Structure of the human clamp loader reveals an autoinhibited conformation of a substrate-bound AAA plus switch

Gaubitz Christl; Liu Xingchen; Magrino JosephStone Nicholas P.Landeck JacobHedglin MarkKelch Brian A.

首页> 外文期刊>Proceedings of the National Academy of Sciences of the United States of America. >Structure of the human clamp loader reveals an autoinhibited conformation of a substrate-bound AAA plus switch

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Structure of the human clamp loader reveals an autoinhibited conformation of a substrate-bound AAA plus switch

机译：Structure of the human clamp loader reveals an autoinhibited conformation of a substrate-bound AAA plus switch

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DNA replication requires the sliding clamp, a ring-shaped protein complex that encircles DNA, where it acts as an essential cofactor for DNA polymerases and other proteins. The sliding clamp needs to be opened and installed onto DNA by a clamp loader ATPase of the AAA+ family. The human clamp loader replication factor C (RFC) and sliding clamp proliferating cell nuclear antigen (PCNA) are both essential and play critical roles in several diseases. Despite decades of study, no structure of human RFC has been resolved. Here, we report the structure of human RFC bound to PCNA by cryogenic electron microscopy to an overall resolution of similar to 3.4 angstrom. The active sites of RFC are fully bound to adenosine 5'-triphosphate (ATP) analogs, which is expected to induce opening of the sliding clamp. However, we observe the complex in a conformation before PCNA opening, with the clamp loader ATPase modules forming an overtwisted spiral that is incapable of binding DNA or hydrolyzing ATP. The autoinhibited conformation observed here has many similarities to a previous yeast RFC:PCNA crystal structure, suggesting that eukaryotic clamp loaders adopt a similar autoinhibited state early on in clamp loading. Our results point to a "limited change/induced fit" mechanism in which the clamp first opens, followed by DNA binding, inducing opening of the loader to release autoinhibition. The proposed change from an overtwisted to an active conformation reveals an additional regulatory mechanism for AAA+ ATPases. Finally, our structural analysis of disease mutations leads to a mechanistic explanation for the role of RFC in human health.

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《Proceedings of the National Academy of Sciences of the United States of America.》 |2020年第38期|23571-23580|共10页
作者
Gaubitz Christl; Liu Xingchen; Magrino JosephStone Nicholas P.Landeck JacobHedglin MarkKelch Brian A.;
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作者单位

Dept Biochem & Mol Pharmacol,Univ Massachusetts;

Dept Chem,Penn State Univ;

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收录信息美国《科学引文索引》(SCI);美国《生物学医学文摘》(MEDLINE);美国《化学文摘》(CA);
原文格式 PDF
正文语种英语
中图分类自然科学总论;
关键词
sliding clamp; DNA replication; AAA; ATPase; clamp loader; REPLICATION FACTOR-C; DNA-POLYMERASE CLAMP; CELL NUCLEAR ANTIGEN; ESCHERICHIA-COLI; SLIDING CLAMP; ATP-BINDING; III HOLOENZYME; DELTA-SUBUNIT; PCNA; MECHANISM;

Structure of the human clamp loader reveals an autoinhibited conformation of a substrate-bound AAA plus switch

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