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Yorodumi- EMDB-1668: Cryo-EM structure of the active yeast 80S ribosome bearing a P-si... -
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-Basic information
Entry | Database: EMDB / ID: EMD-1668 | |||||||||
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Title | Cryo-EM structure of the active yeast 80S ribosome bearing a P-site tRNA and with the rRNA expansion segment ES27 in the exit conformation | |||||||||
Map data | This map represents a yeast 80S ribosome programmed with the first 120 amino acids of the type I signal anchor protein DPAP-B attached to a P-site tRNA. The expansion segment ES27 is in the exit position. | |||||||||
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: 7.9 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_1668.map.gz | 18.8 MB | EMDB map data format | |
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Header (meta data) | emd-1668-v30.xml emd-1668.xml | 11.3 KB 11.3 KB | Display Display | EMDB header |
Images | 1668.gif 1668_EMD_1668_ES27_exit_yeast.jpg | 93.6 KB 236 KB | ||
Archive directory | http://ftp.pdbj.org/pub/emdb/structures/EMD-1668 ftp://ftp.pdbj.org/pub/emdb/structures/EMD-1668 | HTTPS FTP |
-Related structure data
Related structure data | 1651C 1652C 1667C 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_1668.map.gz / Format: CCP4 / Size: 185.7 MB / Type: IMAGE STORED AS FLOATING POINT NUMBER (4 BYTES) | ||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||
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Annotation | This map represents a yeast 80S ribosome programmed with the first 120 amino acids of the type I signal anchor protein DPAP-B attached to a P-site tRNA. The expansion segment ES27 is in the exit position. | ||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||
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 : A programmed yeast ribosome with ES27 in the exit conformation
Entire | Name: A programmed yeast ribosome with ES27 in the exit conformation |
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Components |
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-Supramolecule #1000: A programmed yeast ribosome with ES27 in the exit conformation
Supramolecule | Name: A programmed yeast ribosome with ES27 in the exit conformation 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 Recombinant expression: No / Ribosome-details: ribosome-eukaryote: ALL |
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Source (natural) | Organism: Saccharomyces cerevisiae (brewer's yeast) / synonym: Baker's Yeast |
Molecular weight | Experimental: 4.2 MDa / Theoretical: 4.2 MDa |
-Experimental details
-Structure determination
Method | negative staining, cryo EM |
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Processing | single particle reconstruction |
Aggregation state | particle |
-Sample preparation
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 |
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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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Electron beam | Acceleration voltage: 300 kV / Electron source: FIELD EMISSION GUN |
Electron optics | Calibrated magnification: 38000 / Illumination mode: FLOOD BEAM / Imaging mode: BRIGHT FIELDBright-field microscopy / 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 |
Temperature | Average: 84 K |
Alignment procedure | Legacy - Astigmatism: Objective lens astigmatism 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 |
Experimental equipment | Model: Tecnai Polara / Image courtesy: FEI Company |
-Image processing
CTF correction | Details: Defocus group volumes |
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Final angle assignment | Details: SPIDER |
Final reconstruction | Applied symmetry - Point group: C1 (asymmetric) / Algorithm: OTHER / Resolution.type: BY AUTHOR / Resolution: 7.9 Å / Resolution method: FSC 0.5 CUT-OFF / Software - Name: SPIDER Details: Map was filtered between 8.3 and 10.3 Angstrom to better visualize the rRNA segment ES27 in the exit conformation Number images used: 69000 |
Details | Particles were selected using the program SIGNATURE and visually inspected. This map resulted from sorting against the ES27 exit position and subsequent sorting for tRNA and the Ssh1 complex and represents the datasubset with ES27 in the exit conformation |