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Yorodumi- PDB-6xir: Cryo-EM Structure of K63 Ubiquitinated Yeast Translocating Riboso... -
+Open data
-Basic information
Entry | Database: PDB / ID: 6xir | ||||||
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Title | Cryo-EM Structure of K63 Ubiquitinated Yeast Translocating Ribosome under Oxidative Stress | ||||||
Components |
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Keywords | RIBOSOME / K63 ubiquitin / oxidative stress / translation | ||||||
Function / homology | Function and homology information negative regulation of glucose mediated signaling pathway / negative regulation of translational frameshifting / mTORC1-mediated signalling / Protein hydroxylation / ribosome-associated ubiquitin-dependent protein catabolic process / GDP-dissociation inhibitor activity / pre-mRNA 5'-splice site binding / Formation of the ternary complex, and subsequently, the 43S complex / Translation initiation complex formation / Ribosomal scanning and start codon recognition ...negative regulation of glucose mediated signaling pathway / negative regulation of translational frameshifting / mTORC1-mediated signalling / Protein hydroxylation / ribosome-associated ubiquitin-dependent protein catabolic process / GDP-dissociation inhibitor activity / pre-mRNA 5'-splice site binding / Formation of the ternary complex, and subsequently, the 43S complex / Translation initiation complex formation / Ribosomal scanning and start codon recognition / cleavage in ITS2 between 5.8S rRNA and LSU-rRNA of tricistronic rRNA transcript (SSU-rRNA, 5.8S rRNA, LSU-rRNA) / response to cycloheximide / mRNA destabilization / Major pathway of rRNA processing in the nucleolus and cytosol / SRP-dependent cotranslational protein targeting to membrane / GTP hydrolysis and joining of the 60S ribosomal subunit / Formation of a pool of free 40S subunits / Nonsense Mediated Decay (NMD) independent of the Exon Junction Complex (EJC) / Nonsense Mediated Decay (NMD) enhanced by the Exon Junction Complex (EJC) / negative regulation of mRNA splicing, via spliceosome / L13a-mediated translational silencing of Ceruloplasmin expression / preribosome, large subunit precursor / endonucleolytic cleavage to generate mature 3'-end of SSU-rRNA from (SSU-rRNA, 5.8S rRNA, LSU-rRNA) / G-protein alpha-subunit binding / positive regulation of protein kinase activity / regulation of translational fidelity / protein-RNA complex assembly / translation regulator activity / ribosomal subunit export from nucleus / endonucleolytic cleavage in ITS1 to separate SSU-rRNA from 5.8S rRNA and LSU-rRNA from tricistronic rRNA transcript (SSU-rRNA, 5.8S rRNA, LSU-rRNA) / cellular response to amino acid starvation / rescue of stalled ribosome / maturation of LSU-rRNA from tricistronic rRNA transcript (SSU-rRNA, 5.8S rRNA, LSU-rRNA) / 90S preribosome / maturation of SSU-rRNA from tricistronic rRNA transcript (SSU-rRNA, 5.8S rRNA, LSU-rRNA) / maturation of LSU-rRNA / ribosomal large subunit biogenesis / maturation of SSU-rRNA / macroautophagy / small-subunit processome / positive regulation of apoptotic signaling pathway / protein kinase C binding / maintenance of translational fidelity / ribosomal large subunit assembly / cytoplasmic stress granule / rRNA processing / ribosome biogenesis / ribosome binding / ribosomal small subunit biogenesis / ribosomal small subunit assembly / small ribosomal subunit / small ribosomal subunit rRNA binding / 5S rRNA binding / large ribosomal subunit rRNA binding / cytosolic small ribosomal subunit / cytosolic large ribosomal subunit / cytoplasmic translation / rRNA binding / negative regulation of translation / ribosome / structural constituent of ribosome / translation / positive regulation of protein phosphorylation / G protein-coupled receptor signaling pathway / negative regulation of gene expression / response to antibiotic / mRNA binding / nucleolus / mitochondrion / RNA binding / zinc ion binding / nucleoplasm / nucleus / metal ion binding / cytoplasm / cytosol Similarity search - Function | ||||||
