Hsu J, Bramhill D, Thompson C M
Department of Biochemistry, Merck Research Laboratories, Rahway, New Jersey 07065.
Mol Microbiol. 1994 Mar;11(5):903-11. doi: 10.1111/j.1365-2958.1994.tb00369.x.
The 245 bp chromosomal origin, oriC, of Escherichia coli contains two iterated motifs. Three 13-mers tandemly repeated at one end of the origin and four 9-mers in a nearby segment of oriC are highly conserved in enteric bacteria, as is the distance separating these two sequence clusters. Mutant origins were constructed with altered spacing of the 9-mers relative to the 13-mers. Loss or addition of even a single base drastically reduced replication, both in vivo and in vitro. Spacing mutant origins bound effectively to DnaA protein but failed to support efficient open complex formation. These results suggest that interaction with the 9-mers positions at least one subunit of DnaA to recognize directly the nearest 13-mer for DNA melting.
大肠杆菌245碱基对的染色体复制起点oriC包含两个重复基序。在该起点一端串联重复的三个13聚体以及oriC附近片段中的四个9聚体在肠道细菌中高度保守，这两个序列簇之间的距离也是如此。构建了9聚体相对于13聚体间距改变的突变复制起点。即使只缺失或添加一个碱基，都会在体内和体外极大地降低复制效率。间距突变的复制起点能有效结合DnaA蛋白，但无法支持高效的开放复合物形成。这些结果表明，与9聚体的相互作用使DnaA的至少一个亚基定位，从而直接识别最近的13聚体以进行DNA解链。
