+Open data
-Basic information
Entry | Database: EMDB / ID: EMD-17584 | |||||||||
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Title | RAD51 filament on ssDNA bound by the BRCA2 c-terminus | |||||||||
Map data | CryoEM map of a RAD51 filament on ssDNA bound with the BRCA2 c-terminus | |||||||||
Sample |
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Keywords | RAD51 / BRCA2 / Filament / Complex / RECOMBINATION | |||||||||
Function / homology | Function and homology information BRCA2-MAGE-D1 complex / negative regulation of mammary gland epithelial cell proliferation / presynaptic intermediate filament cytoskeleton / mitotic recombination-dependent replication fork processing / cellular response to camptothecin / chromosome organization involved in meiotic cell cycle / establishment of protein localization to telomere / telomere maintenance via telomere lengthening / positive regulation of DNA ligation / Impaired BRCA2 translocation to the nucleus ...BRCA2-MAGE-D1 complex / negative regulation of mammary gland epithelial cell proliferation / presynaptic intermediate filament cytoskeleton / mitotic recombination-dependent replication fork processing / cellular response to camptothecin / chromosome organization involved in meiotic cell cycle / establishment of protein localization to telomere / telomere maintenance via telomere lengthening / positive regulation of DNA ligation / Impaired BRCA2 translocation to the nucleus / Impaired BRCA2 binding to SEM1 (DSS1) / nuclear ubiquitin ligase complex / double-strand break repair involved in meiotic recombination / cellular response to hydroxyurea / replication-born double-strand break repair via sister chromatid exchange / lateral element / telomere maintenance via recombination / DNA recombinase assembly / histone H4 acetyltransferase activity / histone H3 acetyltransferase activity / regulation of DNA damage checkpoint / mitotic recombination / Impaired BRCA2 binding to PALB2 / DNA strand invasion / HDR through MMEJ (alt-NHEJ) / gamma-tubulin binding / DNA strand exchange activity / reciprocal meiotic recombination / DNA repair complex / Defective homologous recombination repair (HRR) due to BRCA1 loss of function / Defective HDR through Homologous Recombination Repair (HRR) due to PALB2 loss of BRCA1 binding function / Defective HDR through Homologous Recombination Repair (HRR) due to PALB2 loss of BRCA2/RAD51/RAD51C binding function / Homologous DNA Pairing and Strand Exchange / Resolution of D-loop Structures through Synthesis-Dependent Strand Annealing (SDSA) / response to UV-C / oocyte maturation / Resolution of D-loop Structures through Holliday Junction Intermediates / single-stranded DNA helicase activity / inner cell mass cell proliferation / ATP-dependent DNA damage sensor activity / DNA damage response, signal transduction by p53 class mediator resulting in transcription of p21 class mediator / hematopoietic stem cell proliferation / HDR through Single Strand Annealing (SSA) / Impaired BRCA2 binding to RAD51 / regulation of double-strand break repair via homologous recombination / nuclear chromosome / female gonad development / replication fork processing / male meiosis I / DNA unwinding involved in DNA replication / Transcriptional Regulation by E2F6 / Presynaptic phase of homologous DNA pairing and strand exchange / centrosome duplication / response to X-ray / intrinsic apoptotic signaling pathway in response to DNA damage by p53 class mediator / ATP-dependent activity, acting on DNA / interstrand cross-link repair / DNA polymerase binding / condensed chromosome / positive regulation of mitotic cell cycle / regulation of cytokinesis / condensed nuclear chromosome / secretory granule / meiotic cell cycle / male germ cell nucleus / cellular response to ionizing radiation / nucleotide-excision repair / response to gamma radiation / regulation of protein phosphorylation / double-strand break repair via homologous recombination / HDR through Homologous Recombination (HRR) / brain development / PML body / Meiotic recombination / cellular senescence / double-strand break repair / site of double-strand break / single-stranded DNA binding / spermatogenesis / double-stranded DNA binding / protease binding / DNA recombination / chromosome, telomeric region / mitochondrial matrix / DNA repair / centrosome / DNA damage response / chromatin binding / chromatin / regulation of DNA-templated transcription / nucleolus / perinuclear region of cytoplasm / positive regulation of DNA-templated transcription / enzyme binding / ATP hydrolysis activity / protein-containing complex / mitochondrion / nucleoplasm / ATP binding / identical protein binding Similarity search - Function | |||||||||
