Wiggs J L, Gilman M Z, Chamberlin M J
Proc Natl Acad Sci U S A. 1981 May;78(5):2762-6. doi: 10.1073/pnas.78.5.2762.
Preparations of Bacillus subtilis RNA polymerase (nucleosidetriphosphate:RNA nucleotidyltransferase, EC 2.7.7.6) from vegetatively growing cells contain small amounts of an activity (B. subtilis RNA polymerase holoenzyme II) that shows a unique promoter specificity with T7 bacteriophage DNA as compared with the normal B. subtilis holoenzyme (holoenzyme I) and lacks the normal sigma subunit [Jaehning, J. A., Wiggs, J. L. & Chamberlin, M. J. (1979) Proc. Natl. Acad. Sci. USA 76, 5470-5474]. By heparin-agarose chromatography we have obtained holoenzyme II fractions that have no detectable holoenzyme I activity as judged by their failure to utilize promoter sites for holoenzyme I on any template we have tested. These fractions are far more active with B. subtilis DNA than with T7 DNA or other heterologous templates. This high degree of specificity has allowed identification of plasmids containing cloned fragments of B. subtilis DNA that bear strong promoter sites for holoenzyme II. These promoter sites are not used at all by B. subtilis RNA polymerase holoenzyme I. The specificity of holoenzyme II is dictated by a peptide of Mr 28,000 as judged by copurification of the peptide with specific holoenzyme II activity and by reconstitution of the holoenzyme II promoter specificity when the isolated peptide is added to B. subtilis core polymerase. Hence the 28,000 Mr peptide appears to be a sigma factor that determines a promoter specificity distinct from that of RNA polymerase holoenzyme I and all other known bacterial RNA polymerases.
从营养生长细胞中制备的枯草芽孢杆菌RNA聚合酶（核苷三磷酸：RNA核苷酸转移酶，EC 2.7.7.6）含有少量活性物质（枯草芽孢杆菌RNA聚合酶全酶II），与正常的枯草芽孢杆菌全酶（全酶I）相比，它对T7噬菌体DNA表现出独特的启动子特异性，并且缺乏正常的σ亚基[Jaehning, J. A., Wiggs, J. L. & Chamberlin, M. J. (1979) Proc. Natl. Acad. Sci. USA 76, 5470 - 5474]。通过肝素-琼脂糖层析，我们获得了全酶II组分，根据它们在我们测试的任何模板上都不能利用全酶I的启动子位点这一情况判断，这些组分没有可检测到的全酶I活性。这些组分对枯草芽孢杆菌DNA的活性比对T7 DNA或其他异源模板的活性高得多。这种高度的特异性使得能够鉴定出含有枯草芽孢杆菌DNA克隆片段的质粒，这些片段带有全酶II的强启动子位点。枯草芽孢杆菌RNA聚合酶全酶I根本不使用这些启动子位点。根据该肽与全酶II特异性活性的共纯化以及将分离的肽添加到枯草芽孢杆菌核心聚合酶中时全酶II启动子特异性的重建判断，全酶II的特异性由一个分子量为28,000的肽决定。因此，分子量为28,000的肽似乎是一种σ因子，它决定了一种与RNA聚合酶全酶I和所有其他已知细菌RNA聚合酶不同的启动子特异性。
