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Yorodumi- EMDB-2623: Sampling intermediate of Regulation of the mammalian elongation c... -
+Open data
-Basic information
Entry | Database: EMDB / ID: EMD-2623 | |||||||||
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Title | Sampling intermediate of Regulation of the mammalian elongation cycle by 40S subunit rolling: a eukaryotic-specific ribosome rearrangement | |||||||||
Map data | reconstruction of initial codon-sampling state of 80S | |||||||||
Sample |
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Keywords | translation / mammalian 80S ribosome / elongation cycle / tRNA selection / eukaryotic ternary complex / elongation factor eEF1A / cryo-electron microscopy | |||||||||
Function / homology | Function and homology information : / guanyl nucleotide binding / sulfur compound metabolic process / positive regulation by host of viral genome replication / Formation of the ternary complex, and subsequently, the 43S complex / Ribosomal scanning and start codon recognition / Translation initiation complex formation / cortical actin cytoskeleton / Protein hydroxylation / SARS-CoV-1 modulates host translation machinery ...: / guanyl nucleotide binding / sulfur compound metabolic process / positive regulation by host of viral genome replication / Formation of the ternary complex, and subsequently, the 43S complex / Ribosomal scanning and start codon recognition / Translation initiation complex formation / cortical actin cytoskeleton / Protein hydroxylation / SARS-CoV-1 modulates host translation machinery / Peptide chain elongation / Selenocysteine synthesis / Formation of a pool of free 40S subunits / Eukaryotic Translation Termination / Response of EIF2AK4 (GCN2) to amino acid deficiency / SRP-dependent cotranslational protein targeting to membrane / Viral mRNA Translation / Nonsense Mediated Decay (NMD) independent of the Exon Junction Complex (EJC) / GTP hydrolysis and joining of the 60S ribosomal subunit / L13a-mediated translational silencing of Ceruloplasmin expression / Major pathway of rRNA processing in the nucleolus and cytosol / translation elongation factor activity / Nonsense Mediated Decay (NMD) enhanced by the Exon Junction Complex (EJC) / stress granule assembly / rough endoplasmic reticulum / cytosolic ribosome / cellular response to epidermal growth factor stimulus / small-subunit processome / maintenance of translational fidelity / ruffle membrane / Regulation of expression of SLITs and ROBOs / ribosomal small subunit biogenesis / Hydrolases; Acting on acid anhydrides; Acting on GTP to facilitate cellular and subcellular movement / cytosolic small ribosomal subunit / SARS-CoV-2 modulates host translation machinery / cytoplasmic translation / tRNA binding / molecular adaptor activity / ribosome / structural constituent of ribosome / translation / GTPase activity / synapse / GTP binding / nucleolus / protein kinase binding / endoplasmic reticulum / RNA binding / nucleoplasm / membrane / nucleus / plasma membrane / cytosol / cytoplasm Similarity search - Function | |||||||||
Biological species | Oryctolagus cuniculus (rabbit) / Escherichia coli (E. coli) | |||||||||
Method | single particle reconstruction / cryo EM / Resolution: 8.7 Å | |||||||||
Authors | Budkevich TV / Giesebrecht J / Behrmann E / Loerke J / Ramrath D / Mielke T / Ismer J / Hildebrand P / Tung C-S / Nierhaus KH ...Budkevich TV / Giesebrecht J / Behrmann E / Loerke J / Ramrath D / Mielke T / Ismer J / Hildebrand P / Tung C-S / Nierhaus KH / Sanbonmatsu KY / Spahn CMT | |||||||||
Citation | Journal: Cell / Year: 2014 Title: Regulation of the mammalian elongation cycle by subunit rolling: a eukaryotic-specific ribosome rearrangement. Authors: Tatyana V Budkevich / Jan Giesebrecht / Elmar Behrmann / Justus Loerke / David J F Ramrath / Thorsten Mielke / Jochen Ismer / Peter W Hildebrand / Chang-Shung Tung / Knud H Nierhaus / ...Authors: Tatyana V Budkevich / Jan Giesebrecht / Elmar Behrmann / Justus Loerke / David J F Ramrath / Thorsten Mielke / Jochen Ismer / Peter W Hildebrand / Chang-Shung Tung / Knud H Nierhaus / Karissa Y Sanbonmatsu / Christian M T Spahn / Abstract: The extent to which bacterial ribosomes and the significantly larger eukaryotic ribosomes share the same mechanisms of ribosomal elongation is unknown. Here, we present subnanometer resolution ...The extent to which bacterial ribosomes and the significantly larger eukaryotic ribosomes