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- PDB-4q5v: Crystal structure of the catalytic core of human DNA polymerase a... -

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Basic information

Entry
Database: PDB / ID: 4q5v
TitleCrystal structure of the catalytic core of human DNA polymerase alpha in ternary complex with an RNA-primed DNA template and aphidicolin
Components
  • DNA polymerase alpha catalytic subunit
  • DNA template
  • RNA primer
KeywordsTRANSFERASE/DNA/RNA / B-family DNA polymerase / DNA replication / TRANSFERASE-DNA-RNA complex
Function / homology
Function and homology information


DNA replication initiation / Inhibition of replication initiation of damaged DNA by RB1/E2F1 / regulation of type I interferon production / Telomere C-strand synthesis initiation / alpha DNA polymerase:primase complex / Polymerase switching / Processive synthesis on the lagging strand / Removal of the Flap Intermediate / lagging strand elongation / mitotic DNA replication initiation ...DNA replication initiation / Inhibition of replication initiation of damaged DNA by RB1/E2F1 / regulation of type I interferon production / Telomere C-strand synthesis initiation / alpha DNA polymerase:primase complex / Polymerase switching / Processive synthesis on the lagging strand / Removal of the Flap Intermediate / lagging strand elongation / mitotic DNA replication initiation / DNA replication, synthesis of primer / Polymerase switching on the C-strand of the telomere / DNA strand elongation involved in DNA replication / DNA synthesis involved in DNA repair / G1/S-Specific Transcription / leading strand elongation / DNA replication origin binding / DNA replication initiation / Activation of the pre-replicative complex / Defective pyroptosis / double-strand break repair via nonhomologous end joining / nuclear matrix / nuclear envelope / single-stranded DNA binding / DNA-directed DNA polymerase / DNA-directed DNA polymerase activity / DNA replication / DNA repair / nucleotide binding / chromatin binding / protein kinase binding / chromatin / nucleolus / DNA binding / zinc ion binding / nucleoplasm / nucleus / cytosol
Similarity search - Function
OB fold (Dihydrolipoamide Acetyltransferase, E2P) - #730 / Alpha-Beta Plaits - #2820 / B family DNA polymerase, thumb domain / DNA polymerase alpha catalytic subunit, N-terminal domain / DNA polymerase alpha, zinc finger domain superfamily / DNA Polymerase alpha zinc finger / DNA polymerase alpha subunit p180 N terminal / Zinc finger, DNA-directed DNA polymerase, family B, alpha / DNA polymerase alpha catalytic subunit, catalytic domain / DNA polymerase family B, thumb domain ...OB fold (Dihydrolipoamide Acetyltransferase, E2P) - #730 / Alpha-Beta Plaits - #2820 / B family DNA polymerase, thumb domain / DNA polymerase alpha catalytic subunit, N-terminal domain / DNA polymerase alpha, zinc finger domain superfamily / DNA Polymerase alpha zinc finger / DNA polymerase alpha subunit p180 N terminal / Zinc finger, DNA-directed DNA polymerase, family B, alpha / DNA polymerase alpha catalytic subunit, catalytic domain / DNA polymerase family B, thumb domain / DNA-directed DNA polymerase, family B, multifunctional domain / DNA-directed DNA polymerase, family B, conserved site / DNA polymerase family B signature. / DNA polymerase family B / Topoisomerase I; Chain A, domain 4 / DNA polymerase family B, exonuclease domain / DNA-directed DNA polymerase, family B, exonuclease domain / DNA polymerase, palm domain superfamily / DNA polymerase type-B family / DNA-directed DNA polymerase, family B / Ribonuclease H-like superfamily/Ribonuclease H / OB fold (Dihydrolipoamide Acetyltransferase, E2P) / Nucleotidyltransferase; domain 5 / Ribonuclease H superfamily / Ribonuclease H-like superfamily / Alpha-Beta Plaits / DNA/RNA polymerase superfamily / Beta Barrel / 2-Layer Sandwich / Orthogonal Bundle / Mainly Beta / Mainly Alpha / Alpha Beta
Similarity search - Domain/homology
Chem-2ZE / DNA / DNA (> 10) / RNA / RNA (> 10) / DNA polymerase alpha catalytic subunit
Similarity search - Component
Biological speciesHomo sapiens (human)
MethodX-RAY DIFFRACTION / SYNCHROTRON / MOLECULAR REPLACEMENT / Resolution: 2.52 Å
AuthorsBaranovskiy, A.G. / Babayeva, N.D. / Suwa, Y. / Gu, J. / Tahirov, T.H.
CitationJournal: Nucleic Acids Res. / Year: 2014
Title: Structural basis for inhibition of DNA replication by aphidicolin.
Authors: Baranovskiy, A.G. / Babayeva, N.D. / Suwa, Y. / Gu, J. / Pavlov, Y.I. / Tahirov, T.H.
History
DepositionApr 17, 2014Deposition site: RCSB / Processing site: RCSB
Revision 1.0Nov 26, 2014Provider: repository / Type: Initial release
Revision 1.1Dec 10, 2014Group: Database references
Revision 1.2Jan 7, 2015Group: Database references
Revision 1.3Feb 28, 2024Group: Data collection / Database references ...Data collection / Database references / Derived calculations / Structure summary
Category: chem_comp / chem_comp_atom ...chem_comp / chem_comp_atom / chem_comp_bond / database_2 / entity / pdbx_entity_nonpoly / struct_ref_seq_dif / struct_sheet / struct_site
Item: _chem_comp.name / _database_2.pdbx_DOI ..._chem_comp.name / _database_2.pdbx_DOI / _database_2.pdbx_database_accession / _entity.pdbx_description / _pdbx_entity_nonpoly.name / _struct_ref_seq_dif.details / _struct_sheet.number_strands / _struct_site.pdbx_auth_asym_id / _struct_site.pdbx_auth_comp_id / _struct_site.pdbx_auth_seq_id

