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Yorodumi- EMDB-5381: Cdc6-induced Conformational Changes in ORC Bound To Origin DNA Re... -
+Open data
-Basic information
Entry | Database: EMDB / ID: EMD-5381 | |||||||||
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Title | Cdc6-induced Conformational Changes in ORC Bound To Origin DNA Revealed by Cryo-Electron Microscopy. This map may be mirrored (based on comparison to EMD-5625). | |||||||||
Map data | cryoEM structure of Yeast Origin Recognition complex with dsDNA and Cdc6. This map may be mirrored (based on comparison to EMD-5625). | |||||||||
Sample |
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Keywords | ORC / Cdc6 | |||||||||
Biological species | Saccharomyces cerevisiae (brewer's yeast) | |||||||||
Method | single particle reconstruction / cryo EM / Resolution: 15.0 Å | |||||||||
Authors | Sun J / Kawakami H / Zech J / Speck C / Stillman B / Li H | |||||||||
Citation | Journal: Structure / Year: 2012 Title: Cdc6-induced conformational changes in ORC bound to origin DNA revealed by cryo-electron microscopy. Authors: Jingchuan Sun / Hironori Kawakami / Juergen Zech / Christian Speck / Bruce Stillman / Huilin Li / Abstract: The eukaryotic origin recognition complex (ORC) interacts with and remodels origins of DNA replication prior to initiation in S phase. Here, we report a single-particle cryo-EM-derived structure of ...The eukaryotic origin recognition complex (ORC) interacts with and remodels origins of DNA replication prior to initiation in S phase. Here, we report a single-particle cryo-EM-derived structure of the supramolecular assembly comprising Saccharomyces cerevisiae ORC, the replication initiation factor Cdc6, and double-stranded ARS1 origin DNA in the presence of ATPγS. The six subunits of ORC are arranged as Orc1:Orc4:Orc5:Orc2:Orc3, with Orc6 binding to Orc2. Cdc6 binding changes the conformation of ORC, in particular reorienting the Orc1 N-terminal BAH domain. Segmentation of the 3D map of ORC-Cdc6 on DNA and docking with the crystal structure of the homologous archaeal Orc1/Cdc6 protein suggest an origin DNA binding model in which the DNA tracks along the interior surface of the crescent-like ORC. Thus, ORC bends and wraps the DNA. This model is consistent with the observation that binding of a single Cdc6 extends the ORC footprint on origin DNA from both ends. | |||||||||
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_5381.map.gz | 390.2 KB | EMDB map data format | |
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Header (meta data) | emd-5381-v30.xml emd-5381.xml | 9.6 KB 9.6 KB | Display Display | EMDB header |
Images | emd_5381_1.jpg | 27.3 KB | ||
Archive directory | http://ftp.pdbj.org/pub/emdb/structures/EMD-5381 ftp://ftp.pdbj.org/pub/emdb/structures/EMD-5381 | HTTPS FTP |
-Validation report
Summary document | emd_5381_validation.pdf.gz | 78.2 KB | Display | EMDB validaton report |
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Full document | emd_5381_full_validation.pdf.gz | 77.3 KB | Display | |
Data in XML | emd_5381_validation.xml.gz | 493 B | Display | |
Arichive directory | https://ftp.pdbj.org/pub/emdb/validation_reports/EMD-5381 ftp://ftp.pdbj.org/pub/emdb/validation_reports/EMD-5381 | HTTPS FTP |
-Related structure data
Similar structure data |
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-Links
EMDB pages | EMDB (EBI/PDBe) / EMDataResource |
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-Map
File | Download / File: emd_5381.map.gz / Format: CCP4 / Size: 422.9 KB / Type: IMAGE STORED AS FLOATING POINT NUMBER (4 BYTES) | ||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||
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Annotation | cryoEM structure of Yeast Origin Recognition complex with dsDNA and Cdc6. This map may be mirrored (based on comparison to EMD-5625). | ||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||
Voxel size | X=Y=Z: 4.23 Å | ||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||
Density |
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Symmetry | Space group: 1 | ||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||
Details | EMDB XML:
CCP4 map header:
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-Supplemental data
-Sample components
-Entire : Yeast Origin Recognition Complex with dsDNA and Cdc6
Entire | Name: Yeast Origin Recognition Complex with dsDNA and Cdc6 |
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Components |
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-Supramolecule #1000: Yeast Origin Recognition Complex with dsDNA and Cdc6
Supramolecule | Name: Yeast Origin Recognition Complex with dsDNA and Cdc6 / type: sample / ID: 1000 Details: ORC and Cdc6 are mixed with 66 bp DNA with ARS1 in ATPrS Number unique components: 8 |
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Molecular weight | Experimental: 500 KDa / Theoretical: 500 KDa |
-Macromolecule #1: Yeast Origin Recognition Complex and Cdc6
Macromolecule | Name: Yeast Origin Recognition Complex and Cdc6 / type: protein_or_peptide / ID: 1 / Name.synonym: ORC, Cdc6 / Number of copies: 1 / Recombinant expression: Yes |
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Source (natural) | Organism: Saccharomyces cerevisiae (brewer's yeast) / synonym: ORC, Cdc6 |
Molecular weight | Experimental: 500 KDa / Theoretical: 500 KDa |
Recombinant expression | Organism: Saccharomyces cerevisiae (brewer's yeast) / Recombinant plasmid: pCITE-2a |
-Experimental details
-Structure determination
Method | cryo EM |
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Processing | single particle reconstruction |
Aggregation state | particle |
-Sample preparation
Concentration | 0.4 mg/mL |
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Buffer | pH: 7.5 Details: 50 mM HEPES/KOH, pH 7.5, 1 mM EDTA, 1 mM EGTA, 1 mM DTT, 100 mM KGlu |
Grid | Details: 300 mesh lacey grid with thin continuous carbon |
Vitrification | Cryogen name: ETHANE / Chamber humidity: 70 % / Instrument: OTHER / Details: Vitrification instrument: FEI Vitrobot / Method: Blot 5 seconds |
-Electron microscopy
Microscope | JEOL 2010F |
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Image recording | Category: FILM / Film or detector model: KODAK SO-163 FILM / Digitization - Scanner: NIKON COOLSCAN / Digitization - Sampling interval: 6.35 µm / Average electron dose: 15 e/Å2 / Bits/pixel: 8 |
Electron beam | Acceleration voltage: 200 kV / Electron source: FIELD EMISSION GUN |
Electron optics | Calibrated magnification: 60000 / Illumination mode: FLOOD BEAM / Imaging mode: BRIGHT FIELD / Cs: 2 mm / Nominal defocus max: 8.0 µm / Nominal defocus min: 3.0 µm / Nominal magnification: 60000 |
Sample stage | Specimen holder: Gatan 626200 / Specimen holder model: GATAN LIQUID NITROGEN |
-Image processing
CTF correction | Details: Each particle set |
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Final reconstruction | Algorithm: OTHER / Resolution.type: BY AUTHOR / Resolution: 15.0 Å / Resolution method: FSC 0.5 CUT-OFF / Software - Name: EMAN1.8 / Number images used: 54000 |