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- EMDB-3734: Class 1 of rotated ribosomes (rotated 1 state) with A/A- and P/E-... -

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Basic information

Entry
Database: EMDB / ID: EMD-3734
TitleClass 1 of rotated ribosomes (rotated 1 state) with A/A- and P/E-site tRNAs of the Rqt1-FTP pulldown
Map dataClass 1 of rotated ribosomes (rotated 1 state) with A/A- and P/E-site tRNAs of the Rqt1-FTP pulldown
Sample
  • Complex: Class 1 of rotated ribosomes (rotated 1 state) with A/A- and P/E-site tRNAs of the Rqt1-FTP pulldown
Biological speciesSaccharomyces cerevisiae (brewer's yeast)
Methodsingle particle reconstruction / cryo EM / Resolution: 7.0 Å
AuthorsMatsuo Y / Ikeuchi K / Saeki Y / Iwasaki S / Schmidt C / Udagawa T / Sato F / Tsuchiya H / Becker T / Tanaka K ...Matsuo Y / Ikeuchi K / Saeki Y / Iwasaki S / Schmidt C / Udagawa T / Sato F / Tsuchiya H / Becker T / Tanaka K / Ingolia N / Beckmann R / Inada T
CitationJournal: Nat Commun / Year: 2017
Title: Ubiquitination of stalled ribosome triggers ribosome-associated quality control.
Authors: Yoshitaka Matsuo / Ken Ikeuchi / Yasushi Saeki / Shintaro Iwasaki / Christian Schmidt / Tsuyoshi Udagawa / Fumiya Sato / Hikaru Tsuchiya / Thomas Becker / Keiji Tanaka / Nicholas T Ingolia / ...Authors: Yoshitaka Matsuo / Ken Ikeuchi / Yasushi Saeki / Shintaro Iwasaki / Christian Schmidt / Tsuyoshi Udagawa / Fumiya Sato / Hikaru Tsuchiya / Thomas Becker / Keiji Tanaka / Nicholas T Ingolia / Roland Beckmann / Toshifumi Inada /
Abstract: Translation arrest by polybasic sequences induces ribosome stalling, and the arrest product is degraded by the ribosome-mediated quality control (RQC) system. Here we report that ubiquitination of ...Translation arrest by polybasic sequences induces ribosome stalling, and the arrest product is degraded by the ribosome-mediated quality control (RQC) system. Here we report that ubiquitination of the 40S ribosomal protein uS10 by the E3 ubiquitin ligase Hel2 (or RQT1) is required for RQC. We identify a RQC-trigger (RQT) subcomplex composed of the RNA helicase-family protein Slh1/Rqt2, the ubiquitin-binding protein Cue3/Rqt3, and yKR023W/Rqt4 that is required for RQC. The defects in RQC of the RQT mutants correlate with sensitivity to anisomycin, which stalls ribosome at the rotated form. Cryo-electron microscopy analysis reveals that Hel2-bound ribosome are dominantly the rotated form with hybrid tRNAs. Ribosome profiling reveals that ribosomes stalled at the rotated state with specific pairs of codons at P-A sites serve as RQC substrates. Rqt1 specifically ubiquitinates these arrested ribosomes to target them to the RQT complex, allowing subsequent RQC reactions including dissociation of the stalled ribosome into subunits.Several protein quality control mechanisms are in place to trigger the rapid degradation of aberrant polypeptides and mRNAs. Here the authors describe a mechanism of ribosome-mediated quality control that involves the ubiquitination of ribosomal proteins by the E3 ubiquitin ligase Hel2/RQT1.
History
DepositionMay 23, 2017-
Header (metadata) releaseMay 31, 2017-
Map releaseAug 9, 2017-
UpdateAug 9, 2017-
Current statusAug 9, 2017Processing site: PDBe / Status: Released

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Structure visualization

Movie
  • Surface view with section colored by density value
  • Surface level: 0.027
  • Imaged by UCSF Chimera
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  • Surface view colored by height
  • Surface level: 0.027
  • Imaged by UCSF Chimera
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Movie viewer
Structure viewerEM map:
SurfViewMolmilJmol/JSmol
Supplemental images

emd_3734.png

Downloads & links

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Map

FileDownload / File: emd_3734.map.gz / Format: CCP4 / Size: 8.4 MB / Type: IMAGE STORED AS FLOATING POINT NUMBER (4 BYTES)
AnnotationClass 1 of rotated ribosomes (rotated 1 state) with A/A- and P/E-site tRNAs of the Rqt1-FTP pulldown
Voxel sizeX=Y=Z: 3.252 Å
Density
Contour LevelBy AUTHOR: 0.027 / Movie #1: 0.027
Minimum - Maximum-0.079807095 - 0.23610805
Average (Standard dev.)-0.00047539812 (±0.023795936)
SymmetrySpace group: 1
Details

EMDB XML:

Map geometry
Axis orderXYZ
Origin000
Dimensions130130130
Spacing130130130
CellA=B=C: 422.76 Å
α=β=γ: 90.0 °

CCP4 map header:

modeImage stored as Reals
Å/pix. X/Y/Z3.2523.2523.252
M x/y/z130130130
origin x/y/z0.0000.0000.000
length x/y/z422.760422.760422.760
α/β/γ90.00090.00090.000
MAP C/R/S123
start NC/NR/NS000
NC/NR/NS130130130
D min/max/mean-0.0800.236-0.000

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Supplemental data

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Sample components

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Entire : Class 1 of rotated ribosomes (rotated 1 state) with A/A- and P/E-...

EntireName: Class 1 of rotated ribosomes (rotated 1 state) with A/A- and P/E-site tRNAs of the Rqt1-FTP pulldown
Components
  • Complex: Class 1 of rotated ribosomes (rotated 1 state) with A/A- and P/E-site tRNAs of the Rqt1-FTP pulldown

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Supramolecule #1: Class 1 of rotated ribosomes (rotated 1 state) with A/A- and P/E-...

SupramoleculeName: Class 1 of rotated ribosomes (rotated 1 state) with A/A- and P/E-site tRNAs of the Rqt1-FTP pulldown
type: complex / ID: 1 / Parent: 0
Details: Second class from classification for dataset in Frealign
Source (natural)Organism: Saccharomyces cerevisiae (brewer's yeast)
Molecular weightTheoretical: 3.6 MDa

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Experimental details

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Structure determination

Methodcryo EM
Processingsingle particle reconstruction
Aggregation stateparticle

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Sample preparation

BufferpH: 7.5
VitrificationCryogen name: ETHANE

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Electron microscopy

MicroscopeFEI TITAN KRIOS
Electron beamAcceleration voltage: 300 kV / Electron source: FIELD EMISSION GUN
Electron opticsIllumination mode: FLOOD BEAM / Imaging mode: BRIGHT FIELDBright-field microscopy / Cs: 2.7 mm
Image recordingFilm or detector model: FEI FALCON II (4k x 4k) / Average electron dose: 28.0 e/Å2
Experimental equipment
Model: Titan Krios / Image courtesy: FEI Company

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Image processing

Initial angle assignmentType: PROJECTION MATCHING
Final angle assignmentType: PROJECTION MATCHING
Final reconstructionResolution.type: BY AUTHOR / Resolution: 7.0 Å / Resolution method: FSC 0.143