Dissection of the functional domains of an archaeal Holliday junction helicase
Dissection of the functional domains of an archaeal Holliday junction helicase
复制标题
古菌霍利迪连接解旋酶功能域的剖析
DOI：
10.1016/j.dnarep.2011.10.009
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发表时间：
2012-02-01
期刊：
影响因子：
3.8
通讯作者：
Shen, Yulong
中科院分区：
文献类型：
--
作者：
Hong, Ye;Chu, Mingzhu;Shen, Yulong
解旋酶和核酸酶形成复合物，在DNA修复途径中发挥非常重要的作用，其中一些在Holliday连接处相互作用。在这项研究中，我们提出了在体外和体内分析的Hjm和它的相互作用与Hjc在硫化叶菌。在体外研究采用Hjm从超嗜热古菌Sulfolobus tokodaii（StoHjm）及其截短的衍生物，和StoHjm蛋白的表征表明，N-末端模块（残基1-431）是能够单独的ATP水解和DNA结合，而C-末端的一个（残基415-704）是负责调节解旋酶活性。StoHjm-StoHjc（Hjc来自S. tokodaii）相互作用被鉴定为StoHjm的结构域II、结构域III（有翼受阻基序）和结构域IV（残基366-645）的一部分。我们提出的证据支持StoHjc通过诱导酶的构象变化来调节StoHjm的解旋酶活性。StoHjm还能抑制Hjc/HJ高复合物的形成，提示StoHjm对Hjc活性的调节机制。我们表明，Hjm是必不可少的细胞活力，使用最近开发的遗传系统和突变体繁殖试验，表明Hjm/Hjc介导的决议停滞的复制叉是至关重要的古细菌。一个试探性的途径，HJM/HJC相互作用可能发生在停滞的复制叉进行了讨论。(C)2011爱思唯尔有限公司版权所有。
Helicases and nucleases form complexes that play very important roles in DNA repair pathways some of which interact with each other at Holliday junctions. In this study, we present in vitro and in vivo analysis of Hjm and its interaction with Hjc in Sulfolobus. In vitro studies employed Hjm from the hyperthermophilic archaeon Sulfolobus tokodaii (StoHjm) and its truncated derivatives, and characterization of the StoHjm proteins revealed that the N-terminal module (residues 1-431) alone was capable of ATP hydrolysis and DNA binding, while the C-terminal one (residues 415-704) was responsible for regulating the helicase activity. The region involved in StoHjm-StoHjc (Hjc from S. tokodaii) interaction was identified as part of domain II, domain III (Winged Helix motif), and domain IV (residues 366-645) for StoHjm. We present evidence supporting that StoHjc regulates the helicase activity of StoHjm by inducing conformation change of the enzyme. Furthermore, StoHjm is able to prevent the formation of Hjc/HJ high complex, suggesting a regulation mechanism of Hjm to the activity of Hjc. We show that Hjm is essential for cell viability using recently developed genetic system and mutant propagation assay, suggesting that Hjm/Hjc mediated resolution of stalled replication forks is of crucial importance in archaea. A tentative pathway with which Hjm/Hjc interaction could have occurred at stalled replication forks is discussed. (C) 2011 Elsevier B.V. All rights reserved.