Böhm R, Sauter M, Böck A
Lehrstuhl für Mikrobiologie, Universität München, FRG.
Mol Microbiol. 1990 Feb;4(2):231-43. doi: 10.1111/j.1365-2958.1990.tb00590.x.
An 8kb segment of DNA from the 58/59 min region of the E. coli chromosome, which complements the defect of a mutant devoid of hydrogenase 3 activity, has been sequenced. Eight open reading frames were identified which are arranged in a transcriptional unit; all open reading frames were transcribed and translated in vivo in a T7 promoter/polymerase system. Analysis of the amino acid sequences derived from the nucleic acid sequences revealed that one of them, open reading frame 5 (ORF5), exhibits significant sequence similarity to conserved regions of the large subunit from Ni/Fe hydrogenases. Two of the open reading frames (orf2, orf6) code for proteins apparently carrying iron-sulphur clusters of the 4Fe/4S ferredoxin type. The product of one of the open reading frames, orf7, displays extensive sequence similarity with protein G from the chloroplast electron transport chain. ORF3 and ORF4, on the other hand, are extremely hydrophobic proteins with nine and six putative transmembrane helices, respectively. Over a limited hydrophilic sequence stretch, bordered by putative transmembrane areas, ORF3 and ORF4 exhibit homology with subunits 4 and 1 of mitochondrial and plastid NADH-ubiquinol oxidoreductases, respectively. The operon described, therefore, appears to comprise genes for redox carriers linking formate oxidation to proton reduction and for a hydrogenase of hitherto unique composition.
对大肠杆菌染色体58/59分钟区域的一段8kb DNA片段进行了测序，该片段可弥补缺乏氢化酶3活性的突变体的缺陷。鉴定出八个开放阅读框，它们排列在一个转录单元中；所有开放阅读框在T7启动子/聚合酶系统中均在体内进行转录和翻译。对从核酸序列推导的氨基酸序列的分析表明，其中一个开放阅读框5（ORF5）与Ni/Fe氢化酶大亚基的保守区域具有显著的序列相似性。其中两个开放阅读框（orf2、orf6）编码的蛋白质显然携带4Fe/4S铁氧化还原蛋白类型的铁硫簇。其中一个开放阅读框orf7的产物与叶绿体电子传递链中的蛋白质G具有广泛的序列相似性。另一方面，ORF3和ORF4是极具疏水性的蛋白质，分别具有九个和六个推定的跨膜螺旋。在由推定的跨膜区域界定的有限亲水序列片段上，ORF3和ORF4分别与线粒体和质体NADH-泛醌氧化还原酶的亚基4和亚基1具有同源性。因此，所述操纵子似乎包含将甲酸氧化与质子还原联系起来的氧化还原载体基因以及一种迄今组成独特的氢化酶基因。
