Lee J H, Patel P, Sankar P, Shanmugam K T
J Bacteriol. 1985 Apr;162(1):344-52. doi: 10.1128/jb.162.1.344-352.1985.
A positive selection procedure is described for the isolation of hydrogenase-defective mutant strains of Escherichia coli. Mutant strains isolated by this procedure can be divided into two major classes. Class I mutants produced hydrogenase activity (determined by using a tritium-exchange assay) and formate hydrogenlyase activity but lacked the ability to reduce benzyl viologen or fumarate with H2 as the electron donor. Class II mutants failed to produce active hydrogenase and hydrogenase-dependent activities. All the mutant strains produced detectable levels of formate dehydrogenase-1 and -2 and fumarate reductase. The mutation in class I mutants mapped near 65 min of the E. coli chromosome, whereas the mutation in class II mutants mapped between srl and cys operons (58 and 59 min, respectively) in the genome. The class II Hyd mutants can be further subdivided into two groups (hydA and hydB) based on the cotransduction characteristics with cys and srl. These results indicate that there are two hyd operons and one hup operon in the E. coli chromosome. The two hyd operons are needed for the production of active hydrogenase, and all three are essential for hydrogen-dependent growth of the cell.
描述了一种用于分离大肠杆菌氢化酶缺陷突变菌株的阳性选择程序。通过该程序分离出的突变菌株可分为两大类。I类突变体产生氢化酶活性（通过氚交换测定法确定）和甲酸氢化酶活性，但缺乏以H2作为电子供体还原苄基紫精或富马酸酯的能力。II类突变体未能产生活性氢化酶和依赖氢化酶的活性。所有突变菌株都产生了可检测水平的甲酸脱氢酶-1和-2以及富马酸还原酶。I类突变体中的突变位于大肠杆菌染色体65分钟附近，而II类突变体中的突变位于基因组中的srl和cys操纵子之间（分别为58和59分钟）。基于与cys和srl的共转导特性，II类Hyd突变体可进一步细分为两组（hydA和hydB）。这些结果表明，大肠杆菌染色体中有两个hyd操纵子和一个hup操纵子。这两个hyd操纵子是产生活性氢化酶所必需的，并且所有三个操纵子对于细胞的氢依赖生长都是必不可少的。