Biological species | Saccharomyces cerevisiae (brewer's yeast) | ||||||
Method | ELECTRON MICROSCOPY / single particle reconstruction / cryo EM / Resolution: 3.2 Å | ||||||
Authors | Zhou, Y. / Bartesaghi, A. / Silva, G.M. | ||||||
Funding support | United States, 1items
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Citation | Journal: Proc Natl Acad Sci U S A / Year: 2020 Title: Structural impact of K63 ubiquitin on yeast translocating ribosomes under oxidative stress. Authors: Ye Zhou / Panagiotis L Kastritis / Shannon E Dougherty / Jonathan Bouvette / Allen L Hsu / Laura Burbaum / Shyamal Mosalaganti / Stefan Pfeffer / Wim J H Hagen / Friedrich Förster / Mario J ...Authors: Ye Zhou / Panagiotis L Kastritis / Shannon E Dougherty / Jonathan Bouvette / Allen L Hsu / Laura Burbaum / Shyamal Mosalaganti / Stefan Pfeffer / Wim J H Hagen / Friedrich Förster / Mario J Borgnia / Christine Vogel / Martin Beck / Alberto Bartesaghi / Gustavo M Silva / Abstract: Subpopulations of ribosomes are responsible for fine tuning the control of protein synthesis in dynamic environments. K63 ubiquitination of ribosomes has emerged as a new posttranslational ...Subpopulations of ribosomes are responsible for fine tuning the control of protein synthesis in dynamic environments. K63 ubiquitination of ribosomes has emerged as a new posttranslational modification that regulates protein synthesis during cellular response to oxidative stress. K63 ubiquitin, a type of ubiquitin chain that functions independently of the proteasome, modifies several sites at the surface of the ribosome, however, we lack a molecular understanding on how this modification affects ribosome structure and function. Using cryoelectron microscopy (cryo-EM), we resolved the first three-dimensional (3D) structures of K63 ubiquitinated ribosomes from oxidatively stressed yeast cells at 3.5-3.2 Å resolution. We found that K63 ubiquitinated ribosomes are also present in a polysome arrangement, similar to that observed in yeast polysomes, which we determined using cryoelectron tomography (cryo-ET). We further showed that K63 ubiquitinated ribosomes are captured uniquely at the rotated pretranslocation stage of translation elongation. In contrast, cryo-EM structures of ribosomes from mutant cells lacking K63 ubiquitin resolved at 4.4-2.7 Å showed 80S ribosomes represented in multiple states of translation, suggesting that K63 ubiquitin regulates protein synthesis at a selective stage of elongation. Among the observed structural changes, ubiquitin mediates the destabilization of proteins in the 60S P-stalk and in the 40S beak, two binding regions of the eukaryotic elongation factor eEF2. These changes would impact eEF2 function, thus, inhibiting translocation. Our findings help uncover the molecular effects of K63 ubiquitination on ribosomes, providing a model of translation control during oxidative stress, which supports elongation halt at pretranslocation. | ||||||
History |
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-Structure visualization
Movie |
Movie viewer |
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Structure viewer | Molecule: MolmilJmol/JSmol |
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PDBx/mmCIF format | 6xir.cif.gz | 4.5 MB | Display | PDBx/mmCIF format |
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PDB format | pdb6xir.ent.gz | Display | PDB format | |
PDBx/mmJSON format | 6xir.json.gz | Tree view | PDBx/mmJSON format | |
Others | Other downloads |
-Validation report
Summary document | 6xir_validation.pdf.gz | 1.5 MB | Display | wwPDB validaton report |
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Full document | 6xir_full_validation.pdf.gz | 1.6 MB | Display | |
Data in XML | 6xir_validation.xml.gz | 319.1 KB | Display | |
Data in CIF | 6xir_validation.cif.gz | 563 KB | Display | |
Arichive directory | https://data.pdbj.org/pub/pdb/validation_reports/xi/6xir ftp://data.pdbj.org/pub/pdb/validation_reports/xi/6xir | HTTPS FTP |
-Related structure data
Related structure data | 22198MC 6xiqC C: citing same article (ref.) M: map data used to model this data |
