Optimum pH for substrate reduction of the three enzymes are near 5.9. Asparagusate dehydrogenases I and II have Km values of 21.5 mM and 20.0 mM for asparagusate and 3.0 mM and 3.3 mM for lipoate, respectively. Lipoyl dehydrogenase activity of asparagusate dehydrogenases is enhanced by NAD and surfactants such as lecithin and Tween 80, but asparagusate dehydrogenase activity is not enhanced. Asparagusate dehydrogenases are strongly inhibited by mercuric ion, p-chloromercuribenzoic acid, and N-ethylmaleimide. Amino acid composition of the three enzymes is presented and discussed.
天冬氨酸脱氢酶I和II以及硫辛酰胺脱氢酶已从芦笋线粒体中以均一状态获得。它们是黄素酶，每摩尔蛋白质含有1摩尔FAD。天冬氨酸脱氢酶I和II以及硫辛酰胺脱氢酶的沉降系数s20,w分别为6.22 S、6.39 S和5.91 S，沉降平衡法测得的分子量分别为111,000、110,000和95,000，凝胶过滤法测得的分子量分别为112,000、112,000和92,000。它们是微酸性蛋白质，等电点分别为6.75、5.75和6.80。两种天冬氨酸脱氢酶都催化反应Asg(SH)2 + NAD+ ⇌ AsgS2 + NADH + H+，并表现出硫辛酰胺脱氢酶和黄递酶活性。硫辛酰胺脱氢酶对硫辛酸具有特异性，没有天冬氨酸脱氢酶活性。在任何情况下，NADP都不能取代NAD。这三种酶底物还原的最适pH接近5.9。天冬氨酸脱氢酶I和II对天冬氨酸的Km值分别为21.5 mM和20.0 mM，对硫辛酸的Km值分别为3.0 mM和3.3 mM。NAD以及卵磷脂和吐温80等表面活性剂可增强天冬氨酸脱氢酶的硫辛酰胺脱氢酶活性，但不能增强天冬氨酸脱氢酶活性。天冬氨酸脱氢酶受到汞离子、对氯汞苯甲酸和N-乙基马来酰亚胺的强烈抑制。文中给出并讨论了这三种酶的氨基酸组成。
