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Yorodumi- EMDB-1073: Three-dimensional structures of translating ribosomes by Cryo-EM. -
+Open data
-Basic information
Entry | Database: EMDB / ID: EMD-1073 | |||||||||
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Title | Three-dimensional structures of translating ribosomes by Cryo-EM. | |||||||||
Map data | Average difference map calculated between individual difference maps for Ig2 and GFP ribosome-nascent chain complexes (see entries 1068, 1070, 1071, 1072). | |||||||||
Sample |
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Biological species | Escherichia coli (E. coli) / Dictyostelium discoideum (eukaryote) | |||||||||
Method | single particle reconstruction / cryo EM / Resolution: 16.0 Å | |||||||||
Authors | Gilbert RJC / Fucini P / Connell S / Fuller SD / Nierhaus KH / Robinson CV / Dobson CM / Stuart DI | |||||||||
Citation | Journal: Mol Cell / Year: 2004 Title: Three-dimensional structures of translating ribosomes by Cryo-EM. Authors: Robert J C Gilbert / Paola Fucini / Sean Connell / Stephen D Fuller / Knud H Nierhaus / Carol V Robinson / Christopher M Dobson / David I Stuart / Abstract: Cryo-electron microscopy and image reconstruction techniques have been used to obtain three-dimensional maps for E. coli ribosomes stalled following translation of three representative proteins. ...Cryo-electron microscopy and image reconstruction techniques have been used to obtain three-dimensional maps for E. coli ribosomes stalled following translation of three representative proteins. Comparisons of these electron density maps, at resolutions of between 13 and 16 A, with that of a nontranslating ribosome pinpoint specific structural differences in stalled ribosomes and identify additional material, including tRNAs and mRNA. In addition, the tunnel through the large subunit, the anticipated exit route of newly synthesized proteins, is partially occluded in all the stalled ribosome structures. This observation suggests that significant segments of the nascent polypeptide chains examined here could be located within an expanded tunnel, perhaps in a rudimentary globular conformation. Such behavior could be an important aspect of the folding of at least some proteins in the cellular environment. | |||||||||
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_1073.map.gz | 4.8 MB | EMDB map data format | |
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Header (meta data) | emd-1073-v30.xml emd-1073.xml | 11 KB 11 KB | Display Display | EMDB header |
Images | 1073.gif | 54.8 KB | ||
Archive directory | http://ftp.pdbj.org/pub/emdb/structures/EMD-1073 ftp://ftp.pdbj.org/pub/emdb/structures/EMD-1073 | HTTPS FTP |
-Validation report
Summary document | emd_1073_validation.pdf.gz | 272.2 KB | Display | EMDB validaton report |
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Full document | emd_1073_full_validation.pdf.gz | 271.4 KB | Display | |
Data in XML | emd_1073_validation.xml.gz | 5.9 KB | Display | |
Arichive directory | https://ftp.pdbj.org/pub/emdb/validation_reports/EMD-1073 ftp://ftp.pdbj.org/pub/emdb/validation_reports/EMD-1073 | HTTPS FTP |
-Related 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_1073.map.gz / Format: CCP4 / Size: 7.8 MB / Type: IMAGE STORED AS FLOATING POINT NUMBER (4 BYTES) | ||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||
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Annotation | Average difference map calculated between individual difference maps for Ig2 and GFP ribosome-nascent chain complexes (see entries 1068, 1070, 1071, 1072). | ||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||
Voxel size | X=Y=Z: 3.33 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 : E. coli ribosome translating GFP and tandem Ig domains - differen...
Entire | Name: E. coli ribosome translating GFP and tandem Ig domains - difference map |
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Components |
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-Supramolecule #1000: E. coli ribosome translating GFP and tandem Ig domains - differen...