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Similar structure data |
-Links
-Assembly
Deposited unit |
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1 |
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-Components
+60S ribosomal protein ... , 28 types, 28 molecules AEFLMNOPQRSTUVXYZacdfghijlop
+Protein , 24 types, 24 molecules BCDGHIJWbekrstvyAFAKAMAPAQASATAV
-40S ribosomal protein ... , 17 types, 17 molecules AAABquwxzADAEAGAHAIAJALANAOAR
#26: Protein | Mass: 12757.445 Da / Num. of mol.: 1 / Source method: isolated from a natural source / Source: (natural) Saccharomyces cerevisiae (brewer's yeast) / Strain: SUB413 / References: UniProt: Q08745 |
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#27: Protein | Mass: 17785.934 Da / Num. of mol.: 1 / Source method: isolated from a natural source / Source: (natural) Saccharomyces cerevisiae (brewer's yeast) / Strain: SUB413 / References: UniProt: P0CX47 |
#47: Protein | Mass: 28051.330 Da / Num. of mol.: 1 / Source method: isolated from a natural source / Source: (natural) Saccharomyces cerevisiae (brewer's yeast) / Strain: SUB413 / References: UniProt: P32905 |
#51: Protein | Mass: 29469.330 Da / Num. of mol.: 1 / Source method: isolated from a natural source / Source: (natural) Saccharomyces cerevisiae (brewer's yeast) / Strain: SUB413 / References: UniProt: P0CX35 |
#53: Protein | Mass: 27054.486 Da / Num. of mol.: 1 / Source method: isolated from a natural source / Source: (natural) Saccharomyces cerevisiae (brewer's yeast) / Strain: SUB413 / References: UniProt: P0CX37 |
#54: Protein | Mass: 21658.209 Da / Num. of mol.: 1 / Source method: isolated from a natural source / Source: (natural) Saccharomyces cerevisiae (brewer's yeast) / Strain: SUB413 / References: UniProt: P26786 |
#56: Protein | Mass: 22487.893 Da / Num. of mol.: 1 / Source method: isolated from a natural source / Source: (natural) Saccharomyces cerevisiae (brewer's yeast) / Strain: SUB413 / References: UniProt: O13516 |
#57: Protein | Mass: 17059.945 Da / Num. of mol.: 1 / Source method: isolated from a natural source / Source: (natural) Saccharomyces cerevisiae (brewer's yeast) / Strain: SUB413 / References: UniProt: P05756 |
#58: Protein | Mass: 14675.771 Da / Num. of mol.: 1 / Source method: isolated from a natural source / Source: (natural) Saccharomyces cerevisiae (brewer's yeast) / Strain: SUB413 / References: UniProt: P39516 |
#60: Protein | Mass: 15877.490 Da / Num. of mol.: 1 / Source method: isolated from a natural source / Source: (natural) Saccharomyces cerevisiae (brewer's yeast) / Strain: SUB413 / References: UniProt: P0CX51 |
#61: Protein | Mass: 15835.384 Da / Num. of mol.: 1 / Source method: isolated from a natural source / Source: (natural) Saccharomyces cerevisiae (brewer's yeast) / Strain: SUB413 / References: UniProt: P14127 |
#62: Protein | Mass: 17071.641 Da / Num. of mol.: 1 / Source method: isolated from a natural source / Source: (natural) Saccharomyces cerevisiae (brewer's yeast) / Strain: SUB413 / References: UniProt: P0CX55 |
#63: Protein | Mass: 15942.125 Da / Num. of mol.: 1 / Source method: isolated from a natural source / Source: (natural) Saccharomyces cerevisiae (brewer's yeast) / Strain: SUB413 / References: UniProt: P07280 |
#65: Protein | Mass: 9758.829 Da / Num. of mol.: 1 / Source method: isolated from a natural source / Source: (natural) Saccharomyces cerevisiae (brewer's yeast) / Strain: SUB413 / References: UniProt: P0C0V8 |
#67: Protein | Mass: 16073.896 Da / Num. of mol.: 1 / Source method: isolated from a natural source / Source: (natural) Saccharomyces cerevisiae (brewer's yeast) / Strain: SUB413 / References: UniProt: P0CX29 |
#68: Protein | Mass: 15362.848 Da / Num. of mol.: 1 / Source method: isolated from a natural source / Source: (natural) Saccharomyces cerevisiae (brewer's yeast) / Strain: SUB413 / References: UniProt: P0CX31 |
#71: Protein | Mass: 8893.391 Da / Num. of mol.: 1 / Source method: isolated from a natural source / Source: (natural) Saccharomyces cerevisiae (brewer's yeast) / Strain: SUB413 / References: UniProt: P35997 |
