Reconstitution of the very short patch repair pathway from Escherichia coli.
Reconstitution of the very short patch repair pathway from Escherichia coli.
复制标题
大肠杆菌极短补丁修复途径的重建。
DOI：
10.1074/jbc.m112.384321
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发表时间：
2012
期刊：
影响因子：
--
通讯作者：
Matson,StevenW
中科院分区：
文献类型：
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作者：
Robertson,AdamB;Matson,StevenW
大肠杆菌短斑 (VSP) 修复途径可纠正由 5-甲基胞嘧啶自发水解脱氨损伤导致的胸苷-鸟嘌呤错配。 VSP 修复途径需要 Vsr 核酸内切酶、DNA 聚合酶 I、DNA 连接酶、MutS 和 MutL 才能以最高效率发挥作用。大多数这些蛋白质在 VSP 修复途径中的生化作用已被广泛研究。然而，这些蛋白质尚未在体外系统中的 VSP 修复背景下一起研究。在重构反应中使用 VSP 修复系统的纯化成分，我们已经开始了解这些蛋白质在 VSP 修复途径中所发挥的作用，并深入了解它们的相互作用。在本报告中，我们展示了使用质粒 DNA 底物对 VSP 修复途径进行体外重建。令人惊讶的是，修复轨迹长度可以通过DNA连接酶的浓度来调节。我们提出 MutL 和 MutS 在协调该修复途径中的作用。
TheEscherichia colivery short patch (VSP) repair pathway corrects thymidine-guanine mismatches that result from spontaneous hydrolytic deamination damage of 5-methyl cytosine. The VSP repair pathway requires the Vsr endonuclease, DNA polymerase I, a DNA ligase, MutS, and MutL to function at peak efficiency. The biochemical roles of most of these proteins in the VSP repair pathway have been studied extensively. However, these proteins have not been studied together in the context of VSP repair in anin vitrosystem. Using purified components of the VSP repair system in a reconstitution reaction, we have begun to develop an understanding of the role played by each of these proteins in the VSP repair pathway and have gained insights into their interactions. In this report we demonstrate anin vitroreconstitution of the VSP repair pathway using a plasmid DNA substrate. Surprisingly, the repair track length can be modulated by the concentration of DNA ligase. We propose roles for MutL and MutS in coordination of this repair pathway.