A prominent β-hairpin structure in the winged-helix domain of RECQ1 is required for DNA unwinding and oligomer formation
A prominent β-hairpin structure in the winged-helix domain of RECQ1 is required for DNA unwinding and oligomer formation
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DOI：
10.1093/nar/gkq1031
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发表时间：
2011-03-01
影响因子：
14.9
通讯作者：
Vindigni, Alessandro
中科院分区：
文献类型：
--
作者：
Lucic, Bojana;Zhang, Ying;Vindigni, Alessandro
由于RecQ解旋酶在抑制基因组不稳定和人类疾病中的作用，近年来引起了人们的极大兴趣。这些非典型的解旋酶通过分解一些高度特异的DNA结构来发挥它们的功能。人RECQ1解旋酶催化核心(RECQ1(49-616))的晶体结构显示一个突出的β-发夹，末端有一个芳香残基(Y564)，位于C末端的翼状螺旋结构域。在这里，我们证明了β-发夹是全长RECQ1的DNA解离和Holliday连接(HJ)分辨活性所必需的，证实了它是五种人RecQ解旋酶不同底物特异性的重要决定因素。此外，我们还发现RECQ1(49-616)中的二聚体形成和全长RECQ1中的四聚体形成都需要β-发夹。我们证实了溶液中存在稳定的RECQ1(49-616)二聚体，并证明了二聚体的形成有利于DNA解离；即使RECQ1单体仍然活跃。相反，四聚体是更专业的活动所必需的，例如HJ拆分和链退火法。有趣的是，四聚体的形成需要两个独立的蛋白质-蛋白质接触，一个涉及RECQ1的β-发夹，另一个涉及RECQ1的N-末端，这表明四聚体组装是一种非分级机制。
RecQ helicases have attracted considerable interest in recent years due to their role in the suppression of genome instability and human diseases. These atypical helicases exert their function by resolving a number of highly specific DNA structures. The crystal structure of a truncated catalytic core of the human RECQ1 helicase (RECQ1(49-616)) shows a prominent beta-hairpin, with an aromatic residue (Y564) at the tip, located in the C-terminal winged-helix domain. Here, we show that the beta-hairpin is required for the DNA unwinding and Holliday junction (HJ) resolution activity of full-length RECQ1, confirming that it represents an important determinant for the distinct substrate specificity of the five human RecQ helicases. In addition, we found that the beta-hairpin is required for dimer formation in RECQ1(49-616) and tetramer formation in full-length RECQ1. We confirmed the presence of stable RECQ1(49-616) dimers in solution and demonstrated that dimer formation favours DNA unwinding; even though RECQ1 monomers are still active. Tetramers are instead necessary for more specialized activities such as HJ resolution and strand annealing. Interestingly, two independent protein-protein contacts are required for tetramer formation, one involves the beta-hairpin and the other the N-terminus of RECQ1, suggesting a non-hierarchical mechanism of tetramer assembly.