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Yorodumi- PDB-8b9a: S. cerevisiae replisome + Ctf4, bound by pol alpha primase. Compl... -
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Open data
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Basic information
| Entry | Database: PDB / ID: 8b9a | ||||||
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| Title | S. cerevisiae replisome + Ctf4, bound by pol alpha primase. Complex engaged with a fork DNA substrate containing a 60 nucleotide lagging strand. | ||||||
Components |
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Keywords | REPLICATION / helicase / polymerase / pol alpha / priming | ||||||
| Function / homology | Function and homology informationInhibition of replication initiation of damaged DNA by RB1/E2F1 / establishment of sister chromatid cohesion / H3-H4 histone complex chaperone activity / Unwinding of DNA / maintenance of DNA repeat elements / regulation of nuclear cell cycle DNA replication / DNA replication initiation / replication fork arrest / Cul8-RING ubiquitin ligase complex / meiotic chromosome segregation ...Inhibition of replication initiation of damaged DNA by RB1/E2F1 / establishment of sister chromatid cohesion / H3-H4 histone complex chaperone activity / Unwinding of DNA / maintenance of DNA repeat elements / regulation of nuclear cell cycle DNA replication / DNA replication initiation / replication fork arrest / Cul8-RING ubiquitin ligase complex / meiotic chromosome segregation / RNA-templated DNA biosynthetic process / Processive synthesis on the lagging strand / Removal of the Flap Intermediate / MCM core complex / Assembly of the pre-replicative complex / Switching of origins to a post-replicative state / DNA strand elongation involved in mitotic DNA replication / GINS complex / MCM complex binding / mitotic DNA replication preinitiation complex assembly / nuclear DNA replication / Polymerase switching / premeiotic DNA replication / replication fork protection complex / pre-replicative complex assembly involved in nuclear cell cycle DNA replication / anaphase-promoting complex binding / alpha DNA polymerase:primase complex / Activation of the pre-replicative complex / mitotic DNA replication / DNA replication checkpoint signaling / CMG complex / establishment of mitotic sister chromatid cohesion / nuclear pre-replicative complex / Activation of ATR in response to replication stress / DNA replication preinitiation complex / MCM complex / lagging strand elongation / telomere capping / mitotic DNA replication checkpoint signaling / double-strand break repair via break-induced replication / mitotic DNA replication initiation / DNA replication, synthesis of primer / cellular response to osmotic stress / 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 / mitotic sister chromatid cohesion / DNA biosynthetic process / DNA strand elongation involved in DNA replication / nuclear chromosome / DNA synthesis involved in DNA repair / leading strand elongation / mitotic G2 DNA damage checkpoint signaling / nuclear replication fork / replication fork processing / DNA replication origin binding / DNA replication initiation / Ub-specific processing proteases / subtelomeric heterochromatin formation / telomere maintenance / nuclear periphery / DNA helicase activity / replication fork / meiotic cell cycle / transcription elongation by RNA polymerase II / helicase activity / DNA-templated DNA replication / Transferases; Transferring phosphorus-containing groups; Nucleotidyltransferases / DNA-directed RNA polymerase activity / nuclear envelope / heterochromatin formation / double-strand break repair / mitotic cell cycle / nucleosome assembly / single-stranded DNA binding / 4 iron, 4 sulfur cluster binding / DNA helicase / DNA-directed DNA polymerase / DNA-directed DNA polymerase activity / chromosome, telomeric region / DNA replication / protein stabilization / DNA repair / nucleotide binding / DNA damage response / chromatin binding / ATP hydrolysis activity / mitochondrion / DNA binding / zinc ion binding / nucleoplasm / ATP binding / metal ion binding / identical protein binding / nucleus / cytoplasm / cytosol Similarity search - Function | ||||||
