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- EMDB-9567: Human RAD51 post-synaptic complexes -

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Basic information

Entry
Database: EMDB / ID: EMD-9567
TitleHuman RAD51 post-synaptic complexes
Map data
SampleHuman RAD51 post-synaptic complex:
Human RAD51 / (DNA) x 2 / DNA repair protein RAD51 homolog 1 / (nucleic-acidNucleic acid) x 2 / (ligand) x 2
Function / homology
Function and homology information


response to glucoside / mitotic recombination-dependent replication fork processing / chromosome organization involved in meiotic cell cycle / cellular response to camptothecin / DNA recombinase assembly / mitotic recombination / telomere maintenance via telomere lengthening / telomere maintenance via recombination / replication-born double-strand break repair via sister chromatid exchange / positive regulation of DNA ligation ...response to glucoside / mitotic recombination-dependent replication fork processing / chromosome organization involved in meiotic cell cycle / cellular response to camptothecin / DNA recombinase assembly / mitotic recombination / telomere maintenance via telomere lengthening / telomere maintenance via recombination / replication-born double-strand break repair via sister chromatid exchange / positive regulation of DNA ligation / strand invasion / cellular response to cisplatin / cellular response to hydroxyurea / lateral element / single-stranded DNA helicase activity / recombinase activity / reciprocal meiotic recombination / replication fork processing / DNA unwinding involved in DNA replication / regulation of double-strand break repair via homologous recombination / negative regulation of G0 to G1 transition / DNA-dependent ATPase activity / DNA polymerase binding / response to X-ray / condensed chromosome / nuclear chromosome / condensed nuclear chromosome / regulation of protein phosphorylation / microtubule organizing center / meiotic cell cycle / chromatin / interstrand cross-link repair / cellular response to ionizing radiation / double-strand break repair via homologous recombination / PML body / cellular response to gamma radiation / site of double-strand break / single-stranded DNA binding / protein homooligomerization / double-stranded DNA binding / nuclear chromosome, telomeric region / DNA recombination / protein C-terminus binding / mitochondrial matrix / nuclear chromatin / DNA repair / cellular response to DNA damage stimulus / chromatin binding / perinuclear region of cytoplasm / enzyme binding / protein-containing complex / mitochondrion / nucleoplasm / ATP binding / identical protein binding / nucleus / cytoplasm
AAA+ ATPase domain / DNA recombination and repair protein RecA, monomer-monomer interface / DNA repair Rad51/transcription factor NusA, alpha-helical / DNA recombination/repair protein Rad51 / DNA recombination and repair protein Rad51-like, C-terminal / DNA recombination and repair protein, RecA-like / DNA recombination and repair protein RecA-like, ATP-binding domain / P-loop containing nucleoside triphosphate hydrolase / Rad51/DMC1/RadA
DNA repair protein RAD51 homolog 1
Biological speciesHomo sapiens (human)
Methodhelical reconstruction / cryo EM / Resolution: 4.5 Å
AuthorsXu J / Zhao L / Xu Y / Zhao W / Sung P / Wang HW
CitationJournal: Nat. Struct. Mol. Biol. / Year: 2017
Title: Cryo-EM structures of human RAD51 recombinase filaments during catalysis of DNA-strand exchange.
Authors: Jingfei Xu / Lingyun Zhao / Yuanyuan Xu / Weixing Zhao / Patrick Sung / Hong-Wei Wang /
Abstract: The central step in eukaryotic homologous recombination (HR) is ATP-dependent DNA-strand exchange mediated by the Rad51 recombinase. In this process, Rad51 assembles on single-stranded DNA (ssDNA) ...The central step in eukaryotic homologous recombination (HR) is ATP-dependent DNA-strand exchange mediated by the Rad51 recombinase. In this process, Rad51 assembles on single-stranded DNA (ssDNA) and generates a helical filament that is able to search for and invade homologous double-stranded DNA (dsDNA), thus leading to strand separation and formation of new base pairs between the initiating ssDNA and the complementary strand within the duplex. Here, we used cryo-EM to solve the structures of human RAD51 in complex with DNA molecules, in presynaptic and postsynaptic states, at near-atomic resolution. Our structures reveal both conserved and distinct structural features of the human RAD51-DNA complexes compared with their prokaryotic counterpart. Notably, we also captured the structure of an arrested synaptic complex. Our results provide new insight into the molecular mechanisms of the DNA homology search and strand-exchange processes.
Validation ReportPDB-ID: 5h1c

SummaryFull reportAbout validation report
History
DepositionOct 28, 2016-
Header (metadata) releaseNov 2, 2016-
Map releaseDec 21, 2016-
UpdateOct 18, 2017-
Current statusOct 18, 2017Processing site: PDBj / Status: Released

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Structure visualization

Movie
  • Surface view with section colored by density value
  • Surface level: 0.013
  • Imaged by UCSF Chimera
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  • Surface view colored by cylindrical radius
  • Surface level: 0.013
  • Imaged by UCSF Chimera
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  • Surface view with fitted model
  • Atomic models: : PDB-5h1c
  • Surface level: 0.013
  • Imaged by UCSF Chimera
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  • Surface view with fitted model
  • Atomic models: : PDB-5h1c
  • Surface level: 0.013
  • Imaged by UCSF Chimera
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  • Simplified surface model + fitted atomic model
  • Atomic models: PDB-5h1c
  • Imaged by Jmol
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Movie viewer
Structure viewerEM map:
SurfViewMolmilJmol/JSmol
Supplemental images

Downloads & links

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Map

FileDownload / File: emd_9567.map.gz / Format: CCP4 / Size: 64 MB / Type: IMAGE STORED AS FLOATING POINT NUMBER (4 BYTES)
Projections & slices

Image control

Size
Brightness
Contrast
Others
AxesZ (Sec.)Y (Row.)X (Col.)
1.31 Å/pix.
x 256 pix.
= 334.336 Å
1.31 Å/pix.
x 256 pix.
= 334.336 Å
1.31 Å/pix.
x 256 pix.
= 334.336 Å

Surface

Projections

Slices (1/3)

Slices (1/2)

Slices (2/3)

Images are generated by Spider.