Biological species | Homo sapiens (human) / synthetic construct (others) | |||||||||
Method | helical reconstruction / cryo EM / Resolution: 2.61 Å | |||||||||
Authors | Appleby R / Pellegrini L | |||||||||
Funding support | United Kingdom, 2 items
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Citation | Journal: Nat Commun / Year: 2023 Title: Structural basis for stabilisation of the RAD51 nucleoprotein filament by BRCA2. Authors: Robert Appleby / Luay Joudeh / Katie Cobbett / Luca Pellegrini / Abstract: The BRCA2 tumour suppressor protein preserves genomic integrity via interactions with the DNA-strand exchange RAD51 protein in homology-directed repair. The RAD51-binding TR2 motif at the BRCA2 C- ...The BRCA2 tumour suppressor protein preserves genomic integrity via interactions with the DNA-strand exchange RAD51 protein in homology-directed repair. The RAD51-binding TR2 motif at the BRCA2 C-terminus is essential for protection and restart of stalled replication forks. Biochemical evidence shows that TR2 recognises filamentous RAD51, but existing models of TR2 binding to RAD51 lack a structural basis. Here we used cryo-electron microscopy and structure-guided mutagenesis to elucidate the mechanism of TR2 binding to nucleoprotein filaments of human RAD51. We find that TR2 binds across the protomer interface in the filament, acting as a brace for adjacent RAD51 molecules. TR2 targets an acidic-patch motif on human RAD51 that serves as a recruitment hub in fission yeast Rad51 for recombination mediators Rad52 and Rad55-Rad57. Our findings provide a structural rationale for RAD51 filament stabilisation by BRCA2 and reveal a common recruitment mechanism of recombination mediators to the RAD51 filament. | |||||||||
History |
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-Structure visualization
Supplemental images |
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-Downloads & links
-EMDB archive
Map data | emd_17584.map.gz | 5 MB | EMDB map data format | |
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Header (meta data) | emd-17584-v30.xml emd-17584.xml | 18.7 KB 18.7 KB | Display Display | EMDB header |
Images | emd_17584.png | 66.6 KB | ||
Filedesc metadata | emd-17584.cif.gz | 6.1 KB | ||
Others | emd_17584_half_map_1.map.gz emd_17584_half_map_2.map.gz | 23.5 MB 23.5 MB | ||
Archive directory | http://ftp.pdbj.org/pub/emdb/structures/EMD-17584 ftp://ftp.pdbj.org/pub/emdb/structures/EMD-17584 | HTTPS FTP |
-Validation report
Summary document | emd_17584_validation.pdf.gz | 721.1 KB | Display | EMDB validaton report |
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Full document | emd_17584_full_validation.pdf.gz | 720.7 KB | Display | |
Data in XML | emd_17584_validation.xml.gz | 9.7 KB | Display | |
Data in CIF | emd_17584_validation.cif.gz | 10.7 KB | Display | |
Arichive directory | https://ftp.pdbj.org/pub/emdb/validation_reports/EMD-17584 ftp://ftp.pdbj.org/pub/emdb/validation_reports/EMD-17584 | HTTPS FTP |
-Related structure data
Related structure data | 8pbcMC 8pbdC M: atomic model generated by this map C: citing same article (ref.) |
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Similar structure data | Similarity search - Function & homologyF&H Search |
-Links
EMDB pages | EMDB (EBI/PDBe) / EMDataResource |
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Related items in Molecule of the Month |
-Map
File | Download / File: emd_17584.map.gz / Format: CCP4 / Size: 5.3 MB / Type: IMAGE STORED AS FLOATING POINT NUMBER (4 BYTES) | ||||||||||||||||||||
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Annotation | CryoEM map of a RAD51 filament on ssDNA bound with the BRCA2 c-terminus | ||||||||||||||||||||
Voxel size | X=Y=Z: 1.304 Å | ||||||||||||||||||||
Density |
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Symmetry | Space group: 1 | ||||||||||||||||||||
Details | EMDB XML:
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-Supplemental data
-Half map: Half-map 1
File | emd_17584_half_map_1.map | ||||||||||||
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Annotation | Half-map 1 | ||||||||||||
Projections & Slices |
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Density Histograms |
-Half map: Half-map 2
File | emd_17584_half_map_2.map | ||||||||||||
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Annotation | Half-map 2 | ||||||||||||
Projections & Slices |
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Density Histograms |
-Sample components
-Entire : RAD51 filament formed on ssDNA and bound with a peptide correspon...
