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- PDB-8ouw: Cryo-EM structure of CMG helicase bound to TIM-1/TIPN-1 and homod... -
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Open data
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Basic information
Entry | Database: PDB / ID: 8ouw | |||||||||
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Title | Cryo-EM structure of CMG helicase bound to TIM-1/TIPN-1 and homodimeric DNSN-1 on fork DNA (Caenorhabditis elegans) | |||||||||
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![]() | REPLICATION / DNA Replication / DNA Replication Initiation / Replisome / DONSON | |||||||||
Function / homology | ![]() MCM complex assembly / negative regulation of DNA helicase activity / Unwinding of DNA / Assembly of the pre-replicative complex / Orc1 removal from chromatin / Activation of the pre-replicative complex / Switching of origins to a post-replicative state / regulation of sister chromatid cohesion / : / replication fork arrest ...MCM complex assembly / negative regulation of DNA helicase activity / Unwinding of DNA / Assembly of the pre-replicative complex / Orc1 removal from chromatin / Activation of the pre-replicative complex / Switching of origins to a post-replicative state / regulation of sister chromatid cohesion / : / replication fork arrest / gonad development / cell cycle phase transition / GINS complex / DNA strand elongation involved in mitotic DNA replication / nuclear DNA replication / mitotic DNA replication preinitiation complex assembly / meiotic sister chromatid cohesion / mitotic DNA replication / DNA replication checkpoint signaling / MCM complex / DNA replication preinitiation complex / pronucleus / replication fork protection complex / mitotic DNA replication initiation / double-strand break repair via break-induced replication / locomotion / mitotic intra-S DNA damage checkpoint signaling / embryo development ending in birth or egg hatching / regulation of DNA-templated DNA replication initiation / DNA duplex unwinding / mitotic sister chromatid cohesion / DNA strand elongation involved in DNA replication / replication fork processing / DNA unwinding involved in DNA replication / mitotic sister chromatid segregation / DNA replication origin binding / DNA replication initiation / condensed chromosome / DNA helicase activity / helicase activity / meiotic cell cycle / DNA-templated DNA replication / chromosome / nervous system development / single-stranded DNA binding / DNA helicase / DNA replication / hydrolase activity / cell cycle / cell division / DNA repair / chromatin binding / positive regulation of cell population proliferation / chromatin / ATP hydrolysis activity / DNA binding / ATP binding / nucleus / metal ion binding / cytoplasm / cytosol Similarity search - Function | |||||||||
Biological species | ![]() ![]() ![]() ![]() | |||||||||
Method | ELECTRON MICROSCOPY / single particle reconstruction / cryo EM / Resolution: 3.75 Å | |||||||||
![]() | Jenkyn-Bedford, M. / Yeeles, J.T.P. / Labib, K.P.M. | |||||||||
Funding support | ![]()
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![]() | ![]() Title: DNSN-1 recruits GINS for CMG helicase assembly during DNA replication initiation in . Authors: Yisui Xia / Remi Sonneville / Michael Jenkyn-Bedford / Liqin Ji / Constance Alabert / Ye Hong / Joseph T P Yeeles / Karim P M Labib / ![]() ![]() Abstract: Assembly of the CMG (CDC-45-MCM-2-7-GINS) helicase is the key regulated step during eukaryotic DNA replication initiation. Until now, it was unclear whether metazoa require additional factors that ...Assembly of the CMG (CDC-45-MCM-2-7-GINS) helicase is the key regulated step during eukaryotic DNA replication initiation. Until now, it was unclear whether metazoa require additional factors that are not present in yeast. In this work, we show that DNSN-1, the ortholog of human DONSON, functions during helicase assembly in a complex with MUS-101/TOPBP1. DNSN-1 is required to recruit the GINS complex to chromatin, and a cryo-electron microscopy structure indicates that DNSN-1 positions GINS on the MCM-2-7 helicase motor (comprising the six MCM-2 to MCM-7 proteins), by direct binding of DNSN-1 to GINS and MCM-3, using interfaces that we show are important for initiation and essential for viability. These findings identify DNSN-1 as a missing link in our understanding of DNA replication initiation, suggesting that initiation defects underlie the human disease syndrome that results from DONSON mutations. | |||||||||
History |
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Structure visualization
Structure viewer | Molecule: ![]() ![]() |
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Downloads & links
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Download
PDBx/mmCIF format | ![]() | 1.2 MB | Display | ![]() |
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PDB format | ![]() | Display | ![]() | |
PDBx/mmJSON format | ![]() | Tree view | ![]() | |
Others | ![]() |
-Validation report
Summary document | ![]() | 1.4 MB | Display | ![]() |
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Full document | ![]() | 1.4 MB | Display | |
Data in XML | ![]() | 155.4 KB | Display | |
Data in CIF | ![]() | 247.4 KB | Display | |
Arichive directory | ![]() ![]() | HTTPS FTP |
-Related structure data
Related structure data | ![]() 17204MC M: map data used to model this data C: citing same article ( |
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Similar structure data | Similarity search - Function & homology ![]() |
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Links
