+Open data
-Basic information
Entry | Database: EMDB / ID: EMD-4410 | |||||||||
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Title | Yeast RPA bound to ssDNA | |||||||||
Map data | 4.7 Angstrom map (sharpened) corresponding to Yeast RPA Trimerisation Core (Tri-C) bound by ssDNA. | |||||||||
Sample |
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Keywords | Complex / heterotrimer / DNA binding / OB-fold / DNA BINDING PROTEIN | |||||||||
Function / homology | Function and homology information heteroduplex formation / sporulation / DNA replication factor A complex / Gap-filling DNA repair synthesis and ligation in GG-NER / telomere maintenance via telomere lengthening / Removal of the Flap Intermediate / telomere maintenance via recombination / Translesion Synthesis by POLH / Activation of the pre-replicative complex / Translesion synthesis by REV1 ...heteroduplex formation / sporulation / DNA replication factor A complex / Gap-filling DNA repair synthesis and ligation in GG-NER / telomere maintenance via telomere lengthening / Removal of the Flap Intermediate / telomere maintenance via recombination / Translesion Synthesis by POLH / Activation of the pre-replicative complex / Translesion synthesis by REV1 / Translesion synthesis by POLK / Translesion synthesis by POLI / single-stranded telomeric DNA binding / Activation of ATR in response to replication stress / mitotic recombination / Termination of translesion DNA synthesis / reciprocal meiotic recombination / DNA unwinding involved in DNA replication / Gap-filling DNA repair synthesis and ligation in TC-NER / DNA topological change / Dual incision in TC-NER / telomere maintenance via telomerase / telomere maintenance / condensed nuclear chromosome / nucleotide-excision repair / double-strand break repair via homologous recombination / establishment of protein localization / site of double-strand break / single-stranded DNA binding / double-stranded DNA binding / DNA replication / sequence-specific DNA binding / chromosome, telomeric region / damaged DNA binding / protein ubiquitination / DNA repair / mRNA binding / nucleus / metal ion binding / cytoplasm / cytosol Similarity search - Function | |||||||||
Biological species | Saccharomyces cerevisiae (strain ATCC 204508 / S288c) (yeast) / synthetic construct (others) | |||||||||
Method | single particle reconstruction / cryo EM / Resolution: 4.7 Å | |||||||||
Authors | Yates LA / Aramayo RJ | |||||||||
Funding support | United Kingdom, 1 items
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Citation | Journal: Nat Commun / Year: 2018 Title: A structural and dynamic model for the assembly of Replication Protein A on single-stranded DNA. Authors: Luke A Yates / Ricardo J Aramayo / Nilisha Pokhrel / Colleen C Caldwell / Joshua A Kaplan / Rajika L Perera / Maria Spies / Edwin Antony / Xiaodong Zhang / Abstract: Replication Protein A (RPA), the major eukaryotic single stranded DNA-binding protein, binds to exposed ssDNA to protect it from nucleases, participates in a myriad of nucleic acid transactions and ...Replication Protein A (RPA), the major eukaryotic single stranded DNA-binding protein, binds to exposed ssDNA to protect it from nucleases, participates in a myriad of nucleic acid transactions and coordinates the recruitment of other important players. RPA is a heterotrimer and coats long stretches of single-stranded DNA (ssDNA). The precise molecular architecture of the RPA subunits and its DNA binding domains (DBDs) during assembly is poorly understood. Using cryo electron microscopy we obtained a 3D reconstruction of the RPA trimerisation core bound with ssDNA (∼55 kDa) at ∼4.7 Å resolution and a dimeric RPA assembly on ssDNA. FRET-based solution studies reveal dynamic rearrangements of DBDs during coordinated RPA binding and this activity is regulated by phosphorylation at S178 in RPA70. We present a structural model on how dynamic DBDs promote the cooperative assembly of multiple RPAs on long ssDNA. | |||||||||
History |
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-Structure visualization
Movie |
Movie viewer |
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Structure viewer | EM map: SurfViewMolmilJmol/JSmol |
Supplemental images |
-Downloads & links
-EMDB archive
Map data | emd_4410.map.gz | 6.2 MB | EMDB map data format | |
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Header (meta data) | emd-4410-v30.xml emd-4410.xml | 25.7 KB 25.7 KB | Display Display | EMDB header |
