Fujita M, Amemura A
Department of Biotechnology, Faculty of Engineering, Fukuyama University, Hiroshima, Japan.
Biosci Biotechnol Biochem. 1992 Nov;56(11):1797-800. doi: 10.1271/bbb.56.1797.
DNA-dependent RNA polymerase (EC 2.7.7.6) was purified from Pseudomonas putida. The enzyme had the typical composition of beta',beta,alpha, and sigma subunits of eubacterial RNA polymerases. The molecular masses of the subunits were 156,000 Da, 151,000 Da, 87,000 Da, and 42,000 Da, respectively, as measured by polyacrylamide gel electrophoresis in the presence of sodium dodecyl sulfate. The NH2-terminal amino acid residues of the alpha subunit had a marked homology with those of the alpha subunit of Escherichia coli RNA polymerase. The enzyme activity was dependent on ribonucleoside triphosphates, Mg2+, and a DNA template, and was inhibited in vitro by rifampicin. The enzyme activity was maximal in the presence of 10 mM MgCl2. In an in vitro transcription assay using the tac promoter-controlled DNA as a template, the RNA polymerase of P. putida initiated transcription at the same site as that of E. coli.
从恶臭假单胞菌中纯化出了依赖DNA的RNA聚合酶（EC 2.7.7.6）。该酶具有真细菌RNA聚合酶β′、β、α和σ亚基的典型组成。通过在十二烷基硫酸钠存在下进行聚丙烯酰胺凝胶电泳测定，各亚基的分子量分别为156,000道尔顿、151,000道尔顿、87,000道尔顿和42,000道尔顿。α亚基的NH2末端氨基酸残基与大肠杆菌RNA聚合酶的α亚基具有显著的同源性。酶活性依赖于核糖核苷三磷酸、Mg2+和DNA模板，并且在体外被利福平抑制。在10 mM MgCl2存在下酶活性最高。在以tac启动子控制的DNA为模板的体外转录试验中，恶臭假单胞菌的RNA聚合酶与大肠杆菌的RNA聚合酶在相同位点起始转录。
