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- PDB-6hms: Cryo-EM map of DNA polymerase D from Pyrococcus abyssi in complex... -

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Entry
Database: PDB / ID: 6hms
TitleCryo-EM map of DNA polymerase D from Pyrococcus abyssi in complex with DNA
Components
  • (DNA polymerase II ...) x 2
  • DNA (5'-D(*GP*AP*GP*AP*CP*GP*GP*GP*CP*CP*GP*CP*GP*TP*C)-3')
  • DNA (5'-D(P*TP*GP*AP*CP*GP*CP*GP*GP*CP*CP*CP*GP*TP*CP*TP*C)-3')
KeywordsREPLICATION / DNA / polymerase D / Pyrococcus
Function / homologyDNA polymerase alpha/epsilon subunit B / Intein N-terminal splicing region / Intein C-terminal splicing motif profile. / Intein N-terminal splicing motif profile. / DNA polymerase II large subunit DP2 / Calcineurin-like phosphoesterase / Hint domain superfamily / Intein C-terminal splicing region / DNA polymerase delta/II small subunit family / DNA polymerase II large subunit DP2, N-terminal ...DNA polymerase alpha/epsilon subunit B / Intein N-terminal splicing region / Intein C-terminal splicing motif profile. / Intein N-terminal splicing motif profile. / DNA polymerase II large subunit DP2 / Calcineurin-like phosphoesterase / Hint domain superfamily / Intein C-terminal splicing region / DNA polymerase delta/II small subunit family / DNA polymerase II large subunit DP2, N-terminal / Nucleic acid-binding, OB-fold / DNA polymerase II small subunit, archaeal / Calcineurin-like phosphoesterase domain, ApaH type / DNA polymerase II large subunit DP2 / Hint domain N-terminal / exodeoxyribonuclease I / exodeoxyribonuclease I activity / DNA catabolic process, exonucleolytic / intein-mediated protein splicing / 3'-5' exonuclease activity / DNA-dependent DNA replication / DNA-directed DNA polymerase / DNA-directed DNA polymerase activity / DNA binding / DNA polymerase II small subunit / DNA polymerase II large subunit
Function and homology information
Specimen sourcePyrococcus abyssi (archaea)
MethodELECTRON MICROSCOPY / single particle reconstruction / cryo EM / 7.1 Å resolution
AuthorsRaia, P. / Carroni, M. / Sauguet, L.
CitationJournal: PLoS Biol. / Year: 2019
Title: Structure of the DP1-DP2 PolD complex bound with DNA and its implications for the evolutionary history of DNA and RNA polymerases.
Authors: Pierre Raia / Marta Carroni / Etienne Henry / Gérard Pehau-Arnaudet / Sébastien Brûlé / Pierre Béguin / Ghislaine Henneke / Erik Lindahl / Marc Delarue / Ludovic Sauguet
Validation Report
SummaryFull reportAbout validation report
DateDeposition: Sep 12, 2018 / Release: Jan 30, 2019

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Structure visualization

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  • Superimposition on EM map
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Structure viewerMolecule:
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Assembly

Deposited unit
A: DNA polymerase II small subunit
B: DNA polymerase II large subunit,DNA polymerase II large subunit
P: DNA (5'-D(*GP*AP*GP*AP*CP*GP*GP*GP*CP*CP*GP*CP*GP*TP*C)-3')
T: DNA (5'-D(P*TP*GP*AP*CP*GP*CP*GP*GP*CP*CP*CP*GP*TP*CP*TP*C)-3')
hetero molecules


Theoretical massNumber of molelcules
Total (without water)199,3029
Polyers198,9844
Non-polymers3175
Water0
1


TypeNameSymmetry operationNumber
identity operation1_5551

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Components

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DNA polymerase II ... , 2 types, 2 molecules AB

#1: Protein/peptide DNA polymerase II small subunit / / Pol II / Exodeoxyribonuclease small subunit / Coordinate model: Cα atoms only