share the same mechanisms of ribosomal elongation is unknown. Here, we present subnanometer resolution cryoelectron microscopy maps of the mammalian 80S ribosome in the posttranslocational state and in complex with the eukaryotic eEF1A⋅Val-tRNA⋅GMPPNP ternary complex, revealing significant differences in the elongation mechanism between bacteria and mammals. Surprisingly, and in contrast to bacterial ribosomes, a rotation of the small subunit around its long axis and orthogonal to the well-known intersubunit rotation distinguishes the posttranslocational state from the classical pretranslocational state ribosome. We term this motion "subunit rolling." Correspondingly, a mammalian decoding complex visualized in substates before and after codon recognition reveals structural distinctions from the bacterial system. These findings suggest how codon recognition leads to GTPase activation in the mammalian system and demonstrate that in mammalia subunit rolling occurs during tRNA selection. | |||||||||
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_2623.map.gz | 150.9 MB | EMDB map data format | |
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Header (meta data) | emd-2623-v30.xml emd-2623.xml | 15.7 KB 15.7 KB | Display Display | EMDB header |
Images | EMD-2623_500x500.tif | 732.6 KB | ||
Archive directory | http://ftp.pdbj.org/pub/emdb/structures/EMD-2623 ftp://ftp.pdbj.org/pub/emdb/structures/EMD-2623 | HTTPS FTP |
-Validation report
Summary document | emd_2623_validation.pdf.gz | 243.8 KB | Display | EMDB validaton report |
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Full document | emd_2623_full_validation.pdf.gz | 243 KB | Display | |
Data in XML | emd_2623_validation.xml.gz | 7.2 KB | Display | |
Arichive directory | https://ftp.pdbj.org/pub/emdb/validation_reports/EMD-2623 ftp://ftp.pdbj.org/pub/emdb/validation_reports/EMD-2623 | HTTPS FTP |
-Related structure data
Related structure data | 4cxgMC 2620C 2621C 2622C 2624C 4cxhC 4ujeC M: atomic model generated by this map 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_2623.map.gz / Format: CCP4 / Size: 173.8 MB / Type: IMAGE STORED AS FLOATING POINT NUMBER (4 BYTES) | ||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||
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Annotation | reconstruction of initial codon-sampling state of 80S | ||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||
Projections & slices | Image control
Images are generated by Spider. | ||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||
Voxel size | X=Y=Z: 1.26 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 : Initial codon sampling state of 80S mammalian ribosome
Entire | Name: Initial codon sampling state of 80S mammalian ribosome |
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Components |
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-Supramolecule #1000: Initial codon sampling state of 80S mammalian ribosome
Supramolecule | Name: Initial codon sampling state of 80S mammalian ribosome type: sample / ID: 1000 Details: 80S were re-associated from 40S and 60S subunits and the sample was assembled in vitro from individual components Oligomeric state: one 80S binds three tRNAs, one elongation factor eEF1A and one mRNA Number unique components: 6 |
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Molecular weight | Theoretical: 4.5 MDa |
-Supramolecule #1: Re-associated 80S
Supramolecule | Name: Re-associated 80S / type: complex / ID: 1 / Name.synonym: 80S ribosome / Recombinant expression: No / Ribosome-details: ribosome-eukaryote: ALL |
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Source (natural) | Organism: Oryctolagus cuniculus (rabbit) / synonym: Rabbit / Tissue: Liver |
Molecular weight | Theoretical: 4.5 MDa |
-Macromolecule #1: transfer RNA
Macromolecule | Name: transfer RNA / type: rna / ID: 1 / Name.synonym: tRNA / Details: tRNAPhe / Classification: TRANSFER / Structure: DOUBLE HELIX / Synthetic?: No |
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Source (natural) | Organism: Oryctolagus cuniculus (rabbit) / synonym: Rabbit |
Molecular weight | Theoretical: 25 KDa |
-Macromolecule #2: transfer RNA
Macromolecule | Name: transfer RNA / type: rna / ID: 2 / Name.synonym: tRNA / Classification: TRANSFER / Structure: DOUBLE HELIX / Synthetic?: No |
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Source (natural) | Organism: Oryctolagus cuniculus (rabbit) / synonym: Rabbit |
