Sharma Atul, Nitharwal Ram G, Singh Bhupender, Dar Ashraf, Dasgupta Santanu, Dhar Suman K
Special Centre for Molecular Medicine, Jawaharlal Nehru University, New Delhi, India.
FEBS J. 2009 Jan;276(2):519-31. doi: 10.1111/j.1742-4658.2008.06799.x. Epub 2008 Dec 11.
Helicobacter pylori, an important bacterial pathogen, causes gastric ulcer and gastric adenocarcinoma in humans. The fundamentals of basic biology such as DNA replication are poorly understood in this pathogen. In the present study, we report the cloning and functional characterization of the single-stranded DNA (ssDNA) binding protein from H. pylori. The N-terminal DNA binding domain shows significant homology with E. coli single-stranded DNA binding protein (SSB), whereas the C-terminal domain shows less homology. The overall DNA-binding activity and tetramerization properties, however, remain unaffected. In in vitro experiments with purified proteins, H. pylori (Hp) SSB bound specifically to ssDNA and modulated the enzymatic ATPase and helicase activity of HpDnaB helicase. HpSSB and HpDnaB proteins were co-localized in sharp, distinct foci in exponentially growing H. pylori cells, whereas both were spread over large areas in its dormant coccoid form, suggesting the absence of active replication forks in the latter. These results confirm the multiple roles of SSB during DNA replication and provide evidence for altered replicative metabolism in the spiral and coccoid forms that may be central to the bacterial physiology and pathogenesis.
幽门螺杆菌是一种重要的细菌病原体，可导致人类胃溃疡和胃腺癌。人们对这种病原体的基本生物学原理（如DNA复制）了解甚少。在本研究中，我们报告了幽门螺杆菌单链DNA（ssDNA）结合蛋白的克隆及功能特性。其N端DNA结合结构域与大肠杆菌单链DNA结合蛋白（SSB）具有显著同源性，而C端结构域的同源性较低。然而，其整体DNA结合活性和四聚化特性并未受到影响。在使用纯化蛋白进行的体外实验中，幽门螺杆菌（Hp）SSB特异性结合ssDNA，并调节HpDnaB解旋酶的ATP酶和螺旋酶活性。在指数生长期的幽门螺杆菌细胞中，HpSSB和HpDnaB蛋白共定位在清晰、明显的焦点区域，而在其休眠的球形形式中，两者都分布在大片区域，这表明后者不存在活跃的复制叉。这些结果证实了SSB在DNA复制过程中的多种作用，并为螺旋形和球形形式中复制代谢的改变提供了证据，这可能是细菌生理学和发病机制的核心。
