A mutation in helicase motif III of E. coli RecG protein abolishes branch migration of Holliday junctions.
A mutation in helicase motif III of E. coli RecG protein abolishes branch migration of Holliday junctions.
复制标题
大肠杆菌 RecG 蛋白解旋酶基序 III 的突变消除了霍利迪连接的分支迁移。
DOI：
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发表时间：
1994
影响因子：
14.9
通讯作者：
Robert G. Lloyd
中科院分区：
文献类型：
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作者：
G. Sharples;M. Whitby;L. Ryder;Robert G. Lloyd
大肠杆菌的 RecG 蛋白催化 RecA 产生的霍利迪连接的分支迁移，并在需要 ATP 水解的反应中将合成的 X 连接解离成双链体产物。为了研究该酶的作用模式，对使 recG (recG162) 失活的染色体突变进行了克隆和测序。 recG162 突变是 G:C 到 A:T 的转变，会在蛋白质中产生 Ala428 到 Val 的取代。这种变化影响蛋白质中的基序（基序 III），该基序在 DNA 和 RNA 解旋酶中高度保守。 RecG162 蛋白经过纯化并显示保留了结合合成 X 和 Y 连接的能力。然而，它不会解离这些连接，并且无法催化从 RecA 链交换反应纯化的霍利迪连接中间体的分支迁移。 RecG162 保留了 DNA 依赖性 ATP 酶活性，但相对于野生型蛋白，该活性大大降低，特别是在单链 DNA 作为辅助因子的情况下。这些结果表明 RecG 的分支迁移与靶向连接的 DNA 解旋酶活性有关。
The RecG protein of Escherichia coli catalyses branch migration of Holliday junctions made by RecA and dissociates synthetic X junctions into duplex products in reactions that require hydrolysis of ATP. To investigate the mode of action of this enzyme a chromosomal mutation that inactivates recG (recG162) was cloned and sequenced. The recG162 mutation is a G:C to A:T transition, which produces an Ala428 to Val substitution in the protein. This change affects a motif (motif III) in the protein that is highly conserved in DNA and RNA helicases. RecG162 protein was purified and shown to retain the ability to bind synthetic X and Y junctions. However, it does not dissociate these junctions and fails to catalyse branch migration of Holliday junction intermediates purified from a RecA strand exchange reaction. RecG162 retains a DNA-dependent ATPase activity, but this is much reduced relative to the wild-type protein, especially with single-stranded DNA as a co-factor. These results suggest that branch migration by RecG is related to a junction-targeted DNA helicase activity.