The Phe-X-Glu DNA binding motif of MutS -: The role of hydrogen bonding in mismatch recognition
The Phe-X-Glu DNA binding motif of MutS -: The role of hydrogen bonding in mismatch recognition
复制标题
DOI：
10.1074/jbc.c100449200
复制
发表时间：
2001-12-07
影响因子：
4.8
通讯作者：
Hsieh, P
中科院分区：
文献类型：
--
作者：
Schofield, MJ;Brownewell, FE;Hsieh, P
来自水生栖热菌和大肠杆菌的MutS蛋白与含有错配的DNA双链体的复合物的晶体结构显示，保守的Phe-X-Glu基序中的Glu残基参与与未配对的胸苷或G-T碱基-碱基错配的胸苷的氢键接触。在这里，氢键在错配识别MutS的作用进行了评估。MutS对分别含有A和T的非极性形状模拟物、4-甲基苯并咪唑（Z）和二氟甲苯（F）的DNA双链体（其缺乏氢键供体和受体）的相对亲和力在凝胶迁移率变动测定中测定。结果提供了支持的诱导配合模式的错配结合，其中双链体不稳定的错配是首选的基板扭结MutS。O是Glu的2基团的一个元素，错配碱基之间的氢键对错配识别的贡献很小，并且远不如具有保守的Phe残基的芳环堆叠重要。其中Ala取代Glu（38）的MutS蛋白显示出在体内不具有错配修复。DNA结合的研究揭示了一个新的作用，保守的Glu残基在建立错配歧视MutS。
The crystal structures of MutS protein from Thermus aquaticus and Escherichia coli in a complex with a mismatch-containing DNA duplex reveal that the Glu residue in a conserved Phe-X-Glu motif participates in a hydrogen-bonded contact with either an unpaired thymidine or the thymidine of a G-T base-base mismatch. Here, the role of hydrogen bonding in mismatch recognition by MutS is assessed. The relative affinities of MutS for DNA duplexes containing nonpolar shape mimics of A and T, 4-methylbenzimidazole (Z), and difluoro-toluene (F), respectively, that lack hydrogen bonding donors and acceptors, are determined in gel mobility shift assays. The results provide support for an induced fit mode of mismatch binding in which duplexes destabilized by mismatches are preferred substrates for kinking by MutS. Hydrogen bonding between the O is an element of2 group of Glu and the mismatched base contributes only marginally to mismatch recognition and is significantly less important than the aromatic ring stack with the conserved Phe residue. A MutS protein in which Ala is substituted for Glu(38) is shown to be defective for mismatch repair in vivo. DNA binding studies reveal a novel role for the conserved Glu residue in the establishment of mismatch discrimination by MutS.