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Assembly
Deposited unit | ![]()
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Components
-DNA replication licensing factor ... , 6 types, 6 molecules 234567
#1: Protein | Mass: 99448.586 Da / Num. of mol.: 1 Source method: isolated from a genetically manipulated source Source: (gene. exp.) ![]() ![]() ![]() ![]() |
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#2: Protein | Mass: 90810.664 Da / Num. of mol.: 1 Source method: isolated from a genetically manipulated source Source: (gene. exp.) ![]() ![]() ![]() ![]() |
#3: Protein | Mass: 91687.555 Da / Num. of mol.: 1 Source method: isolated from a genetically manipulated source Source: (gene. exp.) ![]() ![]() ![]() ![]() |
#4: Protein | Mass: 85057.734 Da / Num. of mol.: 1 Source method: isolated from a genetically manipulated source Source: (gene. exp.) ![]() ![]() ![]() ![]() |
#5: Protein | Mass: 91248.320 Da / Num. of mol.: 1 Source method: isolated from a genetically manipulated source Source: (gene. exp.) ![]() ![]() ![]() ![]() |
#6: Protein | Mass: 81721.352 Da / Num. of mol.: 1 Source method: isolated from a genetically manipulated source Source: (gene. exp.) ![]() ![]() ![]() ![]() |
-Probable DNA replication complex GINS protein ... , 2 types, 2 molecules AB
#7: Protein | Mass: 23064.361 Da / Num. of mol.: 1 Mutation: Protease-cleaved N-terminal expression tag (Gly-Pro-Gly-Ser) Source method: isolated from a genetically manipulated source Source: (gene. exp.) ![]() ![]() ![]() ![]() |
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#8: Protein | Mass: 20349.330 Da / Num. of mol.: 1 Source method: isolated from a genetically manipulated source Source: (gene. exp.) ![]() ![]() ![]() ![]() |
-DNA replication complex GINS protein ... , 2 types, 2 molecules CD
#9: Protein | Mass: 21717.668 Da / Num. of mol.: 1 Source method: isolated from a genetically manipulated source Source: (gene. exp.) ![]() ![]() ![]() ![]() |
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#10: Protein | Mass: 25705.123 Da / Num. of mol.: 1 Source method: isolated from a genetically manipulated source Source: (gene. exp.) ![]() ![]() ![]() ![]() |
-Protein , 4 types, 6 molecules EFGHKL
#11: Protein | Mass: 66245.758 Da / Num. of mol.: 1 Source method: isolated from a genetically manipulated source Source: (gene. exp.) ![]() ![]() ![]() ![]() | ||||
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#12: Protein | Mass: 66079.383 Da / Num. of mol.: 3 Source method: isolated from a genetically manipulated source Source: (gene. exp.) ![]() ![]() ![]() ![]() #14: Protein | | Mass: 157256.656 Da / Num. of mol.: 1 Source method: isolated from a genetically manipulated source Source: (gene. exp.) ![]() ![]() ![]() ![]() #15: Protein | | Mass: 27415.215 Da / Num. of mol.: 1 Mutation: Protease-cleaved N-terminal expression tag (Gly-Pro-Gly-Ser) Source method: isolated from a genetically manipulated source Source: (gene. exp.) ![]() ![]() ![]() ![]() |
-DNA chain , 2 types, 3 molecules IJM
#13: DNA chain | Mass: 26396.836 Da / Num. of mol.: 2 / Source method: obtained synthetically / Source: (synth.) ![]() ![]() #16: DNA chain | | Mass: 18524.887 Da / Num. of mol.: 1 / Source method: obtained synthetically / Source: (synth.) ![]() ![]() |
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-Non-polymers , 3 types, 13 molecules ![](data/chem/img/ZN.gif)
![](data/chem/img/MG.gif)
![](data/chem/img/ANP.gif)
![](data/chem/img/MG.gif)
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#17: Chemical | ChemComp-ZN / #18: Chemical | ChemComp-MG / #19: Chemical | ChemComp-ANP / |
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-Details
Has ligand of interest | N |
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-Experimental details
-Experiment
Experiment | Method: ELECTRON MICROSCOPY |
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EM experiment | Aggregation state: PARTICLE / 3D reconstruction method: single particle reconstruction |
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Sample preparation
Component | Name: Complex of CMG helicase bound to TIM-1/TIPN-1 and homodimeric DNSN-1 on fork DNA (Caenorhabditis elegans) Type: COMPLEX / Entity ID: #1-#16 / Source: RECOMBINANT |
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Source (natural) | Organism: ![]() ![]() |
Source (recombinant) | Organism: ![]() ![]() |
Buffer solution | pH: 7.5 |
Specimen | Embedding applied: NO / Shadowing applied: NO / Staining applied: NO / Vitrification applied: YES |
Specimen support | Details: 15 mA / Grid material: COPPER / Grid mesh size: 400 divisions/in. / Grid type: Quantifoil R2/2 |
Vitrification | Cryogen name: ETHANE |
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Electron microscopy imaging
Experimental equipment | ![]() Model: Titan Krios / Image courtesy: FEI Company |
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Microscopy | Model: FEI TITAN KRIOS |
Electron gun | Electron source: ![]() |
Electron lens | Mode: BRIGHT FIELD / Nominal defocus max: 2500 nm / Nominal defocus min: 1000 nm |
Image recording | Average exposure time: 1.7 sec. / Electron dose: 40.1 e/Å2 / Film or detector model: GATAN K3 BIOQUANTUM (6k x 4k) |
EM imaging optics | Energyfilter name: GIF Bioquantum / Energyfilter slit width: 20 eV |
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Processing
EM software |
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CTF correction | Type: PHASE FLIPPING AND AMPLITUDE CORRECTION | ||||||||||||
3D reconstruction | Resolution: 3.75 Å / Resolution method: FSC 0.143 CUT-OFF / Num. of particles: 33900 / Symmetry type: POINT |