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- EMDB-1811: Yeast 80S ribosome stalled by a stem-loop containing mRNA in comp... -
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Open data
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Basic information
Entry | Database: EMDB / ID: EMD-1811 | |||||||||
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Title | Yeast 80S ribosome stalled by a stem-loop containing mRNA in complex with Dom34-Hbs1. The dataset is computationally sorted for presence of P-site tRNA and Dom34-Hbs1. | |||||||||
![]() | This map represents a cryo-EM reconstruction of yeast 80S ribosome stalled by a stable stem-loop structure in complex with Dom34 and Hbs1. Additionally, it contains a P-site tRNA. | |||||||||
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![]() | Ribosome / stalling / mRNA / P-site tRNA / no-go mRNA decay | |||||||||
Function / homology | ![]() Eukaryotic Translation Elongation / Dom34-Hbs1 complex / RNA surveillance / nuclear-transcribed mRNA catabolic process, no-go decay / nuclear-transcribed mRNA catabolic process, non-stop decay / HSF1 activation / Protein methylation / ribosome disassembly / nonfunctional rRNA decay / positive regulation of translational initiation ...Eukaryotic Translation Elongation / Dom34-Hbs1 complex / RNA surveillance / nuclear-transcribed mRNA catabolic process, no-go decay / nuclear-transcribed mRNA catabolic process, non-stop decay / HSF1 activation / Protein methylation / ribosome disassembly / nonfunctional rRNA decay / positive regulation of translational initiation / meiotic cell cycle / translation elongation factor activity / Neutrophil degranulation / RNA endonuclease activity / rescue of stalled ribosome / positive regulation of translation / Hydrolases; Acting on acid anhydrides; Acting on GTP to facilitate cellular and subcellular movement / translation / cell division / GTPase activity / GTP binding / metal ion binding / cytoplasm / cytosol Similarity search - Function | |||||||||
Biological species | ![]() ![]() | |||||||||
Method | single particle reconstruction / cryo EM / Resolution: 9.5 Å | |||||||||
![]() | Becker T / Armache JP / Anger AM / Jarasch A / Villa E / Sieber H / AbdelMotaal B / Berninghausen O / Mielke T / Beckmann R | |||||||||
![]() | ![]() Title: Structure of the no-go mRNA decay complex Dom34-Hbs1 bound to a stalled 80S ribosome. Authors: Thomas Becker / Jean-Paul Armache / Alexander Jarasch / Andreas M Anger / Elizabeth Villa / Heidemarie Sieber / Basma Abdel Motaal / Thorsten Mielke / Otto Berninghausen / Roland Beckmann / ![]() Abstract: No-go decay (NGD) is a mRNA quality-control mechanism in eukaryotic cells that leads to degradation of mRNAs stalled during translational elongation. The key factors triggering NGD are Dom34 and Hbs1. ...No-go decay (NGD) is a mRNA quality-control mechanism in eukaryotic cells that leads to degradation of mRNAs stalled during translational elongation. The key factors triggering NGD are Dom34 and Hbs1. We used cryo-EM to visualize NGD intermediates resulting from binding of the Dom34-Hbs1 complex to stalled ribosomes. At subnanometer resolution, all domains of Dom34 and Hbs1 were identified, allowing the docking of crystal structures and homology models. Moreover, the close structural similarity of Dom34 and Hbs1 to eukaryotic release factors (eRFs) enabled us to propose a model for the ribosome-bound eRF1-eRF3 complex. Collectively, our data provide structural insights into how stalled mRNA is recognized on the ribosome and how the eRF complex can simultaneously recognize stop codons and catalyze peptide release. | |||||||||
History |
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Structure visualization
Movie |
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Structure viewer | EM map: ![]() ![]() ![]() |
Supplemental images |
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Downloads & links
-EMDB archive
Map data | ![]() | 28.9 MB | ![]() | |
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Header (meta data) | ![]() ![]() | 13.8 KB 13.8 KB | Display Display | ![]() |
Images | ![]() | 122.6 KB | ||
Archive directory | ![]() ![]() | HTTPS FTP |
-Related structure data
Related structure data | ![]() 3izqMC ![]() 1808C ![]() 1809C ![]() 1812C M: atomic model generated by this map C: citing same article ( |
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Similar structure data |
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Links
EMDB pages | ![]() ![]() |
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Related items in Molecule of the Month |
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Map
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Annotation | This map represents a cryo-EM reconstruction of yeast 80S ribosome stalled by a stable stem-loop structure in complex with Dom34 and Hbs1. Additionally, it contains a P-site tRNA. | ||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||
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
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Sample components
-Entire : Stem-loop stalled yeast 80S ribosome in complex with Dom34-Hbs1 a...
Entire | Name: Stem-loop stalled yeast 80S ribosome in complex with Dom34-Hbs1 and P-site tRNA. |
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Components |
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-Supramolecule #1000: Stem-loop stalled yeast 80S ribosome in complex with Dom34-Hbs1 a...