Voxel sizeX=Y=Z: 1.306 Å
Density
Contour LevelBy AUTHOR: 0.013 / Movie #1: 0.013
Minimum - Maximum-0.018787928 - 0.051937886
Average (Standard dev.)0.00038559092 (±0.0028562339)
SymmetrySpace group: 1
Details

EMDB XML:

Map geometry
Axis orderXYZ
Origin000
Dimensions256256256
Spacing256256256
CellA=B=C: 334.336 Å
α=β=γ: 90.0 °

CCP4 map header:

modeImage stored as Reals
Å/pix. X/Y/Z1.3061.3061.306
M x/y/z256256256
origin x/y/z0.0000.0000.000
length x/y/z334.336334.336334.336
α/β/γ90.00090.00090.000
start NX/NY/NZ-2600
NX/NY/NZ264044
MAP C/R/S123
start NC/NR/NS000
NC/NR/NS256256256
D min/max/mean-0.0190.0520.000

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Supplemental data

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Sample components

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Entire Human RAD51 post-synaptic complex

EntireName: Human RAD51 post-synaptic complex / Number of components: 9

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Component #1: protein, Human RAD51 post-synaptic complex

ProteinName: Human RAD51 post-synaptic complex / Recombinant expression: No
MassTheoretical: 24.18 MDa

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Component #2: protein, Human RAD51

ProteinName: Human RAD51 / Recombinant expression: No

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Component #3: protein, DNA

ProteinName: DNA / Recombinant expression: No

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Component #4: protein, DNA

ProteinName: DNA / Recombinant expression: No

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Component #5: protein, DNA repair protein RAD51 homolog 1

ProteinName: DNA repair protein RAD51 homolog 1 / Number of Copies: 3 / Recombinant expression: No
MassTheoretical: 37.008074 kDa
SourceSpecies: Homo sapiens (human)
Source (engineered)Expression System: Escherichia coli 'BL21-Gold(DE3)pLysS AG' (bacteria)

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Component #6: nucleic-acid, DNA (5'-D(P*TP*TP*TP*TP*TP*TP*TP*TP*T)-3')

nucleic acidName: DNA (5'-D(P*TP*TP*TP*TP*TP*TP*TP*TP*T)-3') / Class: DNA / Structure: OTHER / Synthetic: No
Sequence:
(DT)(DT)(DT)(DT)(DT)(DT)(DT)(DT)(DT)
MassTheoretical: 2.692778 kDa
SourceSpecies: Homo sapiens (human)

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Component #7: nucleic-acid, DNA (5'-D(P*AP*AP*AP*AP*AP*AP*AP*AP*A)-3')

nucleic acidName: DNA (5'-D(P*AP*AP*AP*AP*AP*AP*AP*AP*A)-3') / Class: DNA / Structure: OTHER / Synthetic: No
Sequence:
(DA)(DA)(DA)(DA)(DA)(DA)(DA)(DA)(DA)
MassTheoretical: 2.773904 kDa
SourceSpecies: Homo sapiens (human)

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Component #8: ligand, MAGNESIUM ION

LigandName: MAGNESIUM ION / Number of Copies: 3 / Recombinant expression: No
MassTheoretical: 2.430505 MDa

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Component #9: ligand, PHOSPHOAMINOPHOSPHONIC ACID-ADENYLATE ESTER

LigandName: PHOSPHOAMINOPHOSPHONIC ACID-ADENYLATE ESTER / Number of Copies: 3 / Recombinant expression: No
MassTheoretical: 0.506196 kDa

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Experimental details

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Sample preparation

SpecimenSpecimen state: Filament / Method: cryo EM
Helical parametersAxial symmetry: C1 (asymmetric) / Delta z: 15.8 Å / Delta phi: 56.18 %deg;
Sample solutionSpecimen conc.: 0.075 mg/mL
Buffer solution: 25mM Tris-HCl, pH 7.5, 50mM KCl, 1mM dithiothreitol, 1mM AMP-PNP and 2mM MgCl2
pH: 7.5
VitrificationInstrument: FEI VITROBOT MARK IV / Cryogen name: ETHANE / Temperature: 289 K / Humidity: 100 %

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Electron microscopy imaging

Experimental equipment
Model: Titan Krios / Image courtesy: FEI Company
ImagingMicroscope: FEI TITAN KRIOS
Electron gunElectron source: FIELD EMISSION GUN / Accelerating voltage: 300 kV / Electron dose: 50 e/Å2 / Illumination mode: FLOOD BEAM
LensCs: 2.7 mm / Imaging mode: BRIGHT FIELD / Defocus: 1500.0 - 2500.0 nm
Specimen HolderModel: OTHER
CameraDetector: GATAN K2 SUMMIT (4k x 4k)

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Image acquisition

Image acquisitionNumber of digital images: 528 / Sampling size: 5 µm

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Image processing

ProcessingMethod: helical reconstruction
3D reconstructionAlgorithm: BACK PROJECTION / Software: SPIDER, RELION / Resolution: 4.5 Å / Resolution method: FSC 0.143 CUT-OFF

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Atomic model buiding

Modeling #1Refinement space: REAL
Input PDB model: 1SZP
Chain ID: E
Output model

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