Switching of origins to a post-replicative state / Unwinding of DNA / DNA strand elongation involved in mitotic DNA replication / GINS complex / mitotic DNA replication preinitiation complex assembly / nuclear origin of replication recognition complex / Regulation of MITF-M-dependent genes involved in DNA replication, damage repair and senescence / alpha DNA polymerase:primase complex / mitotic DNA replication / DNA replication checkpoint signaling ...Switching of origins to a post-replicative state / Unwinding of DNA / DNA strand elongation involved in mitotic DNA replication / GINS complex / mitotic DNA replication preinitiation complex assembly / nuclear origin of replication recognition complex / Regulation of MITF-M-dependent genes involved in DNA replication, damage repair and senescence / alpha DNA polymerase:primase complex / mitotic DNA replication / DNA replication checkpoint signaling / CMG complex / regulation of phosphorylation / DNA replication preinitiation complex / MCM complex / double-strand break repair via break-induced replication / mitotic DNA replication initiation / regulation of DNA-templated DNA replication initiation / inner cell mass cell proliferation / DNA strand elongation involved in DNA replication / cochlea development / G1/S-Specific Transcription / DNA replication origin binding / Activation of the pre-replicative complex / DNA replication initiation / Activation of ATR in response to replication stress / cellular response to epidermal growth factor stimulus / cellular response to interleukin-4 / Assembly of the pre-replicative complex / multicellular organism growth / DNA-templated DNA replication / Orc1 removal from chromatin / cellular senescence / cellular response to xenobiotic stimulus / nucleosome assembly / single-stranded DNA binding / DNA helicase / histone binding / forked DNA-dependent helicase activity / single-stranded 3'-5' DNA helicase activity / four-way junction helicase activity / double-stranded DNA helicase activity / chromosome, telomeric region / cell population proliferation / DNA replication / ciliary basal body / cilium / intracellular membrane-bounded organelle / apoptotic process / centrosome / DNA damage response / chromatin binding / chromatin / perinuclear region of cytoplasm / enzyme binding / ATP hydrolysis activity / DNA binding / zinc ion binding / nucleoplasm / ATP binding / identical protein binding / nucleus / membrane / cytosol / cytoplasm 類似検索 - 分子機能
DNA replication licensing factor MCM2-like, winged-helix domain / : / PSF3 N-terminal domain / CDC45 family / DNA replication complex GINS protein Psf2 / CDC45 / GINS complex, subunit Psf3 / DNA replication complex GINS protein SLD5, C-terminal / GINS complex, subunit Psf3 superfamily / GINS complex protein Sld5, alpha-helical domain ...DNA replication licensing factor MCM2-like, winged-helix domain / : / PSF3 N-terminal domain / CDC45 family / DNA replication complex GINS protein Psf2 / CDC45 / GINS complex, subunit Psf3 / DNA replication complex GINS protein SLD5, C-terminal / GINS complex, subunit Psf3 superfamily / GINS complex protein Sld5, alpha-helical domain / DNA replication complex GINS protein SLD5 C-terminus / GINS complex, subunit Psf1 / GINS complex subunit Sld5 / GINS subunit, domain A / GINS, helical bundle-like domain superfamily / GINS complex protein helical bundle domain / : / MCM5, C-terminal domain / DNA replication licensing factor Mcm5 / DNA replication licensing factor Mcm3 / Mini-chromosome maintenance complex protein 4 / MCM3-like, winged helix domain / DNA replication licensing factor Mcm6 / DNA replication licensing factor Mcm7 / Mcm6, C-terminal winged-helix domain / MCM6 C-terminal winged-helix domain / DNA replication