+
Open data
-
Basic information
Entry | Database: PDB / ID: 7pfo | |||||||||
---|---|---|---|---|---|---|---|---|---|---|
Title | Core human replisome | |||||||||
![]() |
| |||||||||
![]() | REPLICATION / DNA replication / helicase / CMG / TIMELESS / TIPIN / CLASPIN / AND-1 / EPSILON / POLYMERASE / REPLISOME | |||||||||
Function / homology | ![]() cellular response to bleomycin / DNA secondary structure binding / Switching of origins to a post-replicative state / detection of abiotic stimulus / replication fork arrest / regulation of nuclear cell cycle DNA replication / Unwinding of DNA / cell cycle phase transition / cellular response to cisplatin / DNA replication initiation ...cellular response to bleomycin / DNA secondary structure binding / Switching of origins to a post-replicative state / detection of abiotic stimulus / replication fork arrest / regulation of nuclear cell cycle DNA replication / Unwinding of DNA / cell cycle phase transition / cellular response to cisplatin / DNA replication initiation / epsilon DNA polymerase complex / GINS complex / DNA strand elongation involved in mitotic DNA replication / mitotic DNA replication preinitiation complex assembly / nuclear origin of replication recognition complex / cellular response to hydroxyurea / alpha DNA polymerase:primase complex / mitotic DNA replication / anaphase-promoting complex binding / CMG complex / DNA replication checkpoint signaling / single-stranded DNA 3'-5' DNA exonuclease activity / MCM complex / DNA replication preinitiation complex / regulation of phosphorylation / mitotic DNA replication checkpoint signaling / replication fork protection complex / double-strand break repair via break-induced replication / mitotic DNA replication initiation / mitotic intra-S DNA damage checkpoint signaling / regulation of DNA-templated DNA replication initiation / positive regulation of double-strand break repair / Hydrolases; Acting on ester bonds; Exodeoxyribonucleases producing 5'-phosphomonoesters / inner cell mass cell proliferation / nucleotide-excision repair, DNA gap filling / DNA replication proofreading / DNA strand elongation involved in DNA replication / branching morphogenesis of an epithelial tube / cochlea development / DNA synthesis involved in DNA repair / G1/S-Specific Transcription / activation of protein kinase activity / DNA unwinding involved in DNA replication / replication fork processing / leading strand elongation / Apoptotic cleavage of cellular proteins / nuclear replication fork / mitotic G2 DNA damage checkpoint signaling / DNA replication origin binding / positive regulation of double-strand break repair via homologous recombination / Activation of the pre-replicative complex / DNA replication initiation / PCNA-Dependent Long Patch Base Excision Repair / cellular response to interleukin-4 / embryonic organ development / error-prone translesion synthesis / Activation of ATR in response to replication stress / response to UV / DNA helicase activity / base-excision repair, gap-filling / Gap-filling DNA repair synthesis and ligation in GG-NER / ciliary basal body / DNA damage checkpoint signaling / helicase activity / Assembly of the pre-replicative complex / morphogenesis of an epithelium / Recognition of DNA damage by PCNA-containing replication complex / Termination of translesion DNA synthesis / lung development / HDR through Homologous Recombination (HRR) / regulation of circadian rhythm / Dual Incision in GG-NER / DNA-templated DNA replication / Dual incision in TC-NER / Orc1 removal from chromatin / Gap-filling DNA repair synthesis and ligation in TC-NER / circadian rhythm / G1/S transition of mitotic cell cycle / cellular response to xenobiotic stimulus / nucleosome assembly / mitotic cell cycle / site of double-strand break / single-stranded DNA binding / Processing of DNA double-strand break ends / 4 iron, 4 sulfur cluster binding / histone binding / peptidyl-serine phosphorylation / DNA helicase / DNA replication / cell population proliferation / chromosome, telomeric region / DNA-directed DNA polymerase / DNA-directed DNA polymerase activity / nuclear body / Ub-specific processing proteases / cell division / intracellular membrane-bounded organelle / DNA repair / nucleotide binding / negative regulation of DNA-templated transcription Similarity search - Function | |||||||||
Biological species | ![]() | |||||||||
Method | ELECTRON MICROSCOPY / single particle reconstruction / cryo EM / Resolution: 3.2 Å | |||||||||
![]() | Jones, M.J. / Yeeles, J.T.P. | |||||||||
Funding support | ![]()
| |||||||||
