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Yorodumi- EMDB-1651: Cryo-EM structure of the programmed yeast 80 ribosome bound the S... -
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-Basic information
Entry | Database: EMDB / ID: EMD-1651 | |||||||||
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Title | Cryo-EM structure of the programmed yeast 80 ribosome bound the Ssh1 complex | |||||||||
Map data | This map is derived from all particles representing a yeast 80S ribosome bound the Ssh1 complex in a unratcheted conformation before sorting for the conformation of ES27, and the presence of Ssh and tRNA (see also Fig S1 and Fig S2A in the accompanying paper). | |||||||||
Sample |
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Keywords | Ribosome / protein exit tunnel / cotranslational protein translocation / protein conducting channel / signal sequence | |||||||||
Biological species | Saccharomyces cerevisiae (brewer's yeast) | |||||||||
Method | single particle reconstruction / cryo EM / negative staining / Resolution: 6.1 Å | |||||||||
Authors | Becker T / Mandon E / Bhushan S / Jarasch A / Armache JP / Funes S / Jossinet F / Gumbart J / Mielke T / Berninghausen O ...Becker T / Mandon E / Bhushan S / Jarasch A / Armache JP / Funes S / Jossinet F / Gumbart J / Mielke T / Berninghausen O / Schulten K / Westhof E / Gilmore R / Beckmann R | |||||||||
Citation | Journal: Science / Year: 2009 Title: Structure of monomeric yeast and mammalian Sec61 complexes interacting with the translating ribosome. Authors: Thomas Becker / Shashi Bhushan / Alexander Jarasch / Jean-Paul Armache / Soledad Funes / Fabrice Jossinet / James Gumbart / Thorsten Mielke / Otto Berninghausen / Klaus Schulten / Eric ...Authors: Thomas Becker / Shashi Bhushan / Alexander Jarasch / Jean-Paul Armache / Soledad Funes / Fabrice Jossinet / James Gumbart / Thorsten Mielke / Otto Berninghausen / Klaus Schulten / Eric Westhof / Reid Gilmore / Elisabet C Mandon / Roland Beckmann / Abstract: The trimeric Sec61/SecY complex is a protein-conducting channel (PCC) for secretory and membrane proteins. Although Sec complexes can form oligomers, it has been suggested that a single copy may ...The trimeric Sec61/SecY complex is a protein-conducting channel (PCC) for secretory and membrane proteins. Although Sec complexes can form oligomers, it has been suggested that a single copy may serve as an active PCC. We determined subnanometer-resolution cryo-electron microscopy structures of eukaryotic ribosome-Sec61 complexes. In combination with biochemical data, we found that in both idle and active states, the Sec complex is not oligomeric and interacts mainly via two cytoplasmic loops with the universal ribosomal adaptor site. In the active state, the ribosomal tunnel and a central pore of the monomeric PCC were occupied by the nascent chain, contacting loop 6 of the Sec complex. This provides a structural basis for the activity of a solitary Sec complex in cotranslational protein translocation. | |||||||||
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_1651.map.gz | 16.8 MB | EMDB map data format | |
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Header (meta data) | emd-1651-v30.xml emd-1651.xml | 12.5 KB 12.5 KB | Display Display | EMDB header |
Images | 1651.gif EMD-1651.jpg | 73.7 KB 91.5 KB | ||
Archive directory | http://ftp.pdbj.org/pub/emdb/structures/EMD-1651 ftp://ftp.pdbj.org/pub/emdb/structures/EMD-1651 | HTTPS FTP |
-Validation report
Summary document | emd_1651_validation.pdf.gz | 258.2 KB | Display | EMDB validaton report |
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Full document | emd_1651_full_validation.pdf.gz | 257.3 KB | Display | |
Data in XML | emd_1651_validation.xml.gz | 7 KB | Display | |
Arichive directory | https://ftp.pdbj.org/pub/emdb/validation_reports/EMD-1651 ftp://ftp.pdbj.org/pub/emdb/validation_reports/EMD-1651 | HTTPS FTP |
-Related structure data
Related structure data | 1652C 1667C 1668C 1669C 2ww9C 2wwaC 2wwbC 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_1651.map.gz / Format: CCP4 / Size: 185.7 MB / Type: IMAGE STORED AS FLOATING POINT NUMBER (4 BYTES) | ||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||
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Annotation | This map is derived from all particles representing a yeast 80S ribosome bound the Ssh1 complex in a unratcheted conformation before sorting for the conformation of ES27, and the presence of Ssh and tRNA (see also Fig S1 and Fig S2A in the accompanying paper). | ||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||
Voxel size | X=Y=Z: 1.2375 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 : This map represents a yeast 80S ribosome in the unratcheted confo...
Entire | Name: This map represents a yeast 80S ribosome in the unratcheted conformation with the yeast Ssh1 complex bound after sorting the entire dataset against an empty (ratcheted) yeast 80S ribosome |
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Components |
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-Supramolecule #1000: This map represents a yeast 80S ribosome in the unratcheted confo...