| Biological species | ![]() | ||||||
| Method | ELECTRON MICROSCOPY / single particle reconstruction / cryo EM / Resolution: 3.5 Å | ||||||
Authors | Jones, M.L. / Yeeles, J.T.P. | ||||||
| Funding support | United Kingdom, 1items
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Citation | Journal: Mol Cell / Year: 2023Title: How Pol α-primase is targeted to replisomes to prime eukaryotic DNA replication. Authors: Morgan L Jones / Valentina Aria / Yasemin Baris / Joseph T P Yeeles / ![]() Abstract: During eukaryotic DNA replication, Pol α-primase generates primers at replication origins to start leading-strand synthesis and every few hundred nucleotides during discontinuous lagging-strand ...During eukaryotic DNA replication, Pol α-primase generates primers at replication origins to start leading-strand synthesis and every few hundred nucleotides during discontinuous lagging-strand replication. How Pol α-primase is targeted to replication forks to prime DNA synthesis is not fully understood. Here, by determining cryoelectron microscopy (cryo-EM) structures of budding yeast and human replisomes containing Pol α-primase, we reveal a conserved mechanism for the coordination of priming by the replisome. Pol α-primase binds directly to the leading edge of the CMG (CDC45-MCM-GINS) replicative helicase via a complex interaction network. The non-catalytic PRIM2/Pri2 subunit forms two interfaces with CMG that are critical for in vitro DNA replication and yeast cell growth. These interactions position the primase catalytic subunit PRIM1/Pri1 directly above the exit channel for lagging-strand template single-stranded DNA (ssDNA), revealing why priming occurs efficiently only on the lagging-strand template and elucidating a mechanism for Pol α-primase to overcome competition from RPA to initiate primer synthesis. | ||||||
| History |
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Structure visualization
| Structure viewer | Molecule: Molmil Jmol/JSmol |
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Downloads & links
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Download
| PDBx/mmCIF format | 8b9a.cif.gz | 3 MB | Display | PDBx/mmCIF format |
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| PDB format | pdb8b9a.ent.gz | Display | PDB format | |
| PDBx/mmJSON format | 8b9a.json.gz | Tree view | PDBx/mmJSON format | |
| Others | Other downloads |
-Validation report
| Summary document | 8b9a_validation.pdf.gz | 1.8 MB | Display | wwPDB validaton report |
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| Full document | 8b9a_full_validation.pdf.gz | 1.8 MB | Display | |
| Data in XML | 8b9a_validation.xml.gz | 188.9 KB | Display | |
| Data in CIF | 8b9a_validation.cif.gz | 315 KB | Display | |
| Arichive directory | https://data.pdbj.org/pub/pdb/validation_reports/b9/8b9a ftp://data.pdbj.org/pub/pdb/validation_reports/b9/8b9a | HTTPS FTP |
-Related structure data
| Related structure data | ![]() 15309MC ![]() 8b9bC ![]() 8b9cC ![]() 8b9dC M: map data used to model this data C: citing same article ( |
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| Similar structure data | Similarity search - Function & homology F&H Search |
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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: 111987.562 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 , 8 types, 10 molecules 5AGPSXYHLK
| #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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| #7: Protein | Mass: 62348.551 Da / Num. of mol.: 1 Source method: isolated from a genetically manipulated source Source: (gene. exp.) ![]() Gene: PRI2, YKL045W, YKL258 / Production host: ![]() |
| #13: 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: ![]() |
| #15: Protein | Mass: 126078.859 Da / Num. of mol.: 1 Source method: isolated from a genetically manipulated source Source: (gene. exp.) ![]() Gene: MRC1, YCL061C, YCL61C/YCL60C / Production host: ![]() |
