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Yorodumi- PDB-8ppv: Intermediate conformer of Pyrococcus abyssi DNA polymerase D (Pol... -
+Open data
-Basic information
Entry | Database: PDB / ID: 8ppv | ||||||
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Title | Intermediate conformer of Pyrococcus abyssi DNA polymerase D (PolD) bound to a primer/template substrate containing three consecutive mismatches | ||||||
Components |
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Keywords | DNA BINDING PROTEIN / Polymerase / DNA / Replication / PolD / Archaea / Editing / Proofreading / Exonuclease / Nuclease | ||||||
Function / homology | Function and homology information exodeoxyribonuclease I / single-stranded DNA 3'-5' DNA exonuclease activity / DNA catabolic process / DNA strand elongation involved in DNA replication / DNA polymerase processivity factor activity / leading strand elongation / regulation of DNA replication / mismatch repair / translesion synthesis / DNA-directed DNA polymerase ...exodeoxyribonuclease I / single-stranded DNA 3'-5' DNA exonuclease activity / DNA catabolic process / DNA strand elongation involved in DNA replication / DNA polymerase processivity factor activity / leading strand elongation / regulation of DNA replication / mismatch repair / translesion synthesis / DNA-directed DNA polymerase / DNA-directed DNA polymerase activity / DNA binding Similarity search - Function | ||||||
Biological species | Pyrococcus abyssi GE5 (archaea) | ||||||
Method | ELECTRON MICROSCOPY / single particle reconstruction / cryo EM / Resolution: 3.02 Å | ||||||
Authors | Betancurt-Anzola, L. / Martinez-Carranza, M. / Zatopek, K.M. / Gardner, A.F. / Sauguet, L. | ||||||
Funding support | France, 1items
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Citation | Journal: Nat Commun / Year: 2023 Title: Molecular basis for proofreading by the unique exonuclease domain of Family-D DNA polymerases. Authors: Leonardo Betancurt-Anzola / Markel Martínez-Carranza / Marc Delarue / Kelly M Zatopek / Andrew F Gardner / Ludovic Sauguet / Abstract: Replicative DNA polymerases duplicate entire genomes at high fidelity. This feature is shared among the three domains of life and is facilitated by their dual polymerase and exonuclease activities. ...Replicative DNA polymerases duplicate entire genomes at high fidelity. This feature is shared among the three domains of life and is facilitated by their dual polymerase and exonuclease activities. Family D replicative DNA polymerases (PolD), found exclusively in Archaea, contain an unusual RNA polymerase-like catalytic core, and a unique Mre11-like proofreading active site. Here, we present cryo-EM structures of PolD trapped in a proofreading mode, revealing an unanticipated correction mechanism that extends the repertoire of protein domains known to be involved in DNA proofreading. Based on our experimental structures, mutants of PolD were designed and their contribution to mismatch bypass and exonuclease kinetics was determined. This study sheds light on the convergent evolution of structurally distinct families of DNA polymerases, and the domain acquisition and exchange mechanism that occurred during the evolution of the replisome in the three domains of life. | ||||||
History |
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-Structure visualization
Structure viewer | Molecule: MolmilJmol/JSmol |
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-Downloads & links
-Download
PDBx/mmCIF format | 8ppv.cif.gz | 505.7 KB | Display | PDBx/mmCIF format |
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PDB format | pdb8ppv.ent.gz | 396.6 KB | Display | PDB format |
PDBx/mmJSON format | 8ppv.json.gz | Tree view | PDBx/mmJSON format | |
Others | Other downloads |
-Validation report
Arichive directory | https://data.pdbj.org/pub/pdb/validation_reports/pp/8ppv ftp://data.pdbj.org/pub/pdb/validation_reports/pp/8ppv | HTTPS FTP |
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-Related structure data
Related structure data | 17817MC 8pptC 8ppuC M: map data used to model this data C: citing same article (ref.) |
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Similar structure data | Similarity search - Function & homologyF&H Search |
-Links
-Assembly
Deposited unit |
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1 |
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-Components
-DNA polymerase ... , 2 types, 4 molecules ACDE
#1: Protein | Mass: 74009.742 Da / Num. of mol.: 1 Source method: isolated from a genetically manipulated source Source: (gene. exp.) Pyrococcus abyssi GE5 (archaea) / Gene: polB, PYRAB01210, PAB2266 / Production host: Escherichia coli BL21(DE3) (bacteria) References: UniProt: Q9V2F3, DNA-directed DNA polymerase, exodeoxyribonuclease I |
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#3: Protein | Mass: 29471.764 Da / Num. of mol.: 3 Source method: isolated from a genetically manipulated source Source: (gene. exp.) Pyrococcus abyssi GE5 (archaea) / Gene: pcn, PYRAB13790, PAB1465 / Production host: Escherichia coli BL21(DE3) (bacteria) / References: UniProt: Q9UYX8 |
-Protein , 1 types, 1 molecules B
#2: Protein | Mass: 144418.969 Da / Num. of mol.: 1 Source method: isolated from a genetically manipulated source Source: (gene. exp.) Pyrococcus abyssi GE5 (archaea) / Production host: Escherichia coli BL21(DE3) (bacteria) |
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-DNA chain , 2 types, 2 molecules PT
#4: DNA chain | Mass: 6512.451 Da / Num. of mol.: 1 / Source method: obtained synthetically / Source: (synth.) Pyrococcus abyssi GE5 (archaea) |
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#5: DNA chain | Mass: 7717.934 Da / Num. of mol.: 1 / Source method: obtained synthetically / Source: (synth.) Pyrococcus abyssi GE5 (archaea) |
-Non-polymers , 2 types, 6 molecules
#6: Chemical | #7: Chemical | |
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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 |
-Sample preparation
Component | Name: Binary complex of PolD bound to PCNA and a primer/template duplex containing three consecutive mismatches Type: COMPLEX / Entity ID: #1-#2, #4, #3 / Source: RECOMBINANT |
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Source (natural) | Organism: Pyrococcus abyssi GE5 (archaea) |
Source (recombinant) | Organism: Escherichia coli BL21(DE3) (bacteria) |
Buffer solution | pH: 8 |
Specimen | Embedding applied: NO / Shadowing applied: NO / Staining applied: NO / Vitrification applied: YES |
Vitrification | Cryogen name: ETHANE |
-Electron microscopy imaging
Experimental equipment | Model: Titan Krios / Image courtesy: FEI Company |
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Microscopy | Model: TFS 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: 1000 nm |
Image recording | Electron dose: 40 e/Å2 / Film or detector model: GATAN K3 BIOQUANTUM (6k x 4k) |
-Processing
CTF correction | Type: PHASE FLIPPING AND AMPLITUDE CORRECTION | ||||||||||||||||||||||||
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3D reconstruction | Resolution: 3.02 Å / Resolution method: FSC 0.143 CUT-OFF / Num. of particles: 299325 / Symmetry type: POINT | ||||||||||||||||||||||||
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