Biochemical and genetic study of the regulatory mechanism of the chromosomal DNA replication in Escherichia coli

大肠杆菌染色体DNA复制调控机制的生化和遗传学研究

基本信息

批准号：
06454600
负责人：
SEKIMIZU Kazuhisa
金额：
$ 4.29万
依托单位：
KYUSHU UNIVERSITY
依托单位国家：
日本
项目类别：
Grant-in-Aid for General Scientific Research (B)
财政年份：
1994
资助国家：
日本
起止时间：
1994 至 1995
项目状态：
已结题

来源：
https://kaken.nii.ac.jp/en/grant/KAKENHI-PROJECT-06454600/
关键词：
DnaA protein DNA replication chromosomal DNA Escherichia coli.ATP-binding phospholipid DNA topoisomerase cell cycle oriC DNAポリメラーゼ脂質 dnaA変異株 ATPピリドキサール

项目摘要

DnaA protein is the initiator of chromosomal DNA replication in Escherichia coli. The head investigator and colleagues showed that DnaA protein is activated by acidic phospholipids including cardiolipin. In this study, we examined the influcnce of mixed liposomes composed of two different synthetic lipids on the ATP-binding of DnaA protein. The result showed that the cluster structure of acidic lipid is necessary for the interaction of acidic lipid in the mixed membrane with DnaA protein. This finding suggests a new insight for the regulation of the activity of DnaA protein, the initiator protein of DNA replication, in the cycle of cell division. Namely, acidic phospholipids may form either the cluster of dispersd structures in cell membrane, and that structural alternation of lipid structure may be coupled with cell cycle and it activates DnaA protein resulting the initiation of DNA replication.Further, we first reported the influence of DnaA protein on the supercoiled structure of double stranded DNA.We showed that DNA is more negatively supercoiled when the reaction of DNA topoisomerase was carried out in the presence of DnaA protein. This activity of DnaA protein was not dependent on sequence of DNA and was specifically inhibited by binding of DnaA protein to adenine nucleotide.

DnaA 蛋白是大肠杆菌中染色体 DNA 复制的起始因子。首席研究员和同事表明，DnaA 蛋白可被包括心磷脂在内的酸性磷脂激活。在本研究中，我们研究了由两种不同合成脂质组成的混合脂质体对 DnaA 蛋白 ATP 结合的影响。结果表明，酸性脂质的簇结构是混合膜中酸性脂质与DnaA蛋白相互作用所必需的。这一发现为细胞分裂周期中 DNA 复制的起始蛋白 DnaA 蛋白的活性调节提供了新的见解。也就是说，酸性磷脂可以在细胞膜上形成分散结构簇，并且脂质结构的结构改变可能与细胞周期耦合并激活DnaA蛋白，从而启动DNA复制。此外，我们首次报道了DnaA的影响我们发现，当 DNA 拓扑异构酶的反应在 DnaA 蛋白存在的情况下进行时，DNA 的超螺旋结构更加负。 DnaA 蛋白的这种活性不依赖于 DNA 序列，并且通过 DnaA 蛋白与腺嘌呤核苷酸的结合而被特异性抑制。