Multistep loading of a DNA sliding clamp onto DNA by replication factor C.
Multistep loading of a DNA sliding clamp onto DNA by replication factor C.
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DOI：
10.7554/elife.78253
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发表时间：
2022-08-08
期刊：
影响因子：
7.7
通讯作者：
Hite, Richard K.
中科院分区：
文献类型：
--
作者：
Schrecker, Marina;Castaneda, Juan C.;Devbhandari, Sujan;Kumar, Charanya;Remus, Dirk;Hite, Richard K.
DNA滑动钳增殖细胞核抗原（PCNA）是许多真核生物DNA代谢酶的重要辅助因子。PCNA通过atp依赖性钳夹装载器复制因子C （RFC）装载在DNA周围，RFC作用于单链(ss)/双链DNA （dsDNA）连接处，其中包含一个凹陷的3 ‘端（3 ’ ss/dsDNA连接处）和DNA缺口。为了阐明装载机制，我们利用低温电镜研究了RFC:PCNA与atp - γ s和3 ' s/dsDNA连接或缺口DNA结合的结构。出乎意料的是，我们在RFC:PCNA:DNA复合体中观察到了开放和封闭的PCNA构象，揭示了PCNA可以采用开放的平面构象，允许直接插入dsDNA，并提出了PCNA环闭合是否与ATP水解机制耦合的问题。通过解析RFC:PCNA的多种dna结合状态，我们观察到部分熔融有助于向RFC:PCNA形成的中心通道侧插入。我们还解析了Rfc1 n端结构域，并证明其单一BRCT结构域在插入中央RFC通道之前参与协调DNA，从而促进体外复制叉的滞后链上的PCNA装载。综上所述，我们的数据为rfc催化的PCNA装载到DNA上提供了一个全面的、从根本上修正的模型。
The DNA sliding clamp proliferating cell nuclear antigen (PCNA) is an essential co-factor for many eukaryotic DNA metabolic enzymes. PCNA is loaded around DNA by the ATP-dependent clamp loader replication factor C (RFC), which acts at single-stranded (ss)/double-stranded DNA (dsDNA) junctions harboring a recessed 3’ end (3’ ss/dsDNA junctions) and at DNA nicks. To illuminate the loading mechanism we have investigated the structure of RFC:PCNA bound to ATPγS and 3’ ss/dsDNA junctions or nicked DNA using cryogenic electron microscopy. Unexpectedly, we observe open and closed PCNA conformations in the RFC:PCNA:DNA complex, revealing that PCNA can adopt an open, planar conformation that allows direct insertion of dsDNA, and raising the question of whether PCNA ring closure is mechanistically coupled to ATP hydrolysis. By resolving multiple DNA-bound states of RFC:PCNA we observe that partial melting facilitates lateral insertion into the central channel formed by RFC:PCNA. We also resolve the Rfc1 N-terminal domain and demonstrate that its single BRCT domain participates in coordinating DNA prior to insertion into the central RFC channel, which promotes PCNA loading on the lagging strand of replication forks in vitro. Combined, our data suggest a comprehensive and fundamentally revised model for the RFC-catalyzed loading of PCNA onto DNA.
DOI：
10.1002/bies.201700070
发表时间：
2017-08
期刊：
BioEssays : news and reviews in molecular, cellular and developmental biology
影响因子：
--
作者：
Kunkel TA;Burgers PMJ
通讯作者：
Burgers PMJ
影响因子：
3.5
作者：
Chiraniya A;Finkelstein J;O'Donnell M;Bloom LB
通讯作者：
Bloom LB
影响因子：
7.8
作者：
Tang H;Hilton B;Musich PR;Fang DZ;Zou Y
通讯作者：
Zou Y