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- EMDB-34430: Cryo-EM structure of the human RAD52 protein -

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Basic information

Entry
Database: EMDB / ID: EMD-34430
TitleCryo-EM structure of the human RAD52 protein
Map data
Sample
  • Organelle or cellular component: RAD52
    • Protein or peptide: DNA repair protein RAD52 homolog
Function / homology
Function and homology information


double-strand break repair via single-strand annealing / DNA double-strand break processing involved in repair via single-strand annealing / DNA recombinase assembly / mitotic recombination / regulation of nucleotide-excision repair / HDR through MMEJ (alt-NHEJ) / HDR through Single Strand Annealing (SSA) / SUMOylation of DNA damage response and repair proteins / protein-DNA complex / double-strand break repair via homologous recombination ...double-strand break repair via single-strand annealing / DNA double-strand break processing involved in repair via single-strand annealing / DNA recombinase assembly / mitotic recombination / regulation of nucleotide-excision repair / HDR through MMEJ (alt-NHEJ) / HDR through Single Strand Annealing (SSA) / SUMOylation of DNA damage response and repair proteins / protein-DNA complex / double-strand break repair via homologous recombination / double-strand break repair / single-stranded DNA binding / cellular response to oxidative stress / DNA recombination / protein-containing complex / DNA binding / nucleoplasm / identical protein binding / nucleus
Similarity search - Function
DNA recombination/repair protein Rad52 / DNA repair protein Rad52/59/22 / Rad52 family / DNA repair protein Rad52/59/22 superfamily / Rad52/22 family double-strand break repair protein
Similarity search - Domain/homology
DNA repair protein RAD52 homolog
Similarity search - Component
Biological speciesHomo sapiens (human)
Methodsingle particle reconstruction / cryo EM / Resolution: 3.48 Å
AuthorsKinoshita C / Takizawa Y / Saotome M / Ogino S / Kurumizaka H / Kagawa W
Funding support Japan, 7 items
OrganizationGrant numberCountry
Japan Society for the Promotion of Science (JSPS)JP18H05534 Japan
Japan Society for the Promotion of Science (JSPS)19K12328 Japan
Japan Society for the Promotion of Science (JSPS)20H00449 Japan
Japan Society for the Promotion of Science (JSPS)22H03743 Japan
Japan Society for the Promotion of Science (JSPS)22K06098 Japan
Japan Science and TechnologyJPMJER1901 Japan
Japan Agency for Medical Research and Development (AMED)JP22am121009 Japan
CitationJournal: FEBS Open Bio / Year: 2023
Title: The cryo-EM structure of full-length RAD52 protein contains an undecameric ring.
Authors: Chiaki Kinoshita / Yoshimasa Takizawa / Mika Saotome / Shun Ogino / Hitoshi Kurumizaka / Wataru Kagawa /
Abstract: The human RAD52 protein, which forms an oligomeric ring structure, is involved in DNA double-strand break repair. The N-terminal half of RAD52 is primarily responsible for self-oligomerisation and ...The human RAD52 protein, which forms an oligomeric ring structure, is involved in DNA double-strand break repair. The N-terminal half of RAD52 is primarily responsible for self-oligomerisation and DNA binding. Crystallographic studies have revealed the detailed structure of the N-terminal half. However, only low-resolution structures have been reported for the full-length protein, and thus the structural role of the C-terminal half in self-oligomerisation has remained elusive. In this study, we determined the solution structure of the human RAD52 protein by cryo-electron microscopy (cryo-EM), at an average resolution of 3.5 Å. The structure revealed an undecameric ring that is nearly identical to the crystal structures of the N-terminal half. The cryo-EM map for the C-terminal half was poorly defined, indicating that the region is intrinsically disordered. The present cryo-EM structure provides important insights into the mechanistic roles played by the N-terminal and C-terminal halves of RAD52 during DNA double-strand break repair.
History
DepositionOct 3, 2022-
Header (metadata) releaseFeb 8, 2023-
Map releaseFeb 8, 2023-
UpdateMar 22, 2023-
Current statusMar 22, 2023Processing site: PDBj / Status: Released

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Structure visualization

Supplemental images

emd_34430.png

Downloads & links

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Map

FileDownload / File: emd_34430.map.gz / Format: CCP4 / Size: 30.5 MB / Type: IMAGE STORED AS FLOATING POINT NUMBER (4 BYTES)
Voxel sizeX=Y=Z: 1.06 Å
Density
Contour LevelBy AUTHOR: 3.48
Minimum - Maximum-18.745953 - 30.877308
Average (Standard dev.)-1.873686e-09 (±0.99999994)
SymmetrySpace group: 1
Details

EMDB XML:

Map geometry
Axis orderXYZ
Origin000
Dimensions200200200
Spacing200200200
CellA=B=C: 211.99998 Å
α=β=γ: 90.0 °

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Supplemental data

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Half map: #1

Fileemd_34430_half_map_1.map
Projections & Slices
AxesZYX

Projections

Slices (1/2)
Density Histograms

Histogram

Histogram (log scale)