Supramolecule | Name: E. coli ribosome translating GFP and tandem Ig domains - difference map type: sample / ID: 1000 / Details: The sample was monodisperse. Oligomeric state: 2 tRNA molecules and the average of two nascent proteins Number unique components: 3 |
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-Macromolecule #1: P site tRNA
Macromolecule | Name: P site tRNA / type: rna / ID: 1 / Classification: OTHER / Structure: SINGLE STRANDED / Synthetic?: No |
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Source (natural) | Organism: Escherichia coli (E. coli) |
-Macromolecule #2: E site tRNA
Macromolecule | Name: E site tRNA / type: rna / ID: 2 / Classification: OTHER / Structure: SINGLE STRANDED / Synthetic?: No |
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Source (natural) | Organism: Escherichia coli (E. coli) |
-Macromolecule #3: nascent protein
Macromolecule | Name: nascent protein / type: protein_or_peptide / ID: 3 / Name.synonym: GFP-Ig2 Details: Density for nascent protein is average of that for tandem Ig domains and GFP Number of copies: 1 / Oligomeric state: monomer / Recombinant expression: Yes |
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Source (natural) | Organism: Dictyostelium discoideum (eukaryote) |
Recombinant expression | Organism: Escherichia coli (E. coli) / Recombinant plasmid: pT7-7 |
-Experimental details
-Structure determination
Method | cryo EM |
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Processing | single particle reconstruction |
Aggregation state | particle |
-Sample preparation
Concentration | 5.0 mg/mL |
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Buffer | Details: 20 mM HEPES, 150 mM ammonium acetate, 6 mM magnesium acetate, 2 mM spermidine, 0.05 mM spermine and 4 mM 2-mercaptoethanol. Concentration of ribosomes expressed to A260 units. |
Grid | Details: 300 mesh copper grid with holey carbon film |
Vitrification | Cryogen name: ETHANE / Chamber temperature: 100 K / Instrument: HOMEMADE PLUNGER Details: Vitrification instrument: Standard unmodified guillotine plunger |
-Electron microscopy
Microscope | FEI/PHILIPS CM200FEG |
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Temperature | Average: 100 K |
Date | May 2, 2000 |
Image recording | Category: FILM / Film or detector model: KODAK SO-163 FILM / Digitization - Scanner: OTHER / Digitization - Sampling interval: 8.322 µm / Details: Scanner model: UMAX PowerLook 3000 / Od range: 5 / Bits/pixel: 8 |
Electron beam | Acceleration voltage: 200 kV / Electron source: FIELD EMISSION GUN |
Electron optics | Illumination mode: OTHER / Imaging mode: OTHER / Cs: 2 mm / Nominal magnification: 50000 |
Sample stage | Specimen holder: Eucentric / Specimen holder model: GATAN LIQUID NITROGEN |
-Image processing
CTF correction | Details: Each negative dataset |
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Final reconstruction | Applied symmetry - Point group: C1 (asymmetric) / Algorithm: OTHER / Resolution.type: BY AUTHOR / Resolution: 16.0 Å / Resolution method: FSC 0.5 CUT-OFF / Software - Name: SPIDER, IMAGIC, GAP, CNS, XPLOR Details: See entries 1068, 1070, 1071 and 1072. Vector difference maps were calculated between reconstructions of each nascent protein-ribosome complex and the control inactive map. The difference ...Details: See entries 1068, 1070, 1071 and 1072. Vector difference maps were calculated between reconstructions of each nascent protein-ribosome complex and the control inactive map. The difference densities for the tandem Ig domains and GFP nascent proteins were more similar to each other than either was to the single Ig domain nascent protein. Therefore these two difference maps were averaged together to produce a set of consensus difference densities representing the main differences between inactive ribosomes and ribosomes stalled during translation that contain a nascent polypeptide chain. |
Final angle assignment | Details: SPIDER euler |
-Atomic model buiding 1
Software | Name: GAP |
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Details | Protocol: Rigid body |
Refinement | Space: REAL / Protocol: RIGID BODY FIT / Target criteria: R-factor and correlation coefficient |