![]() | ![]() Title: Structure of a human replisome shows the organisation and interactions of a DNA replication machine. Authors: Morgan L Jones / Yasemin Baris / Martin R G Taylor / Joseph T P Yeeles / ![]() Abstract: The human replisome is an elaborate arrangement of molecular machines responsible for accurate chromosome replication. At its heart is the CDC45-MCM-GINS (CMG) helicase, which, in addition to ...The human replisome is an elaborate arrangement of molecular machines responsible for accurate chromosome replication. At its heart is the CDC45-MCM-GINS (CMG) helicase, which, in addition to unwinding the parental DNA duplex, arranges many proteins including the leading-strand polymerase Pol ε, together with TIMELESS-TIPIN, CLASPIN and AND-1 that have key and varied roles in maintaining smooth replisome progression. How these proteins are coordinated in the human replisome is poorly understood. We have determined a 3.2 Å cryo-EM structure of a human replisome comprising CMG, Pol ε, TIMELESS-TIPIN, CLASPIN and AND-1 bound to replication fork DNA. The structure permits a detailed understanding of how AND-1, TIMELESS-TIPIN and Pol ε engage CMG, reveals how CLASPIN binds to multiple replisome components and identifies the position of the Pol ε catalytic domain. Furthermore, the intricate network of contacts contributed by MCM subunits and TIMELESS-TIPIN with replication fork DNA suggests a mechanism for strand separation. #1: ![]() Title: A conserved mechanism for regulating replisome disassembly in eukaryotes. Authors: Michael Jenkyn-Bedford / Morgan L Jones / Yasemin Baris / Karim P M Labib / Giuseppe Cannone / Joseph T P Yeeles / Tom D Deegan / ![]() Abstract: Replisome disassembly is the final step of eukaryotic DNA replication and is triggered by ubiquitylation of the CDC45-MCM-GINS (CMG) replicative helicase. Despite being driven by evolutionarily ...Replisome disassembly is the final step of eukaryotic DNA replication and is triggered by ubiquitylation of the CDC45-MCM-GINS (CMG) replicative helicase. Despite being driven by evolutionarily diverse E3 ubiquitin ligases in different eukaryotes (SCF in budding yeast, CUL2 in metazoa), replisome disassembly is governed by a common regulatory principle, in which ubiquitylation of CMG is suppressed before replication termination, to prevent replication fork collapse. Recent evidence suggests that this suppression is mediated by replication fork DNA. However, it is unknown how SCF and CUL2 discriminate terminated from elongating replisomes, to selectively ubiquitylate CMG only after termination. Here we used cryo-electron microscopy to solve high-resolution structures of budding yeast and human replisome-E3 ligase assemblies. Our structures show that the leucine-rich repeat domains of Dia2 and LRR1 are structurally distinct, but bind to a common site on CMG, including the MCM3 and MCM5 zinc-finger domains. The LRR-MCM interaction is essential for replisome disassembly and, crucially, is occluded by the excluded DNA strand at replication forks, establishing the structural basis for the suppression of CMG ubiquitylation before termination. Our results elucidate a conserved mechanism for the regulation of replisome disassembly in eukaryotes, and reveal a previously unanticipated role for DNA in preserving replisome integrity. | |||||||||
History |
|
-
Structure visualization
Movie |
![]() |
---|---|
Structure viewer | Molecule: ![]() ![]() |
-
Downloads & links
-
Download
PDBx/mmCIF format | ![]() | 2.9 MB | Display | ![]() |
---|---|---|---|---|
PDB format | ![]() | Display | ![]() | |
PDBx/mmJSON format | ![]() | Tree view | ![]() | |
Others | ![]() |
-Validation report
Arichive directory | ![]() ![]() | HTTPS FTP |
---|
-Related structure data
Related structure data | ![]() 13375MC M: map data used to model this data C: citing same article ( |
---|---|
Similar structure data |
-
Links
-
Assembly
Deposited unit | ![]()
|
---|---|
1 |
|
-
Components
-DNA replication licensing factor ... , 6 types, 6 molecules 234567
#1: Protein | Mass: 102034.102 Da / Num. of mol.: 1 Source method: isolated from a genetically manipulated source Source: (gene. exp.) ![]() ![]() |
---|---|
#2: Protein | Mass: 91110.852 Da / Num. of mol.: 1 Source method: isolated from a genetically manipulated source Source: (gene. exp.) ![]() ![]() |
#3: Protein | Mass: 96684.852 Da / Num. of mol.: 1 Source method: isolated from a genetically manipulated source Source: (gene. exp.) ![]() ![]() |
#4: Protein | Mass: 82406.633 Da / Num. of mol.: 1 Source method: isolated from a genetically manipulated source Source: (gene. exp.) ![]() ![]() |
#5: Protein | Mass: 93010.273 Da / Num. of mol.: 1 Source method: isolated from a genetically manipulated source Source: (gene. exp.) ![]() ![]() |
