Tam J E, Kline B C
Department of Biochemistry and Molecular Biology, Mayo Clinic/Foundation, Rochester, Minnesota 55905.
J Bacteriol. 1989 May;171(5):2353-60. doi: 10.1128/jb.171.5.2353-2360.1989.
The F sex factor plasmid of Escherichia coli contains a pair of genes, ccdA and ccdB, whose protein gene products are involved in an unusual feature of plasmid maintenance. The CcdB protein is a cytotoxin that becomes activated when the F plasmid is lost, thereby killing the F- segregant cells. In F+ cells, the CcdA protein protects against the lethal effects of CcdB. In the present study we show that ccdA and ccdB expressions are negatively autoregulated at the level of transcription. Genetic studies showed that repression required at least ccdB; ccdA alone was without effect, and ccdB alone was not examined because it is lethal. Ccd-operator complexes were purified and contained a mixture of both CcdA and CcdB proteins; however, we could not conclude from our results whether CcdA was necessary for DNA binding or autorepression. By using restriction fragments of the promoter-operator region, we obtained results indicating that at least two DNA-binding sites existed for the Ccd protein(s). Subsequent footprinting of the binding sites showed protection over about a 113-base-pair region encompassing the putative promoter-operator and the beginning of the ccdA gene.
大肠杆菌的F性因子质粒包含一对基因，ccdA和ccdB，其蛋白质基因产物参与质粒维持的一个不同寻常的特性。CcdB蛋白是一种细胞毒素，当F质粒丢失时被激活，从而杀死F - 分离子细胞。在F + 细胞中，CcdA蛋白可防止CcdB的致死作用。在本研究中，我们表明ccdA和ccdB的表达在转录水平上受到负向自动调节。遗传学研究表明，抑制至少需要ccdB；单独的ccdA没有作用，而单独的ccdB由于其致死性未进行检测。Ccd - 操纵子复合物被纯化，并且包含CcdA和CcdB蛋白的混合物；然而，从我们的结果中我们无法得出CcdA对于DNA结合或自动抑制是否必要的结论。通过使用启动子 - 操纵子区域的限制性片段，我们获得的结果表明Ccd蛋白至少存在两个DNA结合位点。随后对结合位点的足迹分析显示在大约113个碱基对的区域受到保护，该区域包含推定的启动子 - 操纵子和ccdA基因的起始部分。