| #18: Protein | Mass: 51827.090 Da / Num. of mol.: 1 Source method: isolated from a genetically manipulated source Source: (gene. exp.) ![]() Gene: PRI1, YIR008C, YIB8C / Production host: ![]() References: UniProt: P10363, Transferases; Transferring phosphorus-containing groups; Nucleotidyltransferases |
| #19: Protein | Mass: 141296.875 Da / Num. of mol.: 1 Source method: isolated from a genetically manipulated source Source: (gene. exp.) ![]() Gene: TOF1, YNL273W, N0636 / Production host: ![]() |
| #20: Protein | Mass: 36588.758 Da / Num. of mol.: 1 Source method: isolated from a genetically manipulated source Source: (gene. exp.) ![]() Gene: CSM3, YMR048W, YM9796.01 / Production host: ![]() |
| #21: Protein | Mass: 108610.148 Da / Num. of mol.: 3 Source method: isolated from a genetically manipulated source Source: (gene. exp.) ![]() Gene: CTF4, CHL15, POB1, YPR135W, P9659.7 / Production host: ![]() |
-DNA polymerase alpha ... , 2 types, 2 molecules BJ
| #8: Protein | Mass: 78865.938 Da / Num. of mol.: 1 Source method: isolated from a genetically manipulated source Source: (gene. exp.) ![]() Gene: POL12, YBL035C, YBL0414 / Production host: ![]() |
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| #14: Protein | Mass: 167027.766 Da / Num. of mol.: 1 Source method: isolated from a genetically manipulated source Source: (gene. exp.) ![]() Gene: POL1, CDC17, YNL102W, N2181 / Production host: ![]() |
-DNA replication complex GINS protein ... , 4 types, 4 molecules CDEF
| #9: 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: ![]() |
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| #10: 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: ![]() |
| #11: Protein | Mass: 24437.859 Da / Num. of mol.: 1 Source method: isolated from a genetically manipulated source Source: (gene. exp.) ![]() Gene: PSF3, YOL146W / Production host: ![]() |
| #12: 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: ![]() |
-DNA chain , 2 types, 2 molecules QR
| #16: DNA chain | Mass: 26092.645 Da / Num. of mol.: 1 / Source method: obtained synthetically / Source: (synth.) ![]() |
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| #17: DNA chain | Mass: 32046.451 Da / Num. of mol.: 1 / Source method: obtained synthetically / Source: (synth.) ![]() |
-Non-polymers , 3 types, 12 molecules 




| #22: Chemical | ChemComp-ANP / #23: Chemical | ChemComp-MG / #24: Chemical | ChemComp-ZN / |
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-Details
| Has ligand of interest | Y |
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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: S. cerevisiae replisome + Ctf4, bound by pol alpha primase. Complex engaged with a fork DNA substrate containing a 60 nucleotide lagging strand. Type: COMPLEX / Entity ID: #1-#13, #18, #21, #15-#17, #19-#20, #14 / Source: RECOMBINANT |
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| Source (natural) | Organism: ![]() |
| Source (recombinant) | Organism: ![]() |
| Buffer solution | pH: 7.6 |
| Specimen | Embedding applied: NO / Shadowing applied: NO / Staining applied: NO / Vitrification applied: YES |
| 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: FIELD EMISSION GUN / Accelerating voltage: 300 kV / Illumination mode: FLOOD BEAM |
| Electron lens | Mode: BRIGHT FIELD / Nominal defocus max: 3000 nm / Nominal defocus min: 1500 nm |
| Image recording | Electron dose: 39.2 e/Å2 / Film or detector model: GATAN K3 BIOQUANTUM (6k x 4k) |
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Processing
| EM software | Name: PHENIX / Version: 1.20_4459: / Category: model refinement | ||||||||||||||||||||||||
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| CTF correction | Type: PHASE FLIPPING AND AMPLITUDE CORRECTION | ||||||||||||||||||||||||
| 3D reconstruction | Resolution: 3.5 Å / Resolution method: FSC 0.143 CUT-OFF / Num. of particles: 54970 / Symmetry type: POINT | ||||||||||||||||||||||||
| Refinement | Cross valid method: NONE Stereochemistry target values: GeoStd + Monomer Library + CDL v1.2 | ||||||||||||||||||||||||
| Displacement parameters | Biso mean: 97.22 Å2 | ||||||||||||||||||||||||
| Refine LS restraints |
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