Interaction between the replicational initiator DnaA and the DNA polymerase components

复制起始子 DnaA 和 DNA 聚合酶组分之间的相互作用

基本信息

批准号：
10680616
负责人：
KATAYAMA Tsutomu
金额：
$ 2.18万
依托单位：
KYUSHU UNIVERSITY
依托单位国家：
日本
项目类别：
Grant-in-Aid for Scientific Research (C)
财政年份：
1998
资助国家：
日本
起止时间：
1998 至 1999
项目状态：
已结题

来源：
https://kaken.nii.ac.jp/grant/KAKENHI-PROJECT-10680616/
关键词：
chromosome DNA replication ATP DnaA DNA polymerase Sliding clamp Replicational initiation Cell cycle スライディング・クランプ染色体複製 otiC 大腸菌

项目摘要

In E. coli, DnaA protein forms a complex with the chromosomal replication origin (oriC). In this complex, DnaA opens the duplex DNA. This initiation activity of DnaA protein depends on ATP binding. On the generated single stranded region, DNA polymerase III, the replicase of chromosome, is loaded.We recently revealed that the hydrolysis of ATP bound to DnaA is accelerated by interaction with the sliding clamp of DNA polymerase III and a partially purified IdaB protein fraction. The sliding clamp is a ring-like configuration that is comprised from a dimer of the β subunit of the replicase and tethers DNA strand to the replicase. By this reaction, the ADP form of DnaA, an inactive form for initiation, is rapidly generated. We named this system RIDA for Regulatory Inactivation of DnaA, and suggested that RIDA is a main system to prevent extra initiations for maintaining the frequency of initiation only once per chromosome per cell cycle. Indeed, certain DnaA mutant proteins that are insensitive to a RIDA causes overinitiation of chromosomal replication. Moreover, we had in vivo evidence that RIDA depends on genes necessary for DNA replication. Content of the ATP form of DnaA in randomly dividing cells is 15 - 30% of total ATP/ADP-bound DnaA molecules, and when genes required for DNA replication were inactivated, the ATP-DnaA level increased up to 80%. These results suggest that the ATP-DnaA level is restrained during the cell cycle depending on DNA replication, which may be a key event to control the initiation cycle of chromosomal replication.

在大肠杆菌中，DnaA 蛋白与染色体复制起点 (oriC) 形成复合物，在该复合物中，DnaA 蛋白的起始活性取决于生成的单链区域 DNA 聚合酶 III。加载染色体的复制酶。我们最近发现，通过与 DNA 聚合酶 III 的滑动夹和部分纯化的 IdaB 蛋白组分的相互作用，可以加速与 DnaA 结合的 ATP 的水解。夹子是一种环状结构，由复制酶的 β 亚基二聚体组成，并将 DNA 链束缚在复制酶上。通过该反应，快速生成 DnaA 的 ADP 形式（一种起始非活性形式）。该系统 RIDA 用于 DnaA 的调节失活，并表明 RIDA 是防止额外启动的主要系统，以维持每个细胞周期每条染色体仅一次的启动频率。对 RIDA 不敏感会导致染色体复制过度启动。此外，我们有体内证据表明 RIDA 依赖于 DNA 复制所需的基因。随机分裂的细胞中 ATP 形式的 DnaA 含量占 ATP/ADP 结合总量的 15 - 30%。 DnaA 分子，当 DNA 复制所需的基因失活时，ATP-DnaA 水平增加高达 80%。这些结果表明 ATP-DnaA 水平细胞周期中的抑制取决于DNA复制，这可能是控制染色体复制起始周期的关键事件。