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- PDB-8p63: S. cerevisiae consensus-sCMGE on ssDNA after DNA replication init... -
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Open data
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Basic information
Entry | Database: PDB / ID: 8p63 | |||||||||||||||
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Title | S. cerevisiae consensus-sCMGE on ssDNA after DNA replication initiation | |||||||||||||||
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![]() | REPLICATION / Saccharomyces cerevisiae / helicase / CMGE / initiation of DNA replication / DNA / DNA unwinding | |||||||||||||||
Function / homology | ![]() DNA-templated DNA replication maintenance of fidelity / gene conversion / Unwinding of DNA / DNA replication initiation / epsilon DNA polymerase complex / MCM core complex / Assembly of the pre-replicative complex / Switching of origins to a post-replicative state / MCM complex binding / GINS complex ...DNA-templated DNA replication maintenance of fidelity / gene conversion / Unwinding of DNA / DNA replication initiation / epsilon DNA polymerase complex / MCM core complex / Assembly of the pre-replicative complex / Switching of origins to a post-replicative state / MCM complex binding / GINS complex / DNA strand elongation involved in mitotic DNA replication / nuclear DNA replication / mitotic DNA replication preinitiation complex assembly / premeiotic DNA replication / pre-replicative complex assembly involved in nuclear cell cycle DNA replication / mitotic DNA replication / SUMO binding / Activation of the pre-replicative complex / CMG complex / nuclear pre-replicative complex / single-stranded DNA 3'-5' DNA exonuclease activity / MCM complex / Activation of ATR in response to replication stress / DNA replication preinitiation complex / Termination of translesion DNA synthesis / mitotic DNA replication checkpoint signaling / replication fork protection complex / double-strand break repair via break-induced replication / mitotic DNA replication initiation / single-stranded DNA helicase activity / mitotic intra-S DNA damage checkpoint signaling / silent mating-type cassette heterochromatin formation / regulation of DNA-templated DNA replication initiation / Hydrolases; Acting on ester bonds; Exodeoxyribonucleases producing 5'-phosphomonoesters / nucleotide-excision repair, DNA gap filling / mitotic sister chromatid cohesion / DNA replication proofreading / DNA strand elongation involved in DNA replication / DNA unwinding involved in DNA replication / leading strand elongation / nuclear replication fork / DNA replication origin binding / Dual incision in TC-NER / DNA replication initiation / subtelomeric heterochromatin formation / error-prone translesion synthesis / DNA helicase activity / base-excision repair, gap-filling / replication fork / helicase activity / transcription elongation by RNA polymerase II / base-excision repair / heterochromatin formation / DNA-templated DNA replication / double-strand break repair via nonhomologous end joining / double-strand break repair / mitotic cell cycle / single-stranded DNA binding / 4 iron, 4 sulfur cluster binding / double-stranded DNA binding / DNA helicase / chromosome, telomeric region / DNA-directed DNA polymerase / DNA-directed DNA polymerase activity / nucleotide binding / mRNA binding / DNA damage response / chromatin binding / ATP hydrolysis activity / DNA binding / zinc ion binding / nucleoplasm / ATP binding / nucleus / metal ion binding / cytoplasm Similarity search - Function | |||||||||||||||
Biological species | ![]() ![]() synthetic construct (others) | |||||||||||||||
Method | ELECTRON MICROSCOPY / single particle reconstruction / cryo EM / Resolution: 3.7 Å | |||||||||||||||
![]() | Henrikus, S.S. / Willhoft, O. | |||||||||||||||
Funding support | ![]() ![]()
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![]() | ![]() Title: Unwinding of a eukaryotic origin of replication visualized by cryo-EM. Authors: Sarah S Henrikus / Marta H Gross / Oliver Willhoft / Thomas Pühringer / Jacob S Lewis / Allison W McClure / Julia F Greiwe / Giacomo Palm / Andrea Nans / John F X Diffley / Alessandro Costa / ![]() ![]() ![]() Abstract: To prevent detrimental chromosome re-replication, DNA loading of a double hexamer of the minichromosome maintenance (MCM) replicative helicase is temporally separated from DNA unwinding. Upon S-phase ...To prevent detrimental chromosome re-replication, DNA loading of a double hexamer of the minichromosome maintenance (MCM) replicative helicase is temporally separated from DNA unwinding. Upon S-phase transition in yeast, DNA unwinding is achieved in two steps: limited opening of the double helix and topological separation of the two DNA strands. First, Cdc45, GINS and Polε engage MCM to assemble a double CMGE with two partially separated hexamers that nucleate DNA melting. In the second step, triggered by Mcm10, two CMGEs separate completely, eject the lagging-strand template and cross paths. To understand Mcm10 during helicase activation, we used biochemical reconstitution with cryogenic electron microscopy. We found that Mcm10 splits the double CMGE by engaging the N-terminal homo-dimerization face of MCM. To eject the lagging strand, DNA unwinding is started from the N-terminal side of MCM while the hexamer channel becomes too narrow to harbor duplex DNA. | |||||||||||||||