Molecular weight | Theoretical: 25 KDa |
-Macromolecule #3: transfer RNA
Macromolecule | Name: transfer RNA / type: rna / ID: 3 / Name.synonym: tRNA / Details: Val-tRNAVal / Classification: TRANSFER / Structure: DOUBLE HELIX / Synthetic?: No |
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Source (natural) | Organism: Oryctolagus cuniculus (rabbit) / synonym: Rabbit |
Molecular weight | Theoretical: 25 KDa |
-Macromolecule #5: messenger RNA
Macromolecule | Name: messenger RNA / type: rna / ID: 5 / Name.synonym: mRNA / Classification: OTHER / Structure: SINGLE STRANDED / Synthetic?: Yes |
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Source (natural) | Organism: Escherichia coli (E. coli) / Strain: XL-1 blue |
Molecular weight | Theoretical: 15 KDa |
Sequence | String: GGGAAAAGAA AAGAAAAGAA AAUGUUCGUU AAAGAAAAGA AAAGAAAU |
-Macromolecule #4: elongation factor 1A
Macromolecule | Name: elongation factor 1A / type: protein_or_peptide / ID: 4 / Name.synonym: eEF1A / Number of copies: 1 / Oligomeric state: monomer / Recombinant expression: No |
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Source (natural) | Organism: Oryctolagus cuniculus (rabbit) / synonym: Rabbit |
Molecular weight | Theoretical: 50 KDa |
Sequence | UniProtKB: Elongation factor 1-alpha 1 |
-Experimental details
-Structure determination
Method | cryo EM |
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Processing | single particle reconstruction |
Aggregation state | particle |
-Sample preparation
Concentration | 1.3 mg/mL |
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Buffer | pH: 7.5 Details: 20 mM Hepes, 5 mM MgCl2, 100 mM NH4Cl, 6 mM beta-mercaptoethanol, 0.8 mM spermidine, 0.6 mM spermine |
Grid | Details: Quantifoil grids with additional continuous carbon support |
Vitrification | Cryogen name: ETHANE / Chamber humidity: 80 % / Instrument: FEI VITROBOT MARK II / Method: blot 2/4 sec before plunging |
-Electron microscopy
Microscope | FEI POLARA 300 |
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Temperature | Min: 77 K |
Alignment procedure | Legacy - Astigmatism: objective lens astigmatism was corrected at 200,000 times magnification |
Details | minimal dose system |
Date | Sep 22, 2008 |
Image recording | Category: FILM / Film or detector model: KODAK SO-163 FILM / Digitization - Scanner: OTHER / Number real images: 230 / Average electron dose: 20 e/Å2 / Bits/pixel: 16 |
Electron beam | Acceleration voltage: 300 kV / Electron source: FIELD EMISSION GUN |
Electron optics | Calibrated magnification: 65520 / Illumination mode: FLOOD BEAM / Imaging mode: BRIGHT FIELD / Cs: 2 mm / Nominal defocus max: 4.0 µm / Nominal defocus min: 2.0 µm / Nominal magnification: 39000 |
Sample stage | Specimen holder: nitrogen cooled / Specimen holder model: GATAN LIQUID NITROGEN |
Experimental equipment | Model: Tecnai Polara / Image courtesy: FEI Company |
-Image processing
Details | The particles were selected using SIGNATURE and processed by using SPIDER and SPARX. |
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CTF correction | Details: defocus group |
Final reconstruction | Applied symmetry - Point group: C1 (asymmetric) / Algorithm: OTHER / Resolution.type: BY AUTHOR / Resolution: 8.7 Å / Resolution method: OTHER / Software - Name: Spider, Sparx / Number images used: 79705 |
-Atomic model buiding 1
Initial model | PDB ID: Chain - Chain ID: A |
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Software | Name: Chimera |
Refinement | Space: REAL / Protocol: RIGID BODY FIT / Target criteria: correlation |
Output model | PDB-4cxg: |
-Atomic model buiding 2
Initial model | PDB ID: 2xqd Chain - Chain ID: Y |
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Software | Name: Chimera, Coot |
Details | Acceptor stem loop (2XQD) and tRNA body (1TTT)(cutting points 26-27, 43-44) were docked separately as rigid bodies. Cutting points were restored using Coot. |
Refinement | Space: REAL / Protocol: RIGID BODY FIT / Target criteria: correlation |
Output model | PDB-4cxg: |
-Atomic model buiding 3
Initial model | PDB ID: Chain - Chain ID: D |
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Software | Name: Chimera, Coot |
Details | Acceptor stem loop (2XQD) and tRNA body (1TTT)(cutting points 26-27, 43-44) were docked separately as rigid bodies. Cutting points were restored using Coot. |
Refinement | Space: REAL / Protocol: RIGID BODY FIT / Target criteria: correlation |
Output model | PDB-4cxg: |