Entire | Name: RAD51 filament formed on ssDNA and bound with a peptide corresponding to the BRCA2 c-terminus |
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Components |
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-Supramolecule #1: RAD51 filament formed on ssDNA and bound with a peptide correspon...
Supramolecule | Name: RAD51 filament formed on ssDNA and bound with a peptide corresponding to the BRCA2 c-terminus type: complex / ID: 1 / Parent: 0 / Macromolecule list: #1-#3 |
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-Supramolecule #2: DNA repair protein RAD51 homolog 1
Supramolecule | Name: DNA repair protein RAD51 homolog 1 / type: complex / ID: 2 / Parent: 1 / Macromolecule list: #1 Details: RAD51 was recombinantly purified BRCA2 c-terminus peptide was synthesised 30-mer ssDNA was synthesised |
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Source (natural) | Organism: Homo sapiens (human) |
-Supramolecule #3: Breast cancer type 2 susceptibility protein
Supramolecule | Name: Breast cancer type 2 susceptibility protein / type: complex / ID: 3 / Parent: 1 / Macromolecule list: #2 |
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Source (natural) | Organism: Homo sapiens (human) |
-Supramolecule #4: DNA (30-MER)
Supramolecule | Name: DNA (30-MER) / type: complex / ID: 4 / Parent: 1 / Macromolecule list: #3 |
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Source (natural) | Organism: synthetic construct (others) |
-Macromolecule #1: DNA repair protein RAD51 homolog 1
Macromolecule | Name: DNA repair protein RAD51 homolog 1 / type: protein_or_peptide / ID: 1 / Number of copies: 11 / Enantiomer: LEVO |
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Source (natural) | Organism: Homo sapiens (human) |
Molecular weight | Theoretical: 37.009125 KDa |
Recombinant expression | Organism: Escherichia coli (E. coli) |
Sequence | String: MAMQMQLEAN ADTSVEEESF GPQPISRLEQ CGINANDVKK LEEAGFHTVE AVAYAPKKEL INIKGISEAK ADKILAEAAK LVPMGFTTA TEFHQRRSEI IQITTGSKEL DKLLQGGIET GSITEMFGEF RTGKTQICHT LAVTCQLPID RGGGEGKAMY I DTEGTFRP ...String: MAMQMQLEAN ADTSVEEESF GPQPISRLEQ CGINANDVKK LEEAGFHTVE AVAYAPKKEL INIKGISEAK ADKILAEAAK LVPMGFTTA TEFHQRRSEI IQITTGSKEL DKLLQGGIET GSITEMFGEF RTGKTQICHT LAVTCQLPID RGGGEGKAMY I DTEGTFRP ERLLAVAERY GLSGSDVLDN VAYARAFNTD HQTQLLYQAS AMMVESRYAL LIVDSATALY RTDYSGRGEL SA RQMHLAR FLRMLLRLAD EFGVAVVITN QVVAQVDGAA MFAADPKKPI GGNIIAHAST TRLYLRKGRG ETRICKIYDS PCL PEAEAM FAINADGVGD AKD UniProtKB: DNA repair protein RAD51 homolog 1 |
-Macromolecule #2: Breast cancer type 2 susceptibility protein
Macromolecule | Name: Breast cancer type 2 susceptibility protein / type: protein_or_peptide / ID: 2 / Number of copies: 10 / Enantiomer: LEVO |
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Source (natural) | Organism: Homo sapiens (human) |