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Structure visualization

Structure viewerMolecule:
MolmilJmol/JSmol

Downloads & links

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Assembly

Deposited unit
A: DNA polymerase alpha catalytic subunit
B: RNA primer
C: DNA template
E: DNA polymerase alpha catalytic subunit
F: RNA primer
G: DNA template
hetero molecules


Theoretical massNumber of molelcules
Total (without water)231,7338
Polymers231,0566
Non-polymers6772
Water2,720151
1
A: DNA polymerase alpha catalytic subunit
B: RNA primer
C: DNA template
hetero molecules


Theoretical massNumber of molelcules
Total (without water)115,8674
Polymers115,5283
Non-polymers3381
Water543
TypeNameSymmetry operationNumber
identity operation1_555x,y,z1
Buried area4930 Å2
ΔGint-29 kcal/mol
Surface area42450 Å2
MethodPISA
2
E: DNA polymerase alpha catalytic subunit
F: RNA primer
G: DNA template
hetero molecules


Theoretical massNumber of molelcules
Total (without water)115,8674
Polymers115,5283
Non-polymers3381
Water543
TypeNameSymmetry operationNumber
identity operation1_555x,y,z1
Buried area4960 Å2
ΔGint-28 kcal/mol
Surface area42220 Å2
MethodPISA
Unit cell
Length a, b, c (Å)105.051, 116.432, 233.026
Angle α, β, γ (deg.)90.00, 90.00, 90.00
Int Tables number19
Space group name H-MP212121

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Components

#1: Protein DNA polymerase alpha catalytic subunit / DNA polymerase alpha catalytic subunit p180


Mass: 105583.805 Da / Num. of mol.: 2
Fragment: Human DNA polymerase apha catalytic core domain residues 336-1257
Mutation: V516A
Source method: isolated from a genetically manipulated source
Source: (gene. exp.) Homo sapiens (human) / Gene: POLA1, POLA / Plasmid: pFastBac1 / Production host: Spodoptera frugiperda (fall armyworm) / Strain (production host): SF-21 / References: UniProt: P09884, DNA-directed DNA polymerase
#2: RNA chain RNA primer


Mass: 3521.170 Da / Num. of mol.: 2 / Source method: obtained synthetically
#3: DNA chain DNA template


Mass: 6423.160 Da / Num. of mol.: 2 / Source method: obtained synthetically
#4: Chemical ChemComp-2ZE / (3R,4R,4aR,6aS,8R,9R,11aS,11bS)-4,9-bis(hydroxymethyl)-4,11b-dimethyltetradecahydro-8,11a-methanocyclohepta[a]naphthale ne-3,9-diol / Aphidicolin


Mass: 338.482 Da / Num. of mol.: 2 / Source method: obtained synthetically / Formula: C20H34O4
#5: Water ChemComp-HOH / water


