Dimethylamine Dehydrogenase from Hyphomicrobium X: Purification and Some Properties of a New Enzyme that Oxidizes Secondary Amines
Dimethylamine Dehydrogenase from Hyphomicrobium X: Purification and Some Properties of a New Enzyme that Oxidizes Secondary Amines
复制标题
来自丝菌 X 的二甲胺脱氢酶：氧化仲胺新酶的纯化和一些性质
DOI：
10.1099/00221287-115-1-49
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发表时间：
1979
期刊：
影响因子：
1.5
通讯作者：
W. Harder
中科院分区：
文献类型：
--
作者：
J. Meiberg;W. Harder
总结：通过硫酸铵分级分离和DEAE-纤维素层析，从以二甲胺为唯一碳源厌氧生长的Hyphomicrobium X中纯化了15.6倍的二甲胺脱氢酶。该制剂不含三甲胺脱氢酶。该酶的分子量为176 000和亚基分析十二烷基硫酸钠-聚丙烯酰胺凝胶电泳表明，它由两个，可能是相同的，分子量为91 000的亚基。在441 nm处有最大吸收。用二甲胺还原酶在356 nm处产生新的最大吸收，而在441 nm处的吸收降低。二甲胺氧化的最适pH为8.1。在该反应中，形成化学计量量的甲胺和甲醛作为产物。该酶对仲胺表现出绝对的特异性;二甲胺、甲基乙胺、二乙基胺、甲基丙胺、乙基丙胺和甲基乙醇胺被氧化，而伯胺、叔胺和季铵盐不被氧化。除了吩嗪硫酸甲酯，只有吩嗪硫酸乙酯，沃斯特蓝和亚甲蓝作为人工电子受体。表观K
在pH7.7时，二甲胺的酶活为15.6。1.6微米级
.三甲胺是一种有效的二甲胺氧化的竞争性抑制剂，
1个7.1 μm
.这种抑制二甲胺脱氢酶的三甲胺可能解释了观察到的积累二甲胺在厌氧生长的菌丝微菌X在三甲胺。
Summary: Dimethylamine dehydrogenase was purified 15.6-fold from Hyphomicrobium X grown anaerobically on dimethylamine as sole carbon source by ammonium sulphate fractionation and chromatography on DEAE-cellulose. The preparation was free from trimethylamine dehydrogenase. The molecular weight of the enzyme was 176 000 and subunit analysis by sodium dodecyl sulphate–polyacrylamide gel electrophoresis indicated that it consists of two, probably identical, subunits with molecular weights of 91 000. The absorption spectrum showed a maximum at 441 nm. Reduction of the enzyme with dimethylamine produced a new absorption maximum at 356 nm, while the absorption at 441 nm decreased. The pH optimum for the oxidation of dimethylamine was 8.1. In this reaction, stoicheiometric amounts of methylamine and formaldehyde were formed as products. The enzyme showed absolute specificity towards secondary amines; dimethylamine, methylethylamine, diethyl-amine, methylpropylamine, ethylpropylamine and methylethanolamine were oxidized while primary and tertiary amines and quaternary ammonium salts were not. Apart from phenazine methosulphate, only phenazine ethosulphate, Wurster's blue and methylene blue served as artificial electron acceptors. The apparent K
m of the enzyme for dimethylamine at pH 7.7 was 15.6. 1.6 μm
. Trimethylamine was a potent competitive inhibitor of dimethylamine oxidation with an apparent K
1 of 7.1 μm
. This inhibition of dimethylamine dehydrogenase by trimethylamine probably explains the observed accumulation of dimethylamine during anaerobic growth of Hyphomicrobium X on trimethylamine.