+Open data
-Basic information
Entry | Database: EMDB / ID: EMD-5118 | |||||||||
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Title | Clathrin D6 coat with Hsc70 and Auxilin | |||||||||
Map data | This is a volume of a D6 clathrin coat bound with Hsc70(1-554) and Auxilin(547-910) | |||||||||
Sample |
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Keywords | clathrin / D6 coat / Hsc70 / Auxilin / uncoating | |||||||||
Biological species | Bos taurus (cattle) | |||||||||
Method | single particle reconstruction / cryo EM / Resolution: 15.2 Å | |||||||||
Authors | Xing Y / Boecking T / Wolf M / Kirchhausen T / Harrison SC | |||||||||
Citation | Journal: EMBO J / Year: 2010 Title: Structure of clathrin coat with bound Hsc70 and auxilin: mechanism of Hsc70-facilitated disassembly. Authors: Yi Xing / Till Böcking / Matthias Wolf / Nikolaus Grigorieff / Tomas Kirchhausen / Stephen C Harrison / Abstract: The chaperone Hsc70 drives the clathrin assembly-disassembly cycle forward by stimulating dissociation of a clathrin lattice. A J-domain containing co-chaperone, auxilin, associates with a freshly ...The chaperone Hsc70 drives the clathrin assembly-disassembly cycle forward by stimulating dissociation of a clathrin lattice. A J-domain containing co-chaperone, auxilin, associates with a freshly budded clathrin-coated vesicle, or with an in vitro assembled clathrin coat, and recruits Hsc70 to its specific heavy-chain-binding site. We have determined by electron cryomicroscopy (cryoEM), at about 11 A resolution, the structure of a clathrin coat (in the D6-barrel form) with specifically bound Hsc70 and auxilin. The Hsc70 binds a previously analysed site near the C-terminus of the heavy chain, with a stoichiometry of about one per three-fold vertex. Its binding is accompanied by a distortion of the clathrin lattice, detected by a change in the axial ratio of the D6 barrel. We propose that when Hsc70, recruited to a position close to its target by the auxilin J-domain, splits ATP, it clamps firmly onto its heavy-chain site and locks in place a transient fluctuation. Accumulation of the local strain thus imposed at multiple vertices can then lead to disassembly. | |||||||||
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_5118.map.gz | 12.8 MB | EMDB map data format | |
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Header (meta data) | emd-5118-v30.xml emd-5118.xml | 10.4 KB 10.4 KB | Display Display | EMDB header |
Images | emd_5118_1.tif | 6.1 MB | ||
Archive directory | http://ftp.pdbj.org/pub/emdb/structures/EMD-5118 ftp://ftp.pdbj.org/pub/emdb/structures/EMD-5118 | HTTPS FTP |
-Validation report
Summary document | emd_5118_validation.pdf.gz | 78.5 KB | Display | EMDB validaton report |
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Full document | emd_5118_full_validation.pdf.gz | 77.6 KB | Display | |
Data in XML | emd_5118_validation.xml.gz | 494 B | Display | |
Arichive directory | https://ftp.pdbj.org/pub/emdb/validation_reports/EMD-5118 ftp://ftp.pdbj.org/pub/emdb/validation_reports/EMD-5118 | 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_5118.map.gz / Format: CCP4 / Size: 62.5 MB / Type: IMAGE STORED AS FLOATING POINT NUMBER (4 BYTES) | ||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||
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Annotation | This is a volume of a D6 clathrin coat bound with Hsc70(1-554) and Auxilin(547-910) | ||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||
Projections & slices | Image control
Images are generated by Spider. | ||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||
Voxel size | X=Y=Z: 4.2 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 : Clathrin coats bound with Hsc70(1-554) and Auxilin(547-910)
Entire | Name: Clathrin coats bound with Hsc70(1-554) and Auxilin(547-910) |
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Components |
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-Supramolecule #1000: Clathrin coats bound with Hsc70(1-554) and Auxilin(547-910)
Supramolecule | Name: Clathrin coats bound with Hsc70(1-554) and Auxilin(547-910) type: sample / ID: 1000 Details: clathrin coats assembled from clathrin and AP-2 with excess of Hsc70(1-554) and Auxilin(547-910) added Number unique components: 9 |
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-Macromolecule #1: uncoating intermediate of clathrin coat
Macromolecule | Name: uncoating intermediate of clathrin coat / type: protein_or_peptide / ID: 1 Name.synonym: clathrin coat in complex with Hsc70 and auxilin Number of copies: 108 / Oligomeric state: D6 assemble / Recombinant expression: No / Database: NCBI |
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Source (natural) | Organism: Bos taurus (cattle) / synonym: cattle / Tissue: Brain |
-Experimental details
-Structure determination
Method | cryo EM |
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Processing | single particle reconstruction |
Aggregation state | particle |
-Sample preparation
Concentration | 0.25 mg/mL |
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Buffer | pH: 6 Details: 20mM MES, pH6.0, 2mM MgCl2, 25mM KCl, 10mM (NH4)2SO4, and 2mM DTT |
Grid | Details: holey carbon grid |
Vitrification | Cryogen name: ETHANE / Chamber humidity: 60 % / Chamber temperature: 93 K / Instrument: OTHER / Details: Vitrification instrument: FEI Vitrobot Timed resolved state: Vitrified 30 msec after spraying with effector Method: Blot for 2 seconds before plunging |
-Electron microscopy
Microscope | FEI TECNAI F20 |
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Temperature | Average: 93 K |
Date | Jan 1, 2007 |
Image recording | Category: FILM / Film or detector model: KODAK SO-163 FILM / Digitization - Scanner: ZEISS SCAI / Digitization - Sampling interval: 7 µm / Number real images: 900 / Average electron dose: 20 e/Å2 / Od range: 1.4 / Bits/pixel: 8 |
Electron beam | Acceleration voltage: 200 kV / Electron source: FIELD EMISSION GUN |
Electron optics | Calibrated magnification: 51159 / Illumination mode: FLOOD BEAM / Imaging mode: BRIGHT FIELD / Cs: 2 mm / Nominal defocus max: 5.0 µm / Nominal defocus min: 2.0 µm / Nominal magnification: 50000 |
Sample stage | Specimen holder: side entry liquid nitrogen-cooled cryo specimen holder. Specimen holder model: GATAN LIQUID NITROGEN |
Experimental equipment | Model: Tecnai F20 / Image courtesy: FEI Company |
-Image processing
CTF correction | Details: CTF correction of each particle. |
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Final reconstruction | Algorithm: OTHER / Resolution.type: BY AUTHOR / Resolution: 15.2 Å / Resolution method: FSC 0.143 CUT-OFF / Software - Name: Frealign Details: a raw map without B factor sharpening and n.c.s averaging Number images used: 1500 |