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- PDB-5n8a: Structure of RPA70N in complex with PrimPol (fragment 480-560) -

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Basic information

Entry
Database: PDB / ID: 5n8a
TitleStructure of RPA70N in complex with PrimPol (fragment 480-560)
Components
  • DNA-directed primase/polymerase protein
  • Replication protein A 70 kDa DNA-binding subunit
KeywordsPROTEIN BINDING / Complex / Replication / Basic cleft
Function / homology
Function and homology information


DNA primase AEP / protein localization to chromosome / DNA replication factor A complex / R-loop processing / mitochondrial DNA replication / single-stranded telomeric DNA binding / lateral element / G-rich strand telomeric DNA binding / chromatin-protein adaptor activity / protein localization to site of double-strand break ...DNA primase AEP / protein localization to chromosome / DNA replication factor A complex / R-loop processing / mitochondrial DNA replication / single-stranded telomeric DNA binding / lateral element / G-rich strand telomeric DNA binding / chromatin-protein adaptor activity / protein localization to site of double-strand break / Removal of the Flap Intermediate / Mismatch repair (MMR) directed by MSH2:MSH3 (MutSbeta) / Mismatch repair (MMR) directed by MSH2:MSH6 (MutSalpha) / Removal of the Flap Intermediate from the C-strand / DNA replication, synthesis of primer / HDR through Single Strand Annealing (SSA) / mitochondrial DNA repair / Impaired BRCA2 binding to RAD51 / replication fork processing / hemopoiesis / site of DNA damage / Presynaptic phase of homologous DNA pairing and strand exchange / PCNA-Dependent Long Patch Base Excision Repair / Activation of the pre-replicative complex / Regulation of HSF1-mediated heat shock response / HSF1 activation / error-prone translesion synthesis / telomere maintenance via telomerase / translesion synthesis / mismatch repair / SUMOylation of DNA damage response and repair proteins / Activation of ATR in response to replication stress / response to UV / Translesion synthesis by REV1 / Translesion synthesis by POLK / Translesion synthesis by POLI / Gap-filling DNA repair synthesis and ligation in GG-NER / DNA-directed RNA polymerase complex / homeostasis of number of cells within a tissue / telomere maintenance / Fanconi Anemia Pathway / male germ cell nucleus / Termination of translesion DNA synthesis / replication fork / Recognition of DNA damage by PCNA-containing replication complex / meiotic cell cycle / Translesion Synthesis by POLH / double-strand break repair via homologous recombination / nucleotide-excision repair / HDR through Homologous Recombination (HRR) / Dual Incision in GG-NER / Formation of Incision Complex in GG-NER / G2/M DNA damage checkpoint / PML body / base-excision repair / Meiotic recombination / Dual incision in TC-NER / Gap-filling DNA repair synthesis and ligation in TC-NER / DNA-templated DNA replication / DNA-directed RNA polymerase activity / site of double-strand break / single-stranded DNA binding / Processing of DNA double-strand break ends / manganese ion binding / Regulation of TP53 Activity through Phosphorylation / DNA recombination / damaged DNA binding / in utero embryonic development / DNA-directed DNA polymerase / DNA-directed DNA polymerase activity / chromosome, telomeric region / DNA replication / mitochondrial matrix / DNA repair / positive regulation of cell population proliferation / DNA damage response / chromatin binding / mitochondrion / zinc ion binding / nucleoplasm / nucleus
Similarity search - Function
DNA-directed primase/polymerase protein / Herpesviridae UL52/UL70 DNA primase / Replication factor-A protein 1, N-terminal domain / Replication factor A protein 1 / Replication factor-A protein 1, N-terminal / Replication protein A, OB domain / Replication protein A OB domain / : / Replication factor A, C-terminal / Replication factor-A C terminal domain ...DNA-directed primase/polymerase protein / Herpesviridae UL52/UL70 DNA primase / Replication factor-A protein 1, N-terminal domain / Replication factor A protein 1 / Replication factor-A protein 1, N-terminal / Replication protein A, OB domain / Replication protein A OB domain / : / Replication factor A, C-terminal / Replication factor-A C terminal domain / DNA primase, small subunit / DNA primase small subunit / OB-fold nucleic acid binding domain, AA-tRNA synthetase-type / OB-fold nucleic acid binding domain / Nucleic acid-binding proteins / OB fold (Dihydrolipoamide Acetyltransferase, E2P) / Nucleic acid-binding, OB-fold / Beta Barrel / Mainly Beta
Similarity search - Domain/homology
Replication protein A 70 kDa DNA-binding subunit / DNA-directed primase/polymerase protein
Similarity search - Component
Biological speciesHomo sapiens (human)
MethodX-RAY DIFFRACTION / SYNCHROTRON / MOLECULAR REPLACEMENT / molecular replacement / Resolution: 1.28 Å
AuthorsBrissett, N.C. / Doherty, A.J.
Funding support United Kingdom, 3items
OrganizationGrant numberCountry
Biotechnology and Biological Sciences Research CouncilBB/H019723/1 United Kingdom
Biotechnology and Biological Sciences Research CouncilBB/M008800/1 United Kingdom
Biotechnology and Biological Sciences Research CouncilBB/M004236/1 United Kingdom
CitationJournal: Nat Commun / Year: 2017
Title: Molecular basis for PrimPol recruitment to replication forks by RPA.
Authors: Guilliam, T.A. / Brissett, N.C. / Ehlinger, A. / Keen, B.A. / Kolesar, P. / Taylor, E.M. / Bailey, L.J. / Lindsay, H.D. / Chazin, W.J. / Doherty, A.J.
History
DepositionFeb 23, 2017Deposition site: PDBE / Processing site: PDBE
Revision 1.0Jun 7, 2017Provider: repository / Type: Initial release
Revision 1.1Aug 30, 2017Group: Author supporting evidence / Category: pdbx_audit_support / Item: _pdbx_audit_support.funding_organization
Revision 1.2Jan 17, 2024Group: Data collection / Database references / Refinement description
Category: chem_comp_atom / chem_comp_bond ...chem_comp_atom / chem_comp_bond / database_2 / pdbx_initial_refinement_model
Item: _database_2.pdbx_DOI / _database_2.pdbx_database_accession