Mass: 18.015 Da / Num. of mol.: 151 / 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: 3.08 Å3/Da / Density % sol: 60.1 %
Crystal growTemperature: 295 K / Method: vapor diffusion, sitting drop / pH: 6.5
Details: 100 mM KCl, 12.5 mM MgCl2, 25 mM Na-Cacodylate, pH 6.5, 6 % 2-propanol, 2 mM TCEP, VAPOR DIFFUSION, SITTING DROP, temperature 295 K

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Data collection

DiffractionMean temperature: 100 K
Diffraction sourceSource: SYNCHROTRON / Site: APS / Beamline: 24-ID-C
DetectorType: DECTRIS PILATUS 6M-F / Detector: PIXEL
RadiationProtocol: SINGLE WAVELENGTH / Monochromatic (M) / Laue (L): M / Scattering type: x-ray
Radiation wavelengthRelative weight: 1
ReflectionResolution: 2.5→100 Å / Num. obs: 94074 / % possible obs: 97.3 % / Observed criterion σ(F): 0 / Observed criterion σ(I): 0 / Redundancy: 5 % / Biso Wilson estimate: 41.9 Å2 / Rmerge(I) obs: 0.042

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Processing

Software
NameClassification
CNSrefinement
HKL-2000data reduction
HKL-2000data scaling
CNSphasing
RefinementMethod to determine structure: MOLECULAR REPLACEMENT / Resolution: 2.52→78.02 Å / Rfactor Rfree error: 0.004 / Data cutoff high absF: 4179734.29 / Data cutoff low absF: 0 / Isotropic thermal model: RESTRAINED / Cross valid method: THROUGHOUT / σ(F): 0
RfactorNum. reflection% reflectionSelection details
Rfree0.268 4693 5 %RANDOM
Rwork0.233 ---
obs0.233 93629 96 %-
Solvent computationSolvent model: FLAT MODEL / Bsol: 32.0279 Å2 / ksol: 0.305892 e/Å3
Displacement parametersBiso mean: 59.1 Å2
Baniso -1Baniso -2Baniso -3
1-0.16 Å20 Å20 Å2
2--0.08 Å20 Å2
3----0.24 Å2
Refine analyze
FreeObs
Luzzati coordinate error0.45 Å0.39 Å
Luzzati d res low-5 Å
Luzzati sigma a0.81 Å0.73 Å
Refinement stepCycle: LAST / Resolution: 2.52→78.02 Å
ProteinNucleic acidLigandSolventTotal
Num. atoms13894 992 48 151 15085
Refine LS restraints
Refine-IDTypeDev ideal
X-RAY DIFFRACTIONc_bond_d0.009
X-RAY DIFFRACTIONc_bond_d_na
X-RAY DIFFRACTIONc_bond_d_prot
X-RAY DIFFRACTIONc_angle_d
X-RAY DIFFRACTIONc_angle_d_na
X-RAY DIFFRACTIONc_angle_d_prot
X-RAY DIFFRACTIONc_angle_deg1.4
X-RAY DIFFRACTIONc_angle_deg_na
X-RAY DIFFRACTIONc_angle_deg_prot
X-RAY DIFFRACTIONc_dihedral_angle_d22.6
X-RAY DIFFRACTIONc_dihedral_angle_d_na
X-RAY DIFFRACTIONc_dihedral_angle_d_prot
X-RAY DIFFRACTIONc_improper_angle_d1.03
X-RAY DIFFRACTIONc_improper_angle_d_na
X-RAY DIFFRACTIONc_improper_angle_d_prot
X-RAY DIFFRACTIONc_mcbond_it
X-RAY DIFFRACTIONc_mcangle_it
X-RAY DIFFRACTIONc_scbond_it
X-RAY DIFFRACTIONc_scangle_it
LS refinement shellResolution: 2.5→2.66 Å / Rfactor Rfree error: 0.017 / Total num. of bins used: 6
RfactorNum. reflection% reflection
Rfree0.442 644 5.2 %
Rwork0.441 11811 -
obs--76 %
Xplor file
Refine-IDSerial noParam fileTopol file
X-RAY DIFFRACTION1protein_rep.paramprotein.top
X-RAY DIFFRACTION2water_rep.paramdna-rna.top
X-RAY DIFFRACTION3dna-rna_rep.paramwater.top
X-RAY DIFFRACTION4ion.paramaphi_noH.top
X-RAY DIFFRACTION5aphi_noH.parion.top

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