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Half map: #2

Fileemd_34430_half_map_2.map
Projections & Slices
AxesZYX

Projections

Slices (1/2)
Density Histograms

Histogram

Histogram (log scale)

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Sample components

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Entire : RAD52

EntireName: RAD52
Components
  • Organelle or cellular component: RAD52
    • Protein or peptide: DNA repair protein RAD52 homolog

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Supramolecule #1: RAD52

SupramoleculeName: RAD52 / type: organelle_or_cellular_component / ID: 1 / Parent: 0 / Macromolecule list: all
Source (natural)Organism: Homo sapiens (human)

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Macromolecule #1: DNA repair protein RAD52 homolog

MacromoleculeName: DNA repair protein RAD52 homolog / type: protein_or_peptide / ID: 1 / Number of copies: 11 / Enantiomer: LEVO
Source (natural)Organism: Homo sapiens (human)
Molecular weightTheoretical: 46.514934 KDa
Recombinant expressionOrganism: Escherichia coli (E. coli)
SequenceString: GSHMSGTEEA ILGGRDSHPA AGGGSVLCFG QCQYTAEEYQ AIQKALRQRL GPEYISSRMA GGGQKVCYIE GHRVINLANE MFGYNGWAH SITQQNVDFV DLNNGKFYVG VCAFVRVQLK DGSYHEDVGY GVSEGLKSKA LSLEKARKEA VTDGLKRALR S FGNALGNC ...String:
GSHMSGTEEA ILGGRDSHPA AGGGSVLCFG QCQYTAEEYQ AIQKALRQRL GPEYISSRMA GGGQKVCYIE GHRVINLANE MFGYNGWAH SITQQNVDFV DLNNGKFYVG VCAFVRVQLK DGSYHEDVGY GVSEGLKSKA LSLEKARKEA VTDGLKRALR S FGNALGNC ILDKDYLRSL NKLPRQLPLE VDLTKAKRQD LEPSVEEARY NSCRPNMALG HPQLQQVTSP SRPSHAVIPA DQ DCSSRSL SSSAVESEAT HQRKLRQKQL QQQFRERMEK QQVRVSTPSA EKSEAAPPAP PVTHSTPVTV SEPLLEKDFL AGV TQELIK TLEDNSEKWA VTPDAGDGVV KPSSRADPAQ TSDTLALNNQ MVTQNRTPHS VCHQKPQAKS GSWDLQTYSA DQRT TGNWE SHRKSQDMKK RKYDPS

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Experimental details

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Structure determination

Methodcryo EM
Processingsingle particle reconstruction
Aggregation stateparticle

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Sample preparation

Concentration0.4 mg/mL
BufferpH: 7.5
Component:
ConcentrationFormulaName
20.0 mMHEPES2-[4-(2-hydroxyethyl)piperazin-1-yl]ethanesulfonic acid
0.1 mMEDTAEthylenediaminetetraacetic acid2,2',2'',2'''-(Ethane-1,2-diyldinitrilo)tetraacetic acid
2.0 mM2-mercaptoethanol2-mercaptoethanol
0.4 MKClpotassium chloride
GridModel: Quantifoil R1.2/1.3 / Material: COPPER / Mesh: 200 / Support film - Material: CARBON / Support film - topology: HOLEY / Pretreatment - Type: GLOW DISCHARGE
VitrificationCryogen name: ETHANE / Chamber humidity: 100 % / Chamber temperature: 277 K / Instrument: FEI VITROBOT MARK IV

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Electron microscopy

MicroscopeFEI TITAN KRIOS
Electron beamAcceleration voltage: 300 kV / Electron source: FIELD EMISSION GUN
Electron opticsIllumination mode: FLOOD BEAM / Imaging mode: BRIGHT FIELDBright-field microscopy / Nominal defocus max: 2.5 µm / Nominal defocus min: 1.0 µm
Sample stageSpecimen holder model: FEI TITAN KRIOS AUTOGRID HOLDER / Cooling holder cryogen: NITROGEN
Image recordingFilm or detector model: GATAN K3 (6k x 4k) / Average electron dose: 57.6 e/Å2
Experimental equipment
Model: Titan Krios / Image courtesy: FEI Company

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Image processing

Startup modelType of model: PDB ENTRY
PDB model - PDB ID:

1kn0

Initial angle assignmentType: MAXIMUM LIKELIHOOD / Software - Name: RELION (ver. 3.1)
Final 3D classificationSoftware - Name: RELION (ver. 3.1)
Final angle assignmentType: MAXIMUM LIKELIHOOD / Software - Name: RELION (ver. 3.1)
Final reconstructionAlgorithm: BACK PROJECTION / Resolution.type: BY AUTHOR / Resolution: 3.48 Å / Resolution method: FSC 0.143 CUT-OFF / Software - Name: RELION (ver. 3.1) / Number images used: 39100
FSC plot (resolution estimation)

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