Shinkawa H, Fujita N, Shiina T, Tanaka K, Takahashi H, Ishihama A, Nimi O
Department of Fermentation Technology, Faculty of Engineering, Hiroshima University.
J Biochem. 1995 Sep;118(3):488-93. doi: 10.1093/oxfordjournals.jbchem.a124934.
RNA polymerase was purified from vegetative-phase mycelia of Streptomyces griseus by a series of ion-exchange chromatographies. By western blot analysis using antiserum against S. coelicolor HrdB, which is a principal sigma factor (sigma(hrdB)), the purified holoenzyme was found to contain sigmaB (=sigma(hrdB)) of S. griseus. Significant amounts of HrdB protein were, however, eluted from the DEAE column at lower concentrations of KCl than that required for for elution of the holoenzyme containing sigmaB, suggesting that sigmaB is dissociated from the core enzyme, or an excess amount of sigmaB exists in S.griseus cells. The holoenzyme containing sigmaB (EsigmaB) transcribed in vitro the dagA promoter of S. coelicolor, and the hardB and hsp70 promoters of S. griseus, suggesting that it is involved in transcription of the essential genes. EsigmaB may be a major form of RNA polymerase holoenzyme in the growing phase of S. griseus.
通过一系列离子交换色谱法从灰色链霉菌的营养期菌丝体中纯化出RNA聚合酶。使用抗天蓝色链霉菌HrdB（一种主要的σ因子（σ(hrdB)））的抗血清进行蛋白质免疫印迹分析，发现纯化的全酶含有灰色链霉菌的σB（=σ(hrdB)）。然而，在比洗脱含σB的全酶所需的KCl浓度更低的情况下，大量的HrdB蛋白从DEAE柱上洗脱下来，这表明σB与核心酶解离，或者在灰色链霉菌细胞中存在过量的σB。含σB的全酶（EσB）在体外转录了天蓝色链霉菌的dagA启动子以及灰色链霉菌的hardB和hsp70启动子，这表明它参与了必需基因的转录。EσB可能是灰色链霉菌生长阶段RNA聚合酶全酶的主要形式。
