Manen D, Upegui-Gonzalez L C, Caro L
Department of Molecular Biology, University of Geneva, Switzerland.
Proc Natl Acad Sci U S A. 1992 Oct 1;89(19):8923-7. doi: 10.1073/pnas.89.19.8923.
The replication of plasmid pSC101 requires the plasmid-encoded protein RepA. This protein has a double role: it binds to three directly repeated sequences in the pSC101 origin and promotes replication of the plasmid; it binds to two inversely repeated sequences in its promoter region and regulates its own transcription. A series of RepA protein derivatives carrying deletions of the C-terminal region were assayed for specific binding. We found that the last third of the protein is not needed for binding to the various specific sites. Truncated proteins that still bind can also form heterodimers with a wild-type protein. Analysis of band retardation assays conducted with wild-type and truncated proteins indicates that RepA binds to directly repeated sequences as a monomer and to inversely repeated sequences as a dimer.
质粒pSC101的复制需要质粒编码的蛋白质RepA。这种蛋白质具有双重作用：它与pSC101复制起点的三个直接重复序列结合并促进质粒的复制；它与自身启动子区域的两个反向重复序列结合并调节自身转录。对一系列携带C端区域缺失的RepA蛋白衍生物进行了特异性结合检测。我们发现该蛋白的最后三分之一对于与各种特定位点的结合并非必需。仍能结合的截短蛋白也能与野生型蛋白形成异二聚体。对野生型和截短型蛋白进行的凝胶阻滞分析表明，RepA以单体形式与直接重复序列结合，以二聚体形式与反向重复序列结合。