#6: Protein | Mass: 81411.875 Da / Num. of mol.: 1 Source method: isolated from a genetically manipulated source Source: (gene. exp.) ![]() ![]() |
-DNA polymerase epsilon ... , 2 types, 2 molecules AB
#7: Protein | Mass: 59600.887 Da / Num. of mol.: 1 Source method: isolated from a genetically manipulated source Source: (gene. exp.) ![]() ![]() |
---|---|
#8: Protein | Mass: 261855.266 Da / Num. of mol.: 1 Source method: isolated from a genetically manipulated source Source: (gene. exp.) ![]() ![]() References: UniProt: Q07864, DNA-directed DNA polymerase, Hydrolases; Acting on ester bonds; Exodeoxyribonucleases producing 5'-phosphomonoesters |
-Protein , 5 types, 7 molecules CHIJKLQ
#9: Protein | Mass: 66016.891 Da / Num. of mol.: 1 Source method: isolated from a genetically manipulated source Source: (gene. exp.) ![]() ![]() | ||||||
---|---|---|---|---|---|---|---|
#14: Protein | Mass: 130098.148 Da / Num. of mol.: 3 Source method: isolated from a genetically manipulated source Source: (gene. exp.) ![]() ![]() #15: Protein | | Mass: 138903.031 Da / Num. of mol.: 1 Source method: isolated from a genetically manipulated source Source: (gene. exp.) ![]() ![]() #16: Protein | | Mass: 34600.223 Da / Num. of mol.: 1 Source method: isolated from a genetically manipulated source Source: (gene. exp.) ![]() ![]() #19: Protein | | Mass: 155184.703 Da / Num. of mol.: 1 Source method: isolated from a genetically manipulated source Source: (gene. exp.) ![]() ![]() |
-DNA replication complex GINS protein ... , 4 types, 4 molecules DEFG
#10: Protein | Mass: 23022.469 Da / Num. of mol.: 1 Source method: isolated from a genetically manipulated source Source: (gene. exp.) ![]() ![]() |
---|---|
#11: Protein | Mass: 21453.713 Da / Num. of mol.: 1 Source method: isolated from a genetically manipulated source Source: (gene. exp.) ![]() ![]() |
#12: Protein | Mass: 24562.611 Da / Num. of mol.: 1 Source method: isolated from a genetically manipulated source Source: (gene. exp.) ![]() ![]() |
#13: Protein | Mass: 30114.236 Da / Num. of mol.: 1 Source method: isolated from a genetically manipulated source Source: (gene. exp.) ![]() ![]() |
-DNA chain , 2 types, 2 molecules MN
#17: DNA chain | Mass: 26396.836 Da / Num. of mol.: 1 / Source method: obtained synthetically / Source: (synth.) ![]() |
---|---|
#18: DNA chain | Mass: 16407.533 Da / Num. of mol.: 1 Source method: isolated from a genetically manipulated source Source: (gene. exp.) ![]() ![]() |
-Non-polymers , 4 types, 12 molecules ![](data/chem/img/ZN.gif)
![](data/chem/img/MG.gif)
![](data/chem/img/ANP.gif)
![](data/chem/img/SO4.gif)
![](data/chem/img/MG.gif)
![](data/chem/img/ANP.gif)
![](data/chem/img/SO4.gif)
#20: Chemical | ChemComp-ZN / #21: Chemical | #22: Chemical | #23: Chemical | ChemComp-SO4 / | |
---|
-Details
Has ligand of interest | Y |
---|
-Experimental details
-Experiment
Experiment | Method: ELECTRON MICROSCOPY |
---|---|
EM experiment | Aggregation state: PARTICLE / 3D reconstruction method: single particle reconstruction |
-
Sample preparation
Component | Name: Core human replisome / Type: COMPLEX Entity ID: #8, #7, #3, #5-#6, #2, #1, #4, #10-#13, #9, #15-#17, #14, #18-#19 Source: RECOMBINANT |
---|---|
Molecular weight | Experimental value: NO |
Source (natural) | Organism: ![]() |
Source (recombinant) | Organism: ![]() |
Buffer solution | pH: 7.8 |
Specimen | Embedding applied: NO / Shadowing applied: NO / Staining applied: NO / Vitrification applied: YES |
Specimen support | Grid material: COPPER / Grid mesh size: 400 divisions/in. / Grid type: Quantifoil R2/2 |
Vitrification | Cryogen name: ETHANE |
-
Electron microscopy imaging
Experimental equipment | ![]() Model: Titan Krios / Image courtesy: FEI Company |
---|---|
Microscopy | Model: FEI TITAN KRIOS |
Electron gun | Electron source: ![]() |
Electron lens | Mode: BRIGHT FIELD / Nominal defocus max: 3500 nm / Nominal defocus min: 1500 nm / Cs: 0.1 mm |
Specimen holder | Specimen holder model: FEI TITAN KRIOS AUTOGRID HOLDER |
Image recording | Electron dose: 39.8 e/Å2 / Detector mode: COUNTING / Film or detector model: GATAN K2 SUMMIT (4k x 4k) |
Image scans | Movie frames/image: 40 |
-
Processing
EM software |
| ||||||||||||||||||||||||
---|---|---|---|---|---|---|---|---|---|---|---|---|---|---|---|---|---|---|---|---|---|---|---|---|---|
CTF correction | Type: PHASE FLIPPING AND AMPLITUDE CORRECTION | ||||||||||||||||||||||||
Particle selection | Num. of particles selected: 490000 | ||||||||||||||||||||||||
3D reconstruction | Resolution: 3.2 Å / Resolution method: FSC 0.143 CUT-OFF / Num. of particles: 110000 / Algorithm: FOURIER SPACE / Symmetry type: POINT | ||||||||||||||||||||||||
Atomic model building | Space: REAL | ||||||||||||||||||||||||
Refine LS restraints |
|