FSC (resolution estimation) | emd_4410_fsc_1.xml emd_4410_fsc_2.xml emd_4410_fsc_3.xml emd_4410_fsc_4.xml emd_4410_fsc_5.xml | 10 KB 3 KB 3 KB 3 KB 4.4 KB | Display Display Display Display Display | FSC data file |
Images | emd_4410.png | 95.6 KB | ||
Filedesc metadata | emd-4410.cif.gz | 6.8 KB | ||
Others | emd_4410_additional_1.map.gz emd_4410_additional_2.map.gz emd_4410_additional_3.map.gz emd_4410_additional_4.map.gz | 49.4 MB 1.8 MB 1.8 MB 1.8 MB | ||
Archive directory | http://ftp.pdbj.org/pub/emdb/structures/EMD-4410 ftp://ftp.pdbj.org/pub/emdb/structures/EMD-4410 | HTTPS FTP |
-Validation report
Summary document | emd_4410_validation.pdf.gz | 489.5 KB | Display | EMDB validaton report |
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Full document | emd_4410_full_validation.pdf.gz | 489.1 KB | Display | |
Data in XML | emd_4410_validation.xml.gz | 5.1 KB | Display | |
Data in CIF | emd_4410_validation.cif.gz | 5.7 KB | Display | |
Arichive directory | https://ftp.pdbj.org/pub/emdb/validation_reports/EMD-4410 ftp://ftp.pdbj.org/pub/emdb/validation_reports/EMD-4410 | HTTPS FTP |
-Related structure data
Related structure data | 6i52MC M: atomic model generated by this map C: citing same article (ref.) |
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Similar structure data |
-Links
EMDB pages | EMDB (EBI/PDBe) / EMDataResource |
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Related items in Molecule of the Month |
-Map
File | Download / File: emd_4410.map.gz / Format: CCP4 / Size: 6.6 MB / Type: IMAGE STORED AS FLOATING POINT NUMBER (4 BYTES) | ||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||
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Annotation | 4.7 Angstrom map (sharpened) corresponding to Yeast RPA Trimerisation Core (Tri-C) bound by ssDNA. | ||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||
Voxel size | X=Y=Z: 1.06 Å | ||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||
Density |
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Symmetry | Space group: 1 | ||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||
Details | EMDB XML:
CCP4 map header:
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-Supplemental data
-Additional map: 7.5 Angstrom map (unsharpened) corresponding to yeast RPA dimer
File | emd_4410_additional_1.map | ||||||||||||
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Annotation | 7.5 Angstrom map (unsharpened) corresponding to yeast RPA dimer | ||||||||||||
Projections & Slices |
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Density Histograms |
-Additional map: 7.7 Angstrom map (sharpened) corresponding to Yeast RPA...
File | emd_4410_additional_2.map | ||||||||||||
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Annotation | 7.7 Angstrom map (sharpened) corresponding to Yeast RPA Trimerisation Core (Tri-C) bound by ssDNA with additional DNA-binding domain (DBD-B) | ||||||||||||
Projections & Slices |
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Density Histograms |
-Additional map: 8.5 Angstrom map (sharpened) corresponding to Yeast RPA...
File | emd_4410_additional_3.map | ||||||||||||
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Annotation | 8.5 Angstrom map (sharpened) corresponding to Yeast RPA Trimerisation Core (Tri-C) bound by ssDNA with additional DNA-binding domain (DBD-B) | ||||||||||||
Projections & Slices |
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Density Histograms |
-Additional map: 10 Angstrom map (sharpened) corresponding to Yeast RPA...
File | emd_4410_additional_4.map | ||||||||||||
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Annotation | 10 Angstrom map (sharpened) corresponding to Yeast RPA Trimerisation Core (Tri-C) bound by ssDNA with additional DNA-binding domain (DBD-B) | ||||||||||||
Projections & Slices |
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Density Histograms |
-Sample components
-Entire : Replication Protein A heterotrimeric complex with ssDNA
Entire | Name: Replication Protein A heterotrimeric complex with ssDNA |
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Components |
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-Supramolecule #1: Replication Protein A heterotrimeric complex with ssDNA
Supramolecule | Name: Replication Protein A heterotrimeric complex with ssDNA type: complex / ID: 1 / Parent: 0 / Macromolecule list: all |
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Molecular weight | Theoretical: 120 KDa |
-Supramolecule #2: Replication Protein A heterotrimeric complex with ssDNA
Supramolecule | Name: Replication Protein A heterotrimeric complex with ssDNA type: complex / ID: 2 / Parent: 1 / Macromolecule list: #1-#3 |