-RNA chain , 5 types, 6 molecules 1342AXAZ
#28: RNA chain | Mass: 1097148.625 Da / Num. of mol.: 1 / Source method: isolated from a natural source / Source: (natural) Saccharomyces cerevisiae (brewer's yeast) / Strain: SUB413 / References: GenBank: 380294104 |
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#29: RNA chain | Mass: 38951.105 Da / Num. of mol.: 1 / Source method: isolated from a natural source / Source: (natural) Saccharomyces cerevisiae (brewer's yeast) / Strain: SUB413 / References: GenBank: 176405 |
#30: RNA chain | Mass: 50682.922 Da / Num. of mol.: 1 / Source method: isolated from a natural source / Source: (natural) Saccharomyces cerevisiae (brewer's yeast) / Strain: SUB413 / References: GenBank: 1267176496 |
#46: RNA chain | Mass: 579761.938 Da / Num. of mol.: 1 / Source method: isolated from a natural source / Source: (natural) Saccharomyces cerevisiae (brewer's yeast) / Strain: SUB413 / References: GenBank: 1262303 |
#75: RNA chain | Mass: 24518.570 Da / Num. of mol.: 2 / Source method: isolated from a natural source / Source: (natural) Saccharomyces cerevisiae (brewer's yeast) / Strain: SUB413 / References: GenBank: 176419 |
-Protein/peptide / Non-polymers , 2 types, 7 molecules n
#43: Protein/peptide | Mass: 3354.243 Da / Num. of mol.: 1 / Source method: isolated from a natural source / Source: (natural) Saccharomyces cerevisiae (brewer's yeast) / Strain: SUB413 / References: UniProt: A0A6A5PXU0 |
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#76: Chemical | ChemComp-ZN / |
-Details
Has ligand of interest | Y |
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-Experimental details
-Experiment
Experiment | Method: ELECTRON MICROSCOPY |
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EM experiment | Aggregation state: PARTICLE / 3D reconstruction method: single particle reconstruction |
-Sample preparation
Component | Name: K63 ubiquitinated Ribosome / Type: RIBOSOME Details: K63 ubiquitin modified yeast translocating ribosome under oxidative stress Entity ID: #1-#75 / Source: NATURAL |
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Molecular weight | Experimental value: NO |
Source (natural) | Organism: Saccharomyces cerevisiae (brewer's yeast) / Strain: SUB413 |
Buffer solution | pH: 7.5 |
Specimen | Embedding applied: NO / Shadowing applied: NO / Staining applied: NO / Vitrification applied: YES |
Specimen support | Grid material: COPPER / Grid type: Quantifoil R2/1 |
Vitrification | Cryogen name: ETHANE |
-Electron microscopy imaging
Experimental equipment | Model: Titan Krios / Image courtesy: FEI Company |
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Microscopy | Model: FEI TITAN KRIOS |
Electron gun | Electron source: FIELD EMISSION GUN / Accelerating voltage: 300 kV / Illumination mode: FLOOD BEAM |
Electron lens | Mode: BRIGHT FIELD |
Specimen holder | Cryogen: NITROGEN / Specimen holder model: FEI TITAN KRIOS AUTOGRID HOLDER |
Image recording | Electron dose: 25 e/Å2 / Detector mode: INTEGRATING / Film or detector model: FEI FALCON II (4k x 4k) / Num. of real images: 5243 |
Image scans | Movie frames/image: 7 / Used frames/image: 1-7 |
-Processing
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EM software |
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CTF correction | Type: PHASE FLIPPING AND AMPLITUDE CORRECTION | ||||||||||||||||||||||||
Symmetry | Point symmetry: C1 (asymmetric) | ||||||||||||||||||||||||
3D reconstruction | Resolution: 3.2 Å / Resolution method: FSC 0.143 CUT-OFF / Num. of particles: 87939 / Algorithm: FOURIER SPACE / Num. of class averages: 1 / Symmetry type: POINT | ||||||||||||||||||||||||
Atomic model building | Protocol: FLEXIBLE FIT / Space: REAL / Target criteria: correlation coefficient | ||||||||||||||||||||||||
Atomic model building | PDB-ID: 6GQ1 | ||||||||||||||||||||||||
Refinement | Cross valid method: NONE Stereochemistry target values: GeoStd + Monomer Library + CDL v1.2 | ||||||||||||||||||||||||
Displacement parameters | Biso mean: 157.13 Å2 | ||||||||||||||||||||||||
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