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- PDB-7x5b: Crystal structure of RuvB -

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Basic information

Entry
Database: PDB / ID: 7x5b
TitleCrystal structure of RuvB
ComponentsHolliday junction ATP-dependent DNA helicase RuvB
KeywordsMOTOR PROTEIN / RuvB / Holliday juncition / Homologous recombination / DNA damage repair / ATP hydrolysis
Function / homology
Function and homology information


cellular response to chromate / cellular response to selenite ion / Holliday junction resolvase complex / four-way junction helicase activity / four-way junction DNA binding / Hydrolases; Acting on acid anhydrides; Acting on acid anhydrides to facilitate cellular and subcellular movement / DNA recombination / DNA repair / ATP hydrolysis activity / ATP binding / cytoplasm
Similarity search - Function
DNA helicase, Holliday junction RuvB-type / RuvB C-terminal winged helix domain / RuvB-like P-loop domain / RuvB, AAA lid domain / RuvB C-terminal winged helix domain / Holliday junction DNA helicase RuvB P-loop domain / RuvB AAA lid domain / Winged helix DNA-binding domain superfamily / Winged helix-like DNA-binding domain superfamily / ATPases associated with a variety of cellular activities ...DNA helicase, Holliday junction RuvB-type / RuvB C-terminal winged helix domain / RuvB-like P-loop domain / RuvB, AAA lid domain / RuvB C-terminal winged helix domain / Holliday junction DNA helicase RuvB P-loop domain / RuvB AAA lid domain / Winged helix DNA-binding domain superfamily / Winged helix-like DNA-binding domain superfamily / ATPases associated with a variety of cellular activities / AAA+ ATPase domain / P-loop containing nucleoside triphosphate hydrolase
Similarity search - Domain/homology
ADENOSINE-5'-DIPHOSPHATE / Holliday junction branch migration complex subunit RuvB
Similarity search - Component
Biological speciesPseudomonas aeruginosa PAO1 (bacteria)
MethodX-RAY DIFFRACTION / SYNCHROTRON / MOLECULAR REPLACEMENT / Resolution: 2.16 Å
AuthorsLin, Z. / Qu, Q. / Zhang, X. / Zhou, Z. / Dai, L.
Funding support China, 2items
OrganizationGrant numberCountry
National Natural Science Foundation of China (NSFC)31971222 China
National Natural Science Foundation of China (NSFC)32171194 China
CitationJournal: Front Plant Sci / Year: 2023
Title: Cryo-EM structure of the RuvAB-Holliday junction intermediate complex from .
Authors: Xu Zhang / Zixuan Zhou / Lin Dai / Yulin Chao / Zheng Liu / Mingdong Huang / Qianhui Qu / Zhonghui Lin /
Abstract: Holliday junction (HJ) is a four-way structured DNA intermediate in homologous recombination. In bacteria, the HJ-specific binding protein RuvA and the motor protein RuvB together form the RuvAB ...Holliday junction (HJ) is a four-way structured DNA intermediate in homologous recombination. In bacteria, the HJ-specific binding protein RuvA and the motor protein RuvB together form the RuvAB complex to catalyze HJ branch migration. (, Pa) is a ubiquitous opportunistic bacterial pathogen that can cause serious infection in a variety of host species, including vertebrate animals, insects and plants. Here, we describe the cryo-Electron Microscopy (cryo-EM) structure of the RuvAB-HJ intermediate complex from . The structure shows that two RuvA tetramers sandwich HJ at the junction center and disrupt base pairs at the branch points of RuvB-free HJ arms. Eight RuvB subunits are recruited by the RuvA octameric core and form two open-rings to encircle two opposite HJ arms. Each RuvB subunit individually binds a RuvA domain III. The four RuvB subunits within the ring display distinct subdomain conformations, and two of them engage the central DNA duplex at both strands with their C-terminal β-hairpins. Together with the biochemical analyses, our structure implicates a potential mechanism of RuvB motor assembly onto HJ DNA.
History
DepositionMar 4, 2022Deposition site: PDBJ / Processing site: PDBJ
Revision 1.0Mar 8, 2023Provider: repository / Type: Initial release
Revision 1.1Sep 20, 2023Group: Data collection / Database references
Category: chem_comp_atom / chem_comp_bond ...chem_comp_atom / chem_comp_bond / citation / citation_author
Item: _citation.country / _citation.journal_abbrev ..._citation.country / _citation.journal_abbrev / _citation.journal_id_CSD / _citation.journal_id_ISSN / _citation.journal_volume / _citation.page_first / _citation.page_last / _citation.pdbx_database_id_DOI / _citation.pdbx_database_id_PubMed / _citation.title / _citation.year
Revision 1.2Nov 29, 2023Group: Refinement description / Category: pdbx_initial_refinement_model

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Structure visualization

Structure viewerMolecule:
MolmilJmol/JSmol

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Assembly

Deposited unit
A: Holliday junction ATP-dependent DNA helicase RuvB
hetero molecules


Theoretical massNumber of molelcules
Total (without water)39,3952
Polymers38,9681
Non-polymers4271
Water3,531196
1