Supramolecule | Name: Stem-loop stalled yeast 80S ribosome in complex with Dom34-Hbs1 and P-site tRNA. type: sample / ID: 1000 Details: Mammalian Sec61 was added to saturate the hydrophobic signal sequence present in the nascent polypeptide chain. Oligomeric state: One ribosome / Number unique components: 3 |
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Molecular weight | Theoretical: 3.3 MDa |
-Supramolecule #1: Saccharomyces cerevisiae 80S ribosome
Supramolecule | Name: Saccharomyces cerevisiae 80S ribosome / type: complex / ID: 1 / Name.synonym: yeast 80S ribosome Details: The mRNA stem-loop structure is not visible in the Cryo-EM reconstruction indicating its flexibility Ribosome-details: ribosome-eukaryote: ALL |
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Molecular weight | Experimental: 3.2 MDa / Theoretical: 3.2 MDa |
-Macromolecule #1: Hbs1p
Macromolecule | Name: Hbs1p / type: protein_or_peptide / ID: 1 / Name.synonym: Hbs1p / Number of copies: 1 / Oligomeric state: Monomer / Recombinant expression: Yes |
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Source (natural) | Organism: ![]() ![]() |
Molecular weight | Experimental: 68 KDa / Theoretical: 68 KDa |
Recombinant expression | Organism: ![]() ![]() |
-Macromolecule #2: Dom34p
Macromolecule | Name: Dom34p / type: protein_or_peptide / ID: 2 / Name.synonym: Dom34p / Number of copies: 1 / Oligomeric state: Monomer / Recombinant expression: Yes |
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Source (natural) | Organism: ![]() ![]() |
Molecular weight | Experimental: 44 KDa / Theoretical: 44 KDa |
Recombinant expression | Organism: ![]() ![]() |
-Experimental details
-Structure determination
Method | cryo EM |
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![]() | single particle reconstruction |
Aggregation state | particle |
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Sample preparation
Concentration | 0.02 mg/mL |
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Buffer | pH: 7 Details: 20 mM Tris/HCl, pH 7.0, 80 mM NaCl, 97 mM KOAc, 10 mM Mg(OAc)2, 1.5 mM DTT, 0.02 % Nikkol, 1.8 % Glycerol, 0.01 mg/ml Cycloheximide, 500 0.5 mM GDPNP, 0.3 % Digitonin |
Grid | Details: Quantifoil Grid with 2 nm carbon on top |
Vitrification | Cryogen name: ETHANE / Chamber humidity: 100 % / Instrument: OTHER / Details: Vitrification instrument: Vitrobot Method: Blotted for 10 seconds before plunging, used 2 layer of filter paper |
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Electron microscopy
Microscope | FEI POLARA 300 |
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Temperature | Average: 84 K |
Alignment procedure | Legacy - Astigmatism: Objective lens astigmatism was corrected at 100000 times magnification |
Image recording | Category: CCD / Film or detector model: KODAK SO-163 FILM / Digitization - Sampling interval: 4.76 µm / Number real images: 78 / Average electron dose: 25 e/Å2 Details: Scanned with a Heidelberg PrimeScan drum scanner at 5334 dpi Od range: 1.2 / Bits/pixel: 16 |
Electron beam | Acceleration voltage: 300 kV / Electron source: ![]() |
Electron optics | Calibrated magnification: 38000 / Illumination mode: FLOOD BEAM / Imaging mode: BRIGHT FIELD / Cs: 2.26 mm / Nominal defocus max: 3.78 µm / Nominal defocus min: 1.5 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 |
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Image processing
Details | Mammalian Sec61 complex was added to the sample to saturate the hydrophobic nascent chain |
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CTF correction | Details: CTF correction on the level of 3D volumes (SPIDER TF CTS command) |
Final reconstruction | Applied symmetry - Point group: C1 (asymmetric) / Algorithm: OTHER / Resolution.type: BY AUTHOR / Resolution: 9.5 Å / Resolution method: FSC 0.5 CUT-OFF / Software - Name: SPIDER Details: The dataset was sorted according to presence of Dom34-Hbs1 complex and P-site tRNA. Number images used: 38400 |
-Atomic model buiding 1
Initial model | PDB ID: |
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Software | Name: Molecular Dynamics based flexible fitting MDFF |
Details | Rigid body fitting of individual domains using Coot followed by MDFF |
Refinement | Space: REAL / Protocol: FLEXIBLE FIT |
Output model | ![]() PDB-3izq: |
-Atomic model buiding 2
Initial model | PDB ID: |
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Software | Name: Molecular Dynamics based flexible fitting MDFF |
Details | Rigid body fitting of individual domains using Coot followed by MDFF |
Refinement | Space: REAL / Protocol: FLEXIBLE FIT |
Output model | ![]() PDB-3izq: |