Axley M J, Grahame D A, Stadtman T C
Laboratory of Biochemistry, National Heart, Lung, and Blood Institute, National Institutes of Health, Bethesda, Maryland 20892.
J Biol Chem. 1990 Oct 25;265(30):18213-8.
The formate-hydrogen lyase complex of Escherichia coli decomposes formic acid to hydrogen and carbon dioxide under anaerobic conditions in the absence of exogenous electron acceptors. The complex consists of two separable enzymatic activities: a formate dehydrogenase and a hydrogenase. The formate dehydrogenase component (FDHH) of the formate-hydrogen lyase complex was purified to near homogeneity in two column chromatographic steps. The purified enzyme was composed of a single polypeptide of molecular weight 80,000 as determined by sodium dodecyl sulfate-polyacrylamide gel electrophoresis. Metal analysis showed each mole of enzyme contained 3.3 g atoms of iron. Denaturation of FDHH released a compound which, when oxidized, displayed a fluorescence spectrum similar to that of the molybdopterin cofactor found in certain other enzymes. The enzyme contained selenium in the form of selenocysteine as determined by radioactive labeling of the enzyme with 75Se and amino acid analysis. FDHH activity was maximal between pH 7.5 and 8.5; however, the enzyme was maximally stable at pH 5.3-6.4 and highly unstable above pH 7.5. Nitrate and nitrite salts caused a drastic reduction in activity. Although azide inhibited FDHH activity, it also protected the enzyme from inactivation by oxygen.
大肠杆菌的甲酸 - 氢裂解酶复合物在厌氧条件下且无外源电子受体时，可将甲酸分解为氢气和二氧化碳。该复合物由两种可分离的酶活性组成：一种甲酸脱氢酶和一种氢化酶。通过两步柱色谱法将甲酸 - 氢裂解酶复合物的甲酸脱氢酶组分（FDHH）纯化至接近均一。经十二烷基硫酸钠 - 聚丙烯酰胺凝胶电泳测定，纯化后的酶由一条分子量为80,000的单一多肽组成。金属分析表明，每摩尔酶含有3.3克原子的铁。FDHH变性后释放出一种化合物，该化合物氧化时显示出的荧光光谱与某些其他酶中发现的钼蝶呤辅因子的荧光光谱相似。通过用75Se对酶进行放射性标记和氨基酸分析确定，该酶以硒代半胱氨酸的形式含有硒。FDHH的活性在pH 7.5至8.5之间最大；然而，该酶在pH 5.3 - 6.4时最稳定，在pH 7.5以上高度不稳定。硝酸盐和亚硝酸盐会导致活性急剧降低。虽然叠氮化物抑制FDHH活性，但它也保护酶不被氧气灭活。