licensing factor Mcm2 / Mini-chromosome maintenance protein 2 / Mini-chromosome maintenance, conserved site / MCM family signature. / MCM N-terminal domain / MCM N-terminal domain / MCM OB domain / MCM OB domain / Mini-chromosome maintenance protein / MCM, AAA-lid domain / MCM P-loop domain / MCM AAA-lid domain / MCM family domain profile. / minichromosome maintenance proteins / MCM domain / Nucleic acid-binding, OB-fold / ATPases associated with a variety of cellular activities / AAA+ ATPase domain / P-loop containing nucleoside triphosphate hydrolase 類似検索 - ドメイン・相同性
Cell division control protein 45 homolog / DNA replication licensing factor MCM3 / DNA replication licensing factor MCM4 / DNA replication licensing factor MCM5 / DNA replication licensing factor MCM7 / DNA replication licensing factor MCM2 / DNA replication licensing factor MCM6 / DNA replication complex GINS protein PSF1 / DNA replication complex GINS protein SLD5 / DNA replication complex GINS protein PSF3 / DNA replication complex GINS protein PSF2 類似検索 - 構成要素
National Institutes of Health/National Institute of General Medical Sciences (NIH/NIGMS)
R35GM152094
米国
National Institutes of Health/National Institute of General Medical Sciences (NIH/NIGMS)
R35GM126907
米国
National Institutes of Health/National Cancer Institute (NIH/NCI)
P30 CA008748
米国
引用
ジャーナル: Science / 年: 2025 タイトル: G-quadruplex-stalled eukaryotic replisome structure reveals helical inchworm DNA translocation. 著者: Sahil Batra / Benjamin Allwein / Charanya Kumar / Sujan Devbhandari / Jan-Gert Brüning / Soon Bahng / Chong M Lee / Kenneth J Marians / Richard K Hite / Dirk Remus / 要旨: DNA G-quadruplexes (G4s) are non-B-form DNA secondary structures that threaten genome stability by impeding DNA replication. To elucidate how G4s induce replication fork arrest, we characterized fork ...DNA G-quadruplexes (G4s) are non-B-form DNA secondary structures that threaten genome stability by impeding DNA replication. To elucidate how G4s induce replication fork arrest, we characterized fork collisions with preformed G4s in the parental DNA using reconstituted yeast and human replisomes. We demonstrate that a single G4 in the leading strand template is sufficient to stall replisomes by arresting the CMG helicase. Cryo-electron microscopy structures of stalled yeast and human CMG complexes reveal that the folded G4 is lodged inside the central CMG channel, arresting translocation. The G4 stabilizes the CMG at distinct translocation intermediates, suggesting an unprecedented helical inchworm mechanism for DNA translocation. These findings illuminate the eukaryotic replication fork mechanism under normal and perturbed conditions.
凍結剤: ETHANE / チャンバー内湿度: 95 % / チャンバー内温度: 277 K / 装置: FEI VITROBOT MARK IV 詳細: 30 second wait time after sample application; 30 second blot time, blot force 0.
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電子顕微鏡法
顕微鏡
TFS KRIOS
撮影
フィルム・検出器のモデル: GATAN K3 (6k x 4k) / 撮影したグリッド数: 1 / 実像数: 3325 / 平均露光時間: 3.0 sec. / 平均電子線量: 66.0 e/Å2
タイプ: OTHER / ソフトウェア - 名称: cryoSPARC (ver. 4.0.0) 詳細: Stochastic gradient descent (cryoSPARC ab initio model generation)
最終 角度割当
タイプ: MAXIMUM LIKELIHOOD / ソフトウェア - 名称: cryoSPARC (ver. 4.0.0)
最終 3次元分類
ソフトウェア - 名称: cryoSPARC (ver. 4.0.0)
FSC曲線 (解像度の算出)
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原子モデル構築 1
初期モデル
Chain - Source name: Other / Chain - Initial model type: in silico model / 詳細: ModelAngelo
詳細
Initial fitting was performed de novo by ModelAngelo, then iteratively improved with ChimeraX/ISOLDE, Coot, and Phenix real-space refinement algorithms.
精密化
空間: REAL / プロトコル: AB INITIO MODEL / 温度因子: 49.99
得られたモデル
PDB-9e2z: Cryo-EM structure of human CMG helicase stalled at G4-containing DNA template