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Structure visualization

Structure viewerMolecule:
MolmilJmol/JSmol

Downloads & links

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Assembly

Deposited unit
X: DNA-directed primase/polymerase protein
A: Replication protein A 70 kDa DNA-binding subunit


Theoretical massNumber of molelcules
Total (without water)24,5152
Polymers24,5152
Non-polymers00
Water2,522140
1


  • Idetical with deposited unit
  • defined by author&software
  • Evidence: gel filtration, light scattering, isothermal titration calorimetry
TypeNameSymmetry operationNumber
identity operation1_555x,y,z1
Buried area980 Å2
ΔGint-3 kcal/mol
Surface area6830 Å2
MethodPISA
Unit cell
Length a, b, c (Å)38.050, 53.490, 53.900
Angle α, β, γ (deg.)90.000, 90.000, 90.000
Int Tables number19
Space group name H-MP212121

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Components

#1: Protein DNA-directed primase/polymerase protein / hPrimpol1 / Coiled-coil domain-containing protein 111


Mass: 11161.771 Da / Num. of mol.: 1 / Fragment: UNP residues 479-559
Source method: isolated from a genetically manipulated source
Source: (gene. exp.) Homo sapiens (human) / Gene: PRIMPOL, CCDC111 / Plasmid: pET28a / Production host: Escherichia coli BL21 (bacteria)
References: UniProt: Q96LW4, Transferases; Transferring phosphorus-containing groups; Nucleotidyltransferases
#2: Protein Replication protein A 70 kDa DNA-binding subunit / RP-A p70 / Replication factor A protein 1 / RF-A protein 1 / Single-stranded DNA-binding protein


Mass: 13353.598 Da / Num. of mol.: 1 / Mutation: E7R
Source method: isolated from a genetically manipulated source
Source: (gene. exp.) Homo sapiens (human) / Gene: RPA1, REPA1, RPA70 / Plasmid: pET15b / Production host: Escherichia coli BL21 (bacteria) / References: UniProt: P27694
#3: Water ChemComp-HOH / water


Mass: 18.015 Da / Num. of mol.: 140 / Source method: isolated from a natural source / Formula: H2O

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Experimental details

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Experiment

ExperimentMethod: X-RAY DIFFRACTION / Number of used crystals: 1

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Sample preparation

CrystalDensity Matthews: 1.88 Å3/Da / Density % sol: 34.66 %
Crystal growTemperature: 293.15 K / Method: vapor diffusion, sitting drop / pH: 6 / Details: 0.2M imidazole malate, 30% w/v PEG 4000