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
Arichive directory | ![]() ![]() | HTTPS FTP |
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-Related structure data
Related structure data | ![]() 17459MC ![]() 8p5eC ![]() 8p62C 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 ... , 5 types, 5 molecules 23467
#1: Protein | Mass: 98911.539 Da / Num. of mol.: 1 Source method: isolated from a genetically manipulated source Source: (gene. exp.) ![]() ![]() Gene: MCM2, YBL023C, YBL0438 / Production host: ![]() ![]() |
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#2: Protein | Mass: 111720.242 Da / Num. of mol.: 1 Source method: isolated from a genetically manipulated source Source: (gene. exp.) ![]() ![]() Gene: MCM3, YEL032W, SYGP-ORF23 / Production host: ![]() ![]() |
#3: Protein | Mass: 105138.375 Da / Num. of mol.: 1 Source method: isolated from a genetically manipulated source Source: (gene. exp.) ![]() ![]() Gene: MCM4, CDC54, HCD21, YPR019W, YP9531.13 / Production host: ![]() ![]() |
#5: Protein | Mass: 113110.211 Da / Num. of mol.: 1 Source method: isolated from a genetically manipulated source Source: (gene. exp.) ![]() ![]() Gene: MCM6, YGL201C / Production host: ![]() ![]() |
#6: Protein | Mass: 95049.875 Da / Num. of mol.: 1 Source method: isolated from a genetically manipulated source Source: (gene. exp.) ![]() ![]() Gene: MCM7, CDC47, YBR202W, YBR1441 / Production host: ![]() ![]() |
-Protein , 2 types, 2 molecules 5E
#4: Protein | Mass: 86505.734 Da / Num. of mol.: 1 Source method: isolated from a genetically manipulated source Source: (gene. exp.) ![]() ![]() Gene: MCM5, CDC46, YLR274W, L9328.1 / Production host: ![]() ![]() |
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#10: Protein | Mass: 75154.703 Da / Num. of mol.: 1 Source method: isolated from a genetically manipulated source Source: (gene. exp.) ![]() ![]() Gene: CDC45, SLD4, YLR103C, L8004.11 / Production host: ![]() ![]() |
-DNA chain , 1 types, 1 molecules A
#7: DNA chain | Mass: 2773.904 Da / Num. of mol.: 1 / Source method: obtained synthetically / Source: (synth.) synthetic construct (others) |
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-DNA replication complex GINS protein ... , 4 types, 4 molecules CDHI
#8: Protein | Mass: 25718.070 Da / Num. of mol.: 1 Source method: isolated from a genetically manipulated source Source: (gene. exp.) ![]() ![]() Gene: PSF3, YOL146W / Production host: ![]() ![]() |
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#9: Protein | Mass: 33983.617 Da / Num. of mol.: 1 Source method: isolated from a genetically manipulated source Source: (gene. exp.) ![]() ![]() Gene: SLD5, YDR489W / Production host: ![]() ![]() |
#13: Protein | Mass: 24230.576 Da / Num. of mol.: 1 Source method: isolated from a genetically manipulated source Source: (gene. exp.) ![]() ![]() Gene: PSF1, YDR013W, PZA208, YD8119.18 / Production host: ![]() ![]() |
#14: Protein | Mass: 25096.807 Da / Num. of mol.: 1 Source method: isolated from a genetically manipulated source Source: (gene. exp.) ![]() ![]() Gene: PSF2, YJL072C, HRF213, J1086 / Production host: ![]() ![]() |
-DNA polymerase epsilon ... , 2 types, 2 molecules FG
#11: Protein | Mass: 78425.852 Da / Num. of mol.: 1 Source method: isolated from a genetically manipulated source Source: (gene. exp.) ![]() ![]() Gene: DPB2, YPR175W, P9705.7 / Production host: ![]() ![]() |
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#12: Protein | Mass: 255992.484 Da / Num. of mol.: 1 Source method: isolated from a genetically manipulated source Source: (gene. exp.) ![]() ![]() Gene: POL2, DUN2, YNL262W, N0825 / Production host: ![]() ![]() References: UniProt: P21951, DNA-directed DNA polymerase, Hydrolases; Acting on ester bonds; Exodeoxyribonucleases producing 5'-phosphomonoesters |
-Non-polymers , 4 types, 16 molecules ![](data/chem/img/ATP.gif)
![](data/chem/img/ZN.gif)
![](data/chem/img/MG.gif)
![](data/chem/img/ADP.gif)
![](data/chem/img/ZN.gif)
![](data/chem/img/MG.gif)
![](data/chem/img/ADP.gif)
#15: Chemical | ChemComp-ATP / #16: Chemical | ChemComp-ZN / #17: Chemical | #18: Chemical | |
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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
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Source (natural) |
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Source (recombinant) |
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Buffer solution | pH: 7.6 | ||||||||||||||||||||||||||||||||||||||||||
Specimen | Embedding applied: NO / Shadowing applied: NO / Staining applied: NO / Vitrification applied: YES | ||||||||||||||||||||||||||||||||||||||||||
Vitrification | Instrument: FEI VITROBOT MARK IV / 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: 3300 nm / Nominal defocus min: 1800 nm |
Image recording | Electron dose: 1.67 e/Å2 / Film or detector model: GATAN K2 SUMMIT (4k x 4k) |
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Processing
EM software |
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CTF correction | Type: PHASE FLIPPING AND AMPLITUDE CORRECTION | ||||||||||||||||||||||||
3D reconstruction | Resolution: 3.7 Å / Resolution method: FSC 0.143 CUT-OFF / Num. of particles: 172552 / Symmetry type: POINT |