Molecular weight | Theoretical: 5.483415 KDa |
Sequence | String: DDQKNCKKRR ALDFLSRLPL PPPVSPICTF VSPAAQKAFQ PPRSCGTKY UniProtKB: Breast cancer type 2 susceptibility protein |
-Macromolecule #3: DNA (30-MER)
Macromolecule | Name: DNA (30-MER) / type: dna / ID: 3 / Number of copies: 1 / Classification: DNA |
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Source (natural) | Organism: synthetic construct (others) |
Molecular weight | Theoretical: 9.671227 KDa |
Sequence | String: (DG)(DG)(DA)(DG)(DG)(DA)(DG)(DG)(DA)(DG) (DG)(DA)(DG)(DG)(DA)(DG)(DG)(DA)(DG)(DG) (DA)(DG)(DG)(DA)(DG)(DG)(DA)(DG)(DG) (DA) |
-Macromolecule #4: ADENOSINE-5'-TRIPHOSPHATE
Macromolecule | Name: ADENOSINE-5'-TRIPHOSPHATE / type: ligand / ID: 4 / Number of copies: 11 / Formula: ATP |
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Molecular weight | Theoretical: 507.181 Da |
Chemical component information | ChemComp-ATP: |
-Macromolecule #5: CALCIUM ION
Macromolecule | Name: CALCIUM ION / type: ligand / ID: 5 / Number of copies: 22 / Formula: CA |
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Molecular weight | Theoretical: 40.078 Da |
-Experimental details
-Structure determination
Method | cryo EM |
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Processing | helical reconstruction |
Aggregation state | filament |
-Sample preparation
Buffer | pH: 7.4 |
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Grid | Material: GOLD |
Vitrification | Cryogen name: ETHANE / Instrument: FEI VITROBOT MARK IV |
-Electron microscopy
Microscope | FEI TITAN KRIOS |
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Image recording | Film or detector model: GATAN K3 (6k x 4k) / Average electron dose: 52.6 e/Å2 |
Electron beam | Acceleration voltage: 300 kV / Electron source: FIELD EMISSION GUN |
Electron optics | Illumination mode: FLOOD BEAM / Imaging mode: BRIGHT FIELD / Nominal defocus max: 2.5 µm / Nominal defocus min: 0.9 µm / Nominal magnification: 130000 |
Sample stage | Cooling holder cryogen: NITROGEN |
Experimental equipment | Model: Titan Krios / Image courtesy: FEI Company |
-Image processing
Final reconstruction | Applied symmetry - Helical parameters - Δz: 16.1 Å Applied symmetry - Helical parameters - Δ&Phi: 56.2 ° Applied symmetry - Helical parameters - Axial symmetry: C1 (asymmetric) Resolution.type: BY AUTHOR / Resolution: 2.61 Å / Resolution method: FSC 0.143 CUT-OFF / Software - Name: RELION (ver. 3.1) / Number images used: 1 |
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Startup model | Type of model: OTHER |
Final angle assignment | Type: NOT APPLICABLE / Software - Name: RELION (ver. 3.1) |
-Atomic model buiding 1
Refinement | Space: REAL / Protocol: OTHER |
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Output model | PDB-8pbc: |