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Data collection

DiffractionMean temperature: 100 K
Diffraction sourceSource: SYNCHROTRON / Site: Diamond / Beamline: I04 / Wavelength: 0.91407 Å
DetectorType: DECTRIS PILATUS 6M-F / Detector: PIXEL / Date: Nov 30, 2014
RadiationProtocol: SINGLE WAVELENGTH / Monochromatic (M) / Laue (L): M / Scattering type: x-ray
Radiation wavelengthWavelength: 0.91407 Å / Relative weight: 1
ReflectionResolution: 1.28→16.247 Å / Num. all: 29019 / Num. obs: 351864 / % possible obs: 99.7 % / Observed criterion σ(I): -3 / Redundancy: 6.431 % / CC1/2: 1 / Rmerge(I) obs: 0.043 / Rrim(I) all: 0.047 / Χ2: 0.984 / Net I/σ(I): 20.01 / Num. measured all: 351864 / Scaling rejects: 8
Reflection shell

Diffraction-ID: 1

Resolution (Å)Redundancy (%)Rmerge(I) obsMean I/σ(I) obsNum. unique obsCC1/2Rrim(I) all% possible all
1.28-1.313.8450.6041.8740280.6770.797.7
1.31-1.355.0430.5222.6139150.7990.58399.5
1.35-1.395.8620.4693.2538100.8610.515100
1.39-1.436.7210.3744.4637480.9360.405100
1.43-1.486.7580.3075.5735970.9490.333100
1.48-1.536.7760.241735340.970.26199.9
1.53-1.596.7940.1888.8133400.9820.203100
1.59-1.656.8510.15710.632870.9880.169100
1.65-1.736.8970.12613.0831280.9910.137100
1.73-1.816.8420.10315.9729520.9950.111100
1.81-1.916.9080.07521.6428390.9970.082100
1.91-2.026.8540.05628.226670.9980.061100
2.02-2.166.980.04634.8425100.9990.049100
2.16-2.346.9710.04140.3723780.9990.044100
2.34-2.566.8220.03544.5721450.9990.038100
2.56-2.867.1170.03251.1119490.9990.034100
2.86-3.36.8630.02759.83171710.029100
3.3-4.056.7680.02170.58146010.023100
4.05-5.726.8130.01976.87111410.02100
5.72-16.2476.9310.01973.9559510.0295

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Phasing

PhasingMethod: molecular replacement
Phasing MRModel details: Phaser MODE: MR_AUTO
Highest resolutionLowest resolution
Rotation1.28 Å16.25 Å
Translation1.28 Å16.25 Å

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Processing

Software
NameVersionClassification
PHENIXrefinement
XDSdata reduction
XSCALEdata scaling
PHASER2.5.6phasing
Cootmodel building
PDB_EXTRACT3.22data extraction
RefinementMethod to determine structure: MOLECULAR REPLACEMENT
Starting model: 4IPC

4ipc


Resolution: 1.28→16.247 Å / SU ML: 0.12 / Cross valid method: FREE R-VALUE / σ(F): 1.34 / Phase error: 17.12
RfactorNum. reflection% reflection
Rfree0.1785 1410 4.87 %
Rwork0.1537 --
obs0.1549 28966 99.8 %
Solvent computationShrinkage radii: 0.9 Å / VDW probe radii: 1.11 Å
Displacement parametersBiso max: 65.99 Å2 / Biso mean: 21.1988 Å2 / Biso min: 8.69 Å2
Refinement stepCycle: final / Resolution: 1.28→16.247 Å
ProteinNucleic acidLigandSolventTotal
Num. atoms999 0 0 140 1139
Biso mean---32.95 -
Num. residues----130
Refine LS restraints
Refine-IDTypeDev idealNumber
X-RAY DIFFRACTIONf_bond_d0.0071095
X-RAY DIFFRACTIONf_angle_d0.9121502
X-RAY DIFFRACTIONf_chiral_restr0.086185
X-RAY DIFFRACTIONf_plane_restr0.005195
X-RAY DIFFRACTIONf_dihedral_angle_d18.65444
LS refinement shell

Refine-ID: X-RAY DIFFRACTION / Rfactor Rfree error: 0 / Total num. of bins used: 10

Resolution (Å)Rfactor RfreeNum. reflection RfreeRfactor RworkNum. reflection RworkNum. reflection all% reflection obs (%)
1.28-1.32580.22361310.18832684281598
1.3258-1.37880.2151660.159726712837100
1.3788-1.44150.19531310.141627322863100
1.4415-1.51750.18611220.137227562878100
1.5175-1.61250.16671320.129327382870100
1.6125-1.73680.15091170.134527722889100
1.7368-1.91140.17151530.137727312884100
1.9114-2.18740.17181360.133227702906100
2.1874-2.75370.15241820.16227672949100
2.7537-16.24790.20141400.168929353075100

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