Supramolecule | Name: This map represents a yeast 80S ribosome in the unratcheted conformation with the yeast Ssh1 complex bound after sorting the entire dataset against an empty (ratcheted) yeast 80S ribosome type: sample / ID: 1000 Details: 80S ribosomes and the detergent solubilized Ssh1 complex were reconstituted in vitro by adding 1 pmol of ribosome and Ssh1 complex in 5 fold molar excess Oligomeric state: 80S Ribosome bound to one copy of the heterotrimeric Ssh1 complex Number unique components: 2 |
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Molecular weight | Experimental: 4.2 MDa / Theoretical: 4.2 MDa / Method: Known for 80S ribosomes |
-Supramolecule #1: Yeast 80S ribosome bound to the yeast Ssh1 complex
Supramolecule | Name: Yeast 80S ribosome bound to the yeast Ssh1 complex / type: complex / ID: 1 Name.synonym: Yeast 80S ribosome bound to the yeast Ssh1 complex Ribosome-details: ribosome-eukaryote: ALL |
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Molecular weight | Experimental: 4.2 MDa / Theoretical: 4.2 MDa |
-Macromolecule #1: Ssh1 complex
Macromolecule | Name: Ssh1 complex / type: ligand / ID: 1 / Name.synonym: Ssh1 complex / Details: His FLAG-tagged / Number of copies: 1 / Oligomeric state: heterotrimer / Recombinant expression: Yes |
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Source (natural) | Organism: Saccharomyces cerevisiae (brewer's yeast) / Strain: RGY1592 / synonym: Baker's yeast / Cell: rough microsomes / Organelle: Endoplasmic reticulum membrane / Location in cell: Endoplasmic reticulum |
Molecular weight | Theoretical: 71.5 KDa |
Recombinant expression | Organism: Saccharomyces cerevisiae (brewer's yeast) |
-Experimental details
-Structure determination
Method | negative staining, cryo EM |
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Processing | single particle reconstruction |
Aggregation state | particle |
-Sample preparation
Concentration | 0.02 mg/mL |
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Buffer | pH: 7.5 Details: 20 mM HEPES/KOH, pH 7.5 100 mM KOAc, 10 mM Mg(OAc)2, 1.5 mM DTT, 0.1 % (w/v) digitonin |
Staining | Type: NEGATIVE / Details: Cryo-EM |
Grid | Details: Quantifoil grids (3/3) with 2 nm carbon on top |
Vitrification | Cryogen name: ETHANE / Chamber humidity: 95 % / Instrument: OTHER / Details: Vitrification instrument: Vitrobot Method: Blot for 10 seconds before plunging, use 2 layer of filter paper |
-Electron microscopy
Microscope | FEI POLARA 300 |
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Temperature | Average: 84 K |
Alignment procedure | Legacy - Astigmatism: Objective lens astgnatism was corrected at 100000 times magnification |
Image recording | Category: FILM / Film or detector model: KODAK SO-163 FILM / Digitization - Scanner: PRIMESCAN / Digitization - Sampling interval: 4.76 µm / Number real images: 185 / Average electron dose: 25 e/Å2 / Details: Scanned at 5334 dpi / Od range: 1.2 / Bits/pixel: 16 |
Tilt angle min | 0 |
Tilt angle max | 0 |
Electron beam | Acceleration voltage: 300 kV / Electron source: FIELD EMISSION GUN |
Electron optics | Calibrated magnification: 38000 / Illumination mode: FLOOD BEAM / Imaging mode: BRIGHT FIELD / Cs: 2.26 mm / Nominal defocus max: 4.5 µm / Nominal defocus min: 1.2 µm / Nominal magnification: 39000 |
Sample stage | Specimen holder: FEI Polara cartridge system / Specimen holder model: OTHER |
Experimental equipment | Model: Tecnai Polara / Image courtesy: FEI Company |
-Image processing
Details | Particles were selected using the program SIGNATURE and visually inspected |
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CTF correction | Details: Defocus group volumes |
Final reconstruction | Applied symmetry - Point group: C1 (asymmetric) / Algorithm: OTHER / Resolution.type: BY AUTHOR / Resolution: 6.1 Å / Resolution method: FSC 0.5 CUT-OFF / Software - Name: SPIDER Details: The map representing all unratcheted particles is filtereb between 6.-8 Angstoms. Maps containing active and idle Ssh1 complex and the ES27 in exit-position are filtered between 8.3 and10.3 ...Details: The map representing all unratcheted particles is filtereb between 6.-8 Angstoms. Maps containing active and idle Ssh1 complex and the ES27 in exit-position are filtered between 8.3 and10.3 Angstoms to better visualize the Ssh1 complex ES27. Number images used: 183000 |
Final angle assignment | Details: SPIDER |