Department of Biology, Bldg. 68-530, Massachusetts Institute of Technology, Cambridge, MA 02139, USA.
J Bacteriol. 2011 Feb;193(3):640-8. doi: 10.1128/JB.01253-10. Epub 2010 Nov 19.
The initiation of DNA replication requires the binding of the initiator protein, DnaA, to specific binding sites in the chromosomal origin of replication, oriC. DnaA also binds to many sites around the chromosome, outside oriC, and acts as a transcription factor at several of these. In low-G+C Gram-positive bacteria, the primosomal proteins DnaD and DnaB, in conjunction with loader ATPase DnaI, load the replicative helicase at oriC, and this depends on DnaA. DnaD and DnaB also are required to load the replicative helicase outside oriC during replication restart, independently of DnaA. Using chromatin immunoprecipitation, we found that DnaD and DnaB, but not the replicative helicase, are associated with many of the chromosomal regions bound by DnaA in Bacillus subtilis. This association was dependent on DnaA, and the order of recruitment was the same as that at oriC, but it was independent of a functional oriC and suggests that DnaD and DnaB do not require open complex formation for the stable association with DNA. These secondary binding regions for DnaA could be serving as a reservoir for excess DnaA, DnaD, and DnaB to help properly regulate replication initiation and perhaps are analogous to the proposed function of the datA locus in Escherichia coli. Alternatively, DnaD and DnaB might modulate the activity of DnaA at the secondary binding regions. All three of these proteins are widely conserved and likely have similar functions in a range of organisms.
DNA 复制的起始需要起始蛋白 DnaA 与染色体复制起点 oriC 中的特定结合位点结合。DnaA 还与染色体周围许多 oriC 以外的位点结合，并在其中几个位点作为转录因子发挥作用。在低 GC 革兰氏阳性菌中，原核蛋白 DnaD 和 DnaB 与加载 ATP 酶 DnaI 一起，将复制解旋酶加载到 oriC 上，这取决于 DnaA。DnaD 和 DnaB 还需要在复制重新启动时将复制解旋酶加载到 oriC 以外的地方，这与 DnaA 无关。使用染色质免疫沉淀，我们发现 DnaD 和 DnaB 而不是复制解旋酶与枯草芽孢杆菌中许多与 DnaA 结合的染色体区域相关联。这种关联依赖于 DnaA，招募的顺序与 oriC 相同，但它独立于功能 oriC，表明 DnaD 和 DnaB 不需要开放复合物的形成即可与 DNA 稳定结合。这些 DnaA 的二级结合区域可能充当多余的 DnaA、DnaD 和 DnaB 的储库，以帮助正确调节复制起始，并且可能类似于大肠杆菌中提议的 datA 基因座的功能。或者，DnaD 和 DnaB 可能在二级结合区域调节 DnaA 的活性。这三种蛋白质都广泛保守，可能在多种生物中具有类似的功能。