Mass: 45078.012 Da / Num. of mol.: 1 / Source: (gene. exp.) Pyrococcus abyssi (archaea) / Gene: polB, PYRAB01210, PAB2266 / Production host: Escherichia coli (E. coli)
References: UniProt: Q9V2F3, DNA-directed DNA polymerase, exodeoxyribonuclease I
#2: Protein/peptide DNA polymerase II large subunit,DNA polymerase II large subunit / / Pol II / Exodeoxyribonuclease large subunit / Coordinate model: Cα atoms only


Mass: 144418.969 Da / Num. of mol.: 1 / Source: (gene. exp.) Pyrococcus abyssi (archaea) / Gene: polC, PYRAB01200, PAB2404 / Production host: Escherichia coli (E. coli)
References: UniProt: Q9V2F4, DNA-directed DNA polymerase, exodeoxyribonuclease I

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DNA chain , 2 types, 2 molecules PT

#3: DNA chain DNA (5'-D(*GP*AP*GP*AP*CP*GP*GP*GP*CP*CP*GP*CP*GP*TP*C)-3')


Mass: 4635.998 Da / Num. of mol.: 1 / Source: (synth.) Pyrococcus abyssi (archaea)
#4: DNA chain DNA (5'-D(P*TP*GP*AP*CP*GP*CP*GP*GP*CP*CP*CP*GP*TP*CP*TP*C)-3')


Mass: 4851.129 Da / Num. of mol.: 1 / Source: (synth.) Pyrococcus abyssi (archaea)

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Non-polymers , 2 types, 5 molecules

#5: Chemical ChemComp-FE / FE (III) ION


Mass: 55.845 Da / Num. of mol.: 1 / Formula: Fe / Iron
#6: Chemical
ChemComp-ZN / ZINC ION


Mass: 65.409 Da / Num. of mol.: 4 / Formula: Zn / Zinc

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Experimental details

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Experiment

ExperimentMethod: ELECTRON MICROSCOPY
EM experimentAggregation state: PARTICLE / Reconstruction method: single particle reconstruction

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Sample preparation

Component
IDNameTypeEntity IDParent IDSource
1Complex between DNA polymerase D and DNA duplexCOMPLEX1, 2, 3, 40MULTIPLE SOURCES
2DNA polymerase DCOMPLEX1, 21RECOMBINANT
3DNACOMPLEX3, 41RECOMBINANT
Source (natural)
IDEntity assembly IDNcbi tax IDOrganism
2229292Pyrococcus abyssi (archaea)
3329292Pyrococcus abyssi (archaea)
Source (recombinant)
IDEntity assembly IDNcbi tax IDOrganism
22562Escherichia coli (E. coli)
3332630synthetic construct (others)
Buffer solutionpH: 8
SpecimenEmbedding applied: NO / Shadowing applied: NO / Staining applied: NO / Vitrification applied: YES
VitrificationCryogen name: ETHANE

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Electron microscopy imaging

Experimental equipment
Model: Titan Krios / Image courtesy: FEI Company
MicroscopyMicroscope model: FEI TITAN KRIOS
Electron gunElectron source: FIELD EMISSION GUN / Accelerating voltage: 300 kV / Illumination mode: FLOOD BEAM
Electron lensMode: BRIGHT FIELDBright-field microscopy
Image recordingElectron dose: 1 e/Å2 / Film or detector model: GATAN K2 QUANTUM (4k x 4k)

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Processing

SoftwareName: PHENIX / Version: 1.11.1_2575: / Classification: refinement
EM softwareName: cryoSPARC / Category: 3D reconstruction
CTF correctionType: PHASE FLIPPING AND AMPLITUDE CORRECTION
3D reconstructionResolution: 7.1 Å / Resolution method: FSC 0.143 CUT-OFF / Number of particles: 8774 / Symmetry type: POINT
Least-squares processHighest resolution: 7.1 Å
Refine LS restraints
Refine IDTypeDev idealNumber
ELECTRON MICROSCOPYf_bond_d0.00812947
ELECTRON MICROSCOPYf_angle_d1.37417637
ELECTRON MICROSCOPYf_dihedral_angle_d9.9647746
ELECTRON MICROSCOPYf_chiral_restr0.0731940
ELECTRON MICROSCOPYf_plane_restr0.0102164

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