NMR structure and dynamics of the C-terminal domain from human Rev1 and its complex with Rev1 interacting region of DNA polymerase η.
NMR structure and dynamics of the C-terminal domain from human Rev1 and its complex with Rev1 interacting region of DNA polymerase η.
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来自人Rev1的C末端结构域的NMR结构和动力学与DNA聚合酶η的Rev1相互作用区域的复合物。
DOI：
10.1021/bi300566z
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发表时间：
2012-07-10
期刊：
影响因子：
2.9
通讯作者：
Korzhnev DM
中科院分区：
文献类型：
--
作者：
Pozhidaeva A;Pustovalova Y;D'Souza S;Bezsonova I;Walker GC;Korzhnev DM
Rev 1是真核生物中DNA损伤耐受所必需的跨损伤合成（TLS）DNA聚合酶。在TLS停滞的过程中，高保真复制DNA聚合酶被专门的TLS酶暂时取代，这些酶可以绕过DNA损伤位点（病变），从而允许复制继续或复制后的缺口被填补。尽管其催化活性有限，但人Rev 1通过充当支架在TLS中发挥关键作用，所述支架提供Y家族TLS聚合酶polη、pol ι和pol κ进入其同源DNA损伤并促进其随后交换为延伸扭曲的DNA引物-模板的pol。Rev 1与其他主要的人类TLS聚合酶polη、ι、κ和polζ的调节亚基Rev 7的相互作用是由Rev 1 C末端结构域（Rev 1-CT）介导的。我们使用NMR光谱来确定Rev 1-CT结构域（残基1157-1251）及其与来自polη（残基524-539）的Rev 1相互作用区（RIR）的复合物的空间结构。该结构域形成一个四螺旋束，其中一个结构良好的N-末端β-发夹对接螺旋1和2，为RIR基序的两个保守Phe残基创建一个结合口袋，结合后折叠成α-螺旋。核磁共振自旋弛豫和核磁共振弛豫分散测量表明，游离Rev 1-CT和Rev 1-CT/polη-RIR复合物表现出涵盖RIR结合位点的μs-ms构象动力学，这可能有助于选择最适合结合的分子构型。这些结果为理解Y家族DNA聚合酶对Rev 1-CT的识别提供了结构基础，从而为人类细胞中TLS的控制提供了新的见解。
Rev1 is a translesion synthesis (TLS) DNA polymerase essential for DNA damage tolerance in eukaryotes. In the process of TLS stalled high-fidelity replicative DNA polymerases are temporarily replaced by specialized TLS enzymes that can bypass sites of DNA damage (lesions), thus allowing replication to continue or postreplicational gaps to be filled. Despite its limited catalytic activity, human Rev1 plays a key role in TLS by serving as a scaffold that provides an access of Y-family TLS polymerases polη, ι, and κ to their cognate DNA lesions and facilitates their subsequent exchange to polζ that extends the distorted DNA primer-template. Rev1 interaction with the other major human TLS polymerases, polη, ι, κ and the regulatory subunit Rev7 of polζ, is mediated by Rev1 C-terminal domain (Rev1-CT). We used NMR spectroscopy to determine the spatial structure of the Rev1-CT domain (residues 1157-1251) and its complex with Rev1 interacting region (RIR) from polη (residues 524-539). The domain forms a four-helix bundle with a well-structured N-terminal β-hairpin docking against helices 1 and 2, creating a binding pocket for the two conserved Phe residues of the RIR motif that upon binding folds into an α-helix. NMR spin-relaxation and NMR relaxation dispersion measurements suggest that free Rev1-CT and Rev1-CT/polη-RIR complex exhibit μs-ms conformational dynamics encompassing the RIR binding site, which might facilitate selection of the molecular configuration optimal for binding. These results offer new insights into the control of TLS in human cells by providing a structural basis for understanding the recognition of the Rev1-CT by Y-family DNA polymerases.
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作者：
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