Functional analysis based on the structure of DNA replication proteins in Escherichia coli

根据该结构，我们检查了与 DNA 复制起始相关的蛋白质的功能。首先，我们检查了 DnaA 的 N 端结构域。 DnaA 蛋白是大肠杆菌中启动染色体复制的关键蛋白。大肠杆菌 DnaA 的结构分为四个功能域。尽管结构域 III 和 IV 的结构已经确定，但含有 DnaA 寡聚活性和 DnaB 解旋酶结合位点的 N 端结构域的结构尚不清楚。为了确定N-末端结构域的结构，我们分配了Dna A的N-末端结构域的^1H、^13C和^15N主链共振(Bio NMR分配2007)。此外，我们使用核磁共振波谱方法确定了 N 端结构域 (1-108) 结构。结构域 I 具有 α-α-β-β-α-β 基序，与 K 同源 (KH) 结构域相似，对 oriC 单链 DNA 具有弱亲和力，与 KH 结构域功能一致。携带 Trp-6 的疏水表面最有可能形成结构域 I 二聚化的界面。 Glu-21 位于结构域 I 与 Trp-6 位点相对的表面，对于 DnaB 解旋酶加载至关重要。这些发现提出了 DnaA 同多聚体形成和 oriC 上 DnaB 解旋酶负载的模型（J. Biol. Chem. 2007）。该论文被 J. Biol 选为本周论文。化学。 2007 年，6 月 15 日。然后，我们检查了与 DnaA 降解相关的热休克蛋白 HSPQ、细胞分裂重新激活剂 CedA 和引发体成分 PriB 的蛋白质功能和结构分析。其中部分成果在多次学术会议上公布。

Based on the structure, we examined the functions of proteins which were associated with DNA replication initiation. Firstly, we examined the N-terminal domain of DnaA. DnaA protein is a key protein in the initiation of chromosomal replication in Escherichia coli. The structure of E. coli DnaA is subdivided into four functional domains. Although the structure of domain III and IV were already determined, the structure of N-terminal domain, which contains DnaA oligomerization activity and DnaB helicase binding sites, was not understood. To determined the structure of N-terminal domain, we assigned the ^1H,^<13>C and ^<15>N backbone resonances of N-terminal domain of Dna A (Bio NMR assignment 2007). Furthermore, we determined the N-terminal domains (1-108) structure using NMR spectroscopic method. Domain I has an α-α-β-β-α-β motif, similar to that of the K homology (KH) domain and has weak affinity for oriC single-stranded DNA, consistent with KH domain function. A hydrophobic surface carrying Trp-6 most likely forms the interface for domain I dimerization. Glu-21 is located on the opposite surface of domain I from the Trp-6 site and is crucial for DnaB helicase loading. These findings suggested a model for DnaA homomultimer formation and DnaB helicase loading on oriC (J. Biol. Chem. 2007) . This paper was selected as Papers of the Week in J. Biol. Chem. 2007, June 15. And then, we examined the protein functional and structural analyses of heat shock protein HSPQ associated with DnaA degradation, cell division re-activator CedA, and the primosome component PriB. Some part of these results was announced at the several academic conferences.