Sutton M D, Carr K M, Vicente M, Kaguni J M
Department of Biochemistry, Michigan State University, East Lansing, Michigan 48824-1319, USA.
J Biol Chem. 1998 Dec 18;273(51):34255-62. doi: 10.1074/jbc.273.51.34255.
Initiation of DNA replication at the Escherichia coli chromosomal origin occurs through an ordered series of events that depends first on the binding of DnaA protein, the replication initiator, to DnaA box sequences followed by unwinding of an AT-rich region. A step that follows is the binding of DnaB helicase at oriC so that it is properly positioned at each replication fork. We show that DnaA protein actively mediates the entry of DnaB at oriC. One region (amino acids 111-148) transiently binds to DnaB as determined by surface plasmon resonance. A second functional domain, possibly involving formation of a unique nucleoprotein structure, promotes the stable binding of DnaB during the initiation process and is inactivated in forming an intermediate termed the prepriming complex by removal of the N-terminal 62 residues. Based on similarities in the replication process between prokaryotes and eukaryotes, these results suggest that a similar mechanism may load the eukaryotic replicative helicase.
大肠杆菌染色体复制起点处的DNA复制起始是通过一系列有序事件发生的，这些事件首先依赖于复制起始因子DnaA蛋白与DnaA框序列的结合，随后富含AT的区域解旋。接下来的一个步骤是DnaB解旋酶在oriC处结合，使其正确定位在每个复制叉处。我们发现DnaA蛋白积极介导DnaB在oriC处的进入。通过表面等离子体共振测定，一个区域（氨基酸111 - 148）与DnaB短暂结合。第二个功能域可能涉及形成独特的核蛋白结构，在起始过程中促进DnaB的稳定结合，并通过去除N端62个残基在形成称为预引发复合物的中间体时失活。基于原核生物和真核生物复制过程的相似性，这些结果表明可能存在类似机制来加载真核生物复制解旋酶。
