- EMDB-25568: Structure of the yeast clamp loader (Replication Factor C RFC) bo... -
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基本情報
登録情報
データベース: EMDB / ID: EMD-25568
タイトル
Structure of the yeast clamp loader (Replication Factor C RFC) bound to the sliding clamp (Proliferating Cell Nuclear Antigen PCNA) in an autoinhibited conformation
マップデータ
Density modified map (with resolve_cryo_em)
試料
複合体: Replication Factor C (RFC), the eukaryotic clamp loader
National Institutes of Health/National Institute of General Medical Sciences (NIH/NIGMS)
R01-GM127776
チェコ
Swiss National Science Foundation
168972
チェコ
Swiss National Science Foundation
177859
チェコ
Ministry of Education (MoE, Czech Republic)
LL2008
チェコ
引用
ジャーナル: Elife / 年: 2022 タイトル: Cryo-EM structures reveal high-resolution mechanism of a DNA polymerase sliding clamp loader. 著者: Christl Gaubitz / Xingchen Liu / Joshua Pajak / Nicholas P Stone / Janelle A Hayes / Gabriel Demo / Brian A Kelch / 要旨: Sliding clamps are ring-shaped protein complexes that are integral to the DNA replication machinery of all life. Sliding clamps are opened and installed onto DNA by clamp loader AAA+ ATPase complexes. ...Sliding clamps are ring-shaped protein complexes that are integral to the DNA replication machinery of all life. Sliding clamps are opened and installed onto DNA by clamp loader AAA+ ATPase complexes. However, how a clamp loader opens and closes the sliding clamp around DNA is still unknown. Here, we describe structures of the clamp loader Replication Factor C (RFC) bound to its cognate sliding clamp Proliferating Cell Nuclear Antigen (PCNA) en route to successful loading. RFC first binds to PCNA in a dynamic, closed conformation that blocks both ATPase activity and DNA binding. RFC then opens the PCNA ring through a large-scale 'crab-claw' expansion of both RFC and PCNA that explains how RFC prefers initial binding of PCNA over DNA. Next, the open RFC:PCNA complex binds DNA and interrogates the primer-template junction using a surprising base-flipping mechanism. Our structures indicate that initial PCNA opening and subsequent closure around DNA do not require ATP hydrolysis, but are driven by binding energy. ATP hydrolysis, which is necessary for RFC release, is triggered by interactions with both PCNA and DNA, explaining RFC's switch-like ATPase activity. Our work reveals how a AAA+ machine undergoes dramatic conformational changes for achieving binding preference and substrate remodeling.
解像度のタイプ: BY AUTHOR / 解像度: 3.9 Å / 解像度の算出法: FSC 0.143 CUT-OFF / ソフトウェア - 名称: RELION 詳細: even/odd maps were refined totally independent (gold standard) 使用した粒子像数: 55308
初期 角度割当
タイプ: RANDOM ASSIGNMENT / ソフトウェア - 名称: cisTEM
最終 角度割当
タイプ: MAXIMUM LIKELIHOOD / ソフトウェア - 名称: RELION
FSC曲線 (解像度の算出)
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原子モデル構築 1
精密化
空間: REAL / プロトコル: FLEXIBLE FIT
得られたモデル
PDB-7thj: Structure of the yeast clamp loader (Replication Factor C RFC) bound to the sliding clamp (Proliferating Cell Nuclear Antigen PCNA) in an autoinhibited conformation