Chan Kun-Wei, Lee Yi-Juan, Wang Chia-Hung, Huang Haimei, Sun Yuh-Ju
Institute of Bioinformatics and Structural Biology, National Tsing Hua University, Hsinchu 300, Taiwan, ROC.
J Mol Biol. 2009 May 8;388(3):508-19. doi: 10.1016/j.jmb.2009.03.022. Epub 2009 Mar 13.
Single-stranded DNA (ssDNA)-binding protein (SSB) plays an important role in DNA replication, recombination, and repair. SSB consists of an N-terminal ssDNA-binding domain with an oligonucleotide/oligosaccharide binding fold and a flexible C-terminal tail involved in protein-protein interactions. SSB from Helicobacter pylori (HpSSB) was isolated, and the ssDNA-binding characteristics of HpSSB were analyzed by fluorescence titration and electrophoretic mobility shift assay. Tryptophan fluorescence quenching was measured as 61%, and the calculated cooperative affinity was 5.4x10(7) M(-1) with an ssDNA-binding length of 25-30 nt. The crystal structure of the C-terminally truncated protein (HpSSBc) in complex with 35-mer ssDNA [HpSSBc-(dT)(35)] was determined at a resolution of 2.3 A. The HpSSBc monomer folds as an oligonucleotide/oligosaccharide binding fold with a Y-shaped conformation. The ssDNA wrapped around the HpSSBc tetramer through a continuous binding path comprising five essential aromatic residues and a positively charged surface formed by numerous basic residues.
单链DNA（ssDNA）结合蛋白（SSB）在DNA复制、重组及修复过程中发挥着重要作用。SSB由一个具有寡核苷酸/寡糖结合折叠的N端ssDNA结合结构域和一个参与蛋白质-蛋白质相互作用的柔性C端尾巴组成。分离出幽门螺杆菌的SSB（HpSSB），并通过荧光滴定和电泳迁移率变动分析对HpSSB的ssDNA结合特性进行了分析。测得色氨酸荧光猝灭率为61%，计算得到的协同亲和力为5.4×10⁷ M⁻¹，ssDNA结合长度为25 - 30 nt。确定了与35聚体ssDNA [HpSSBc-(dT)₃₅] 复合的C端截短蛋白（HpSSBc）的晶体结构，分辨率为2.3 Å。HpSSBc单体折叠成具有Y形构象的寡核苷酸/寡糖结合折叠。ssDNA通过由五个必需芳香族残基和由众多碱性残基形成的带正电表面组成的连续结合路径缠绕在HpSSBc四聚体周围。
