Protein roadblocks and helix discontinuities are barriers to the initiation of mismatch repair
Protein roadblocks and helix discontinuities are barriers to the initiation of mismatch repair
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DOI：
10.1073/pnas.0705129104
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发表时间：
2007-07-31
影响因子：
11.1
通讯作者：
Modrich, Paul
中科院分区：
文献类型：
--
作者：
Pluciennik, Anna;Modrich, Paul
指导大肠杆菌错配修复的半甲基化d（GATC）序列可以位于错配的任一侧，间隔距离为1，000 bp或更大。修复的起始涉及MutH内切核酸酶的错配、MutS和MutL依赖性激活，其在d（GATC）序列处切割未甲基化的链，随后的链断裂用作适当的3 '至5 '或5 '至3 '切除系统的加载位点。然而，负责协调识别的错配和半修饰的d（GATC）网站的机制是不确定的。我们表明，蛋白质路障（EcoRl（E111 Q），EcoRl内切酶的水解缺陷形式）放置在两个DNA位点之间的螺旋上抑制MutH激活70-80%，并且逃脱抑制的事件至少部分归因于EcoRl（E111 Q）远离其识别位点的扩散。我们还表明，位于较短的路径连接的错配和d（GATC）在一个环状异源双链位点的双链断裂废除MutH激活，而在较长的路径内的双链断裂是没有效果的。这些发现支持了这样的观点，即错配修复的启动涉及沿着螺旋轮廓的信号传导。
The hemimethylated d(GATC) sequence that directs Escherichia coli mismatch repair can reside on either side of a mismatch at a separation distance of 1,000 bp or more. Initiation of repair involves the mismatch-, MutS-, and MutL-dependent activation of MutH endonuclease, which incises the unmethylated strand at the d(GATC) sequence, with the ensuing strand break serving as the loading site for the appropriate 3 '-to-5 ' or 5 '-to-3 ' excision system. However, the mechanism responsible for the coordinated recognition of the mismatch and a hemimodified d(GATC) site is uncertain. We show that a protein roadblock (EcoRl(E111Q), a hydrolytically defective form of EcoRl enclonuclease) placed on the helix between the two DNA sites inhibits MutH activation by 70-80% and that events that escape inhibition are attributable, at least in part, to diffusion of EcoRl(E111Q) away from its recognition site. We also demonstrate that a double-strand break located within the shorter path linking the mismatch and a d(GATC) site in a circular heteroduplex abolishes MutH activation, whereas a double-strand break within the longer path is without effect. These findings support the idea that initiation of mismatch repair involves signaling along the helix contour.