  • Idetical with deposited unit
  • defined by author&software
TypeNameSymmetry operationNumber
identity operation1_555x,y,z1
Buried area800 Å2
ΔGint-5 kcal/mol
Surface area15570 Å2
MethodPISA
Unit cell
Length a, b, c (Å)85.677, 85.677, 76.973
Angle α, β, γ (deg.)90.000, 90.000, 120.000
Int Tables number170
Space group name H-MP65
Space group name HallP65
Symmetry operation#1: x,y,z
#2: x-y,x,z+5/6
#3: y,-x+y,z+1/6
#4: -y,x-y,z+2/3
#5: -x+y,-x,z+1/3
#6: -x,-y,z+1/2

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Components

#1: Protein Holliday junction ATP-dependent DNA helicase RuvB


Mass: 38967.551 Da / Num. of mol.: 1 / Mutation: A146G
Source method: isolated from a genetically manipulated source
Source: (gene. exp.) Pseudomonas aeruginosa PAO1 (bacteria) / Strain: PAO1 / Gene: ruvB / Production host: Escherichia coli BL21(DE3) (bacteria) / Strain (production host): BL21(DE3) / References: UniProt: Q51426, DNA helicase
#2: Chemical ChemComp-ADP / ADENOSINE-5'-DIPHOSPHATE


Mass: 427.201 Da / Num. of mol.: 1 / Source method: obtained synthetically / Formula: C10H15N5O10P2 / Feature type: SUBJECT OF INVESTIGATION / Comment: ADP, energy-carrying molecule*YM
#3: Water ChemComp-HOH / water


Mass: 18.015 Da / Num. of mol.: 196 / Source method: isolated from a natural source / Formula: H2O
Has ligand of interestY

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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: 2.09 Å3/Da / Density % sol: 41.23 %
Crystal growTemperature: 291 K / Method: vapor diffusion, sitting drop
Details: 0.2 M LProline, 0.1 M HEPES pH 7.5, 10% w/v polyethylene glycol 3350, 10 mM Trimethylamine hydrochloride

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Data collection

DiffractionMean temperature: 80 K / Serial crystal experiment: N
Diffraction sourceSource: SYNCHROTRON / Site: SSRF / Beamline: BL19U1 / Wavelength: 0.978 Å
DetectorType: DECTRIS PILATUS3 6M / Detector: PIXEL / Date: Jul 13, 2020
RadiationProtocol: SINGLE WAVELENGTH / Monochromatic (M) / Laue (L): M / Scattering type: x-ray
Radiation wavelengthWavelength: 0.978 Å / Relative weight: 1
ReflectionResolution: 2.1→50 Å / Num. obs: 17198 / % possible obs: 99.6 % / Redundancy: 20 % / Biso Wilson estimate: 22.37 Å2 / Rmerge(I) obs: 0.14 / Net I/σ(I): 23.9
Reflection shellResolution: 2.1→2.24 Å / CC1/2: 0.774

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Processing

Software
NameVersionClassification
HKL-30001.13_2998data reduction
HKL-30001.13_2998data scaling
PHENIX1.19_4092refinement
MOLREPphasing
RefinementMethod to determine structure: MOLECULAR REPLACEMENT
Starting model: 6BLB
Resolution: 2.16→28.04 Å / SU ML: 0.1876 / Cross valid method: FREE R-VALUE / σ(F): 1.38 / Phase error: 20.9614 / Stereochemistry target values: GeoStd + Monomer Library
RfactorNum. reflection% reflection
Rfree0.2173 1717 9.98 %
Rwork0.1731 15481 -
obs0.1775 17198 99.65 %
Solvent computationShrinkage radii: 0.9 Å / VDW probe radii: 1.11 Å / Solvent model: FLAT BULK SOLVENT MODEL
Displacement parametersBiso mean: 27.14 Å2
Refinement stepCycle: LAST / Resolution: 2.16→28.04 Å
ProteinNucleic acidLigandSolventTotal
Num. atoms2440 0 27 196 2663
Refine LS restraints
Refine-IDTypeDev idealNumber
X-RAY DIFFRACTIONf_bond_d0.00692508
X-RAY DIFFRACTIONf_angle_d0.90823398
X-RAY DIFFRACTIONf_chiral_restr0.0544391
X-RAY DIFFRACTIONf_plane_restr0.0057443
X-RAY DIFFRACTIONf_dihedral_angle_d18.73921533
LS refinement shell
Resolution (Å)Rfactor RfreeNum. reflection RfreeRfactor RworkNum. reflection RworkRefine-ID% reflection obs (%)
2.16-2.230.25651400.19731242X-RAY DIFFRACTION96.58
2.23-2.30.25841420.19261280X-RAY DIFFRACTION99.86
2.3-2.380.23491430.18411303X-RAY DIFFRACTION100
2.38-2.470.24371420.18951273X-RAY DIFFRACTION100
2.47-2.590.23221460.18821291X-RAY DIFFRACTION100
2.59-2.720.27951430.19271278X-RAY DIFFRACTION100
2.72-2.890.19721440.18061296X-RAY DIFFRACTION100
2.89-3.120.22911380.17261302X-RAY DIFFRACTION100
3.12-3.430.18891430.17411301X-RAY DIFFRACTION100
3.43-3.930.21281500.16071287X-RAY DIFFRACTION100
3.93-4.940.17031430.14741297X-RAY DIFFRACTION99.93
4.94-28.040.22441430.17191331X-RAY DIFFRACTION99.59

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