Schmitz R A, Richter M, Linder D, Thauer R K
Laboratorium für Mikrobiologie, Philipps-Universität Marburg, Federal Republic of Germany.
Eur J Biochem. 1992 Jul 15;207(2):559-65. doi: 10.1111/j.1432-1033.1992.tb17082.x.
Methanobacterium wolfei is a thermophilic methanogenic archaeon which requires tungsten or molybdenum for growth. We have found that the organism contains two formylmethanofuran dehydrogenases, one of which is a tungsten enzyme. Indirect evidence indicates that the other formylmethanofuran dehydrogenase is a molybdenum enzyme. The tungsten enzyme was purified and characterized. The native enzyme had an apparent molecular mass of 130 kDa. SDS/PAGE revealed a composition of three subunits of apparent molecular mass 35, 51 and 64 kDa, the N-terminal amino acid sequences of two of which were determined. 0.3-0.4 mol tungsten/mol enzyme was found but no molybdenum. The pterin cofactor was identified as molybdopterin guanine dinucleotide. The purified enzyme exhibited a specific activity of 8.3 mumol.min-1.mg protein-1 and an apparent Km for formylmethanofuran and methylviologen of 13 microM and 0.4 mM, respectively. The optimum temperature for activity was 65 degrees C. At 40-60 degrees C, the rate increased with a Q10 of 1.9; the activation energy of the reaction was 45 kJ/mol. The enzyme was found to require potassium ions for thermostability. The oxygen-sensitive enzyme was not inactivated by cyanide.
沃氏甲烷杆菌是一种嗜热产甲烷古菌，其生长需要钨或钼。我们发现该生物体含有两种甲酰甲烷呋喃脱氢酶，其中一种是钨酶。间接证据表明另一种甲酰甲烷呋喃脱氢酶是钼酶。对钨酶进行了纯化和表征。天然酶的表观分子量为130 kDa。SDS/PAGE显示其由三个亚基组成，表观分子量分别为35、51和64 kDa，并测定了其中两个亚基的N端氨基酸序列。发现每摩尔酶含有0.3 - 0.4摩尔钨，但不含钼。蝶呤辅因子被鉴定为钼蝶呤鸟嘌呤二核苷酸。纯化后的酶比活性为8.3 μmol·min⁻¹·mg蛋白⁻¹，对甲酰甲烷呋喃和甲基紫精的表观Km分别为13 μM和0.4 mM。酶活性的最适温度为65℃。在40 - 60℃时，反应速率随Q10为1.9增加；反应的活化能为45 kJ/mol。发现该酶的热稳定性需要钾离子。这种对氧敏感的酶不会被氰化物灭活。
