Membrane-bound F420H2-dependent heterodisulfide reduction in Methanococcus voltae
Membrane-bound F420H2-dependent heterodisulfide reduction in Methanococcus voltae
复制标题
伏氏甲烷球菌中膜结合的 F420H2 依赖性异二硫键还原
DOI：
10.1007/s002030050686
复制
发表时间：
1999
影响因子：
2.8
通讯作者：
U. Deppenmeier
中科院分区：
文献类型：
--
作者：
Jens Brodersen;G. Gottschalk;U. Deppenmeier
由 H2+CO2 或甲酸盐生长的伏氏甲烷球菌细胞制备的洗涤膜催化辅酶 F420H2 的氧化和 2-巯基乙磺酸盐和 7-巯基庚酰苏氨酸磷酸盐的异二硫键 (CoB-S-S-CoM) 的还原，这是产甲烷途径的末端电子受体。反应遵循 1:1 化学计量，方程式为：F420H2+ COB–S–S–CoM → F420+ CoM–SH + CoB–SH。这些发现表明，该反应取决于膜结合的 F420H2 氧化酶和异二硫键还原酶，后者在细胞裂解后仍部分与膜结合。为了阐明 F420H2 氧化蛋白的性质，用去垢剂溶解洗涤过的膜，并通过蔗糖密度离心、阴离子交换色谱和凝胶过滤纯化酶。多项证据表明 F420H2 氧化是由膜相关的 F420 还原氢化酶催化的。纯化的蛋白质催化甲基紫精和 F420 的 H2 依赖性还原。表观分子量和亚基组成（43、37 和 27 kDa）与已从 Mc 中纯化的 F420 还原氢化酶几乎相同。伏特。此外，37-kDa亚基的N-末端与从编码来自Mc的含硒F420还原氢化酶的操纵子的fruG基因推导的氨基酸序列相同。伏特。在此生物体中未发现甲基营养型产甲烷菌中存在的独特 F420H2 脱氢酶。
Washed membranes prepared from H2+CO2- or formate-grown cells ofMethanococcus voltaecatalyzed the oxidation of coenzyme F420H2and the reduction of the heterodisulfide (CoB–S–S–CoM) of 2-mercaptoethanesulfonate and 7-mercaptoheptanoylthreonine phosphate, which is the terminal electron acceptor of the methanogenic pathway. The reaction followed a 1:1 stoichiometry according to the equation: F420H2+ COB–S–S–CoM → F420+ CoM–SH + CoB–SH. These findings indicate that the reaction depends on a membrane-bound F420H2-oxidizing enzyme and on the heterodisulfide reductase, which remains partly membrane-bound after cell lysis. To elucidate the nature of the F420H2-oxidizing protein, washed membranes were solubilized with detergent, and the enzyme was purified by sucrose density centrifugation, anion-exchange chromatography, and gel filtration. Several lines of evidence indicate that F420H2oxidation is catalyzed by a membrane-associated F420-reducing hydrogenase. The purified protein catalyzed the H2-dependent reduction of methyl viologen and F420. The apparent molecular mass and the subunit composition (43, 37, and 27 kDa) are almost identical to those of the F420-reducing hydrogenase that has already been purified fromMc. voltae. Moreover, the N-terminus of the 37-kDa subunit is identical to the amino acid sequence deduced from thefruGgene of the operon encoding the selenium-containing F420-reducing hydrogenase fromMc. voltae. A distinct F420H2dehydrogenase, which is present in methylotrophic methanogens, was not found in this organism.