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Source (natural) | Organism: Saccharomyces cerevisiae (strain ATCC 204508 / S288c) (yeast) Location in cell: nucleus |
-Supramolecule #3: Replication Protein A heterotrimeric complex with ssDNA
Supramolecule | Name: Replication Protein A heterotrimeric complex with ssDNA type: complex / ID: 3 / Parent: 1 / Macromolecule list: #4 |
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Source (natural) | Organism: synthetic construct (others) |
-Macromolecule #1: Replication factor A protein 3
Macromolecule | Name: Replication factor A protein 3 / type: protein_or_peptide / ID: 1 / Number of copies: 1 / Enantiomer: LEVO |
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Source (natural) | Organism: Saccharomyces cerevisiae (strain ATCC 204508 / S288c) (yeast) |
Molecular weight | Theoretical: 13.827723 KDa |
Recombinant expression | Organism: Escherichia coli BL21(DE3) (bacteria) |
Sequence | String: MASETPRVDP TEISNVNAPV FRIIAQIKSQ PTESQLILQS PTISSKNGSE VEMITLNNIR VSMNKTFEID SWYEFVCRNN DDGELGFLI LDAVLCKFKE NEDLSLNGVV ALQRLCKKYP EIY UniProtKB: Replication factor A protein 3 |
-Macromolecule #2: Replication factor A protein 2
Macromolecule | Name: Replication factor A protein 2 / type: protein_or_peptide / ID: 2 / Number of copies: 1 / Enantiomer: LEVO |
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Source (natural) | Organism: Saccharomyces cerevisiae (strain ATCC 204508 / S288c) (yeast) |
Molecular weight | Theoretical: 14.80987 KDa |
Recombinant expression | Organism: Escherichia coli BL21(DE3) (bacteria) |
Sequence | String: TNTRVNTLTP VTIKQILESK QDIQDGPFVS HNQELHHVCF VGVVRNITDH TANIFLTIED GTGQIEVRKW SEDAAQQFEI GGYVKVFGA LKEFGGKKNI QYAVIKPIDS FNEVLTHHLE VIKCHSIASG MMK UniProtKB: Replication factor A protein 2 |
-Macromolecule #3: Replication factor A protein 1
Macromolecule | Name: Replication factor A protein 1 / type: protein_or_peptide / ID: 3 / Number of copies: 1 / Enantiomer: LEVO |
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Source (natural) | Organism: Saccharomyces cerevisiae (strain ATCC 204508 / S288c) (yeast) |
Molecular weight | Theoretical: 20.643072 KDa |
Recombinant expression | Organism: Escherichia coli BL21(DE3) (bacteria) |
Sequence | String: KFIAQRITIA RAQAENLGRS EKGDFFSVKA AISFLKVDNF AYPACSNENC NKKVLEQPDG TWRCEKCDTN NARPNWRYIL TISIIDETN QLWLTLFDDQ AKQLLGVDAN TLMSLKEEDP NEFTKITQSI QMNEYDFRIR AREDTYNDQS RIRYTVANLH S LNYRAEAD YLADELSKAL UniProtKB: Replication factor A protein 1 |
-Macromolecule #4: DNA (5'-D(P*TP*TP*TP*TP*TP*TP*TP*TP*TP*TP*TP*TP*TP*TP*TP*TP*TP*TP...
Macromolecule | Name: DNA (5'-D(P*TP*TP*TP*TP*TP*TP*TP*TP*TP*TP*TP*TP*TP*TP*TP*TP*TP*TP*TP*T)-3') type: dna / ID: 4 / Number of copies: 1 / Classification: DNA |
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Source (natural) | Organism: synthetic construct (others) |
Molecular weight | Theoretical: 6.038899 KDa |
Sequence | String: (DT)(DT)(DT)(DT)(DT)(DT)(DT)(DT)(DT)(DT) (DT)(DT)(DT)(DT)(DT)(DT)(DT)(DT)(DT)(DT) |
-Experimental details
-Structure determination
Method | cryo EM |
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Processing | single particle reconstruction |
Aggregation state | particle |
-Sample preparation
Concentration | 0.5 mg/mL | ||||||||||||
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Buffer | pH: 7.4 Component:
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Grid | Model: Quantifoil R2/2 / Material: COPPER / Mesh: 300 / Pretreatment - Type: GLOW DISCHARGE / Pretreatment - Time: 30 sec. / Pretreatment - Atmosphere: AIR | ||||||||||||
Vitrification | Cryogen name: ETHANE / Chamber humidity: 100 % / Chamber temperature: 277 K / Instrument: FEI VITROBOT MARK IV Details: glow-discharged grid, wait time 30 seconds, blotting time 1 second. |
-Electron microscopy
Microscope | FEI TITAN KRIOS |
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Image recording | Film or detector model: GATAN K2 SUMMIT (4k x 4k) / Detector mode: COUNTING / Average electron dose: 2.1 e/Å2 |
Electron beam | Acceleration voltage: 300 kV / Electron source: FIELD EMISSION GUN |
Electron optics | Illumination mode: FLOOD BEAM / Imaging mode: BRIGHT FIELD / Cs: 2.7 mm |
Sample stage | Specimen holder model: FEI TITAN KRIOS AUTOGRID HOLDER / Cooling holder cryogen: NITROGEN |
Experimental equipment | Model: Titan Krios / Image courtesy: FEI Company |
+Image processing
-Atomic model buiding 1
Refinement | Space: REAL / Protocol: FLEXIBLE FIT |
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Output model | PDB-6i52: |