Steigerwald V J, Pihl T D, Reeve J N
Department of Microbiology, Ohio State University, Columbus 43210.
Proc Natl Acad Sci U S A. 1992 Aug 1;89(15):6929-33. doi: 10.1073/pnas.89.15.6929.
Sequencing the genes encoding the methyl viologen-reducing hydrogenase, cloned from Methanobacterium thermoautotrophicum strain delta H and Methanothermus fervidus, revealed the presence of tightly linked genes, designated mvhB, which were predicted to encode proteins containing six tandemly arranged bacterial ferrodoxin-like domains. A lacZ-mvhB gene fusion has been constructed and expressed in Escherichia coli. Rabbit antibodies raised against the fusion polypeptide purified from E. coli have been used to identify and isolate the polyferrodoxin from Mb. thermoautotrophicum strain delta H. The polyferredoxin accumulates in cells of the methanogen during exponential growth but decreases rapidly on entry into stationary phase. It is not processed into monoferredoxins and is located primarily in the soluble fraction of cell lysates of Mb. thermoautotrophicum. Metronidazole reduction by crude extracts of Mb. thermoautotrophicum strain delta H cells, dependent on the presence of hydrogen and the heterodisulfide CoM-S-S-HTP [formed from the two coenzymes 2-mercaptoethanesulfonic acid (coenzyme M, HS-CoM) and N-(7-mercaptoheptanoyl)threonine O3-phosphate (HS-HTP)], was not inhibited by the antibodies raised against the LacZ-MvhB fusion polypeptide.
对从嗜热自养甲烷杆菌δH菌株和嗜热甲烷热菌中克隆的编码甲基紫精还原氢化酶的基因进行测序，结果显示存在紧密连锁的基因，命名为mvhB，预计这些基因编码的蛋白质含有六个串联排列的细菌铁氧化还原蛋白样结构域。构建了一个lacZ-mvhB基因融合体并在大肠杆菌中表达。用针对从大肠杆菌中纯化的融合多肽产生的兔抗体来鉴定和分离嗜热自养甲烷杆菌δH菌株中的多铁氧化还原蛋白。多铁氧化还原蛋白在产甲烷菌指数生长期的细胞中积累，但在进入稳定期后迅速减少。它不会被加工成单铁氧化还原蛋白，主要位于嗜热自养甲烷杆菌细胞裂解物的可溶部分。嗜热自养甲烷杆菌δH菌株细胞的粗提物还原甲硝唑，这依赖于氢气和异二硫化物CoM-S-S-HTP[由两种辅酶2-巯基乙烷磺酸（辅酶M，HS-CoM）和N-（7-巯基庚酰基）苏氨酸O3-磷酸（HS-HTP）形成]的存在，不受针对LacZ-MvhB融合多肽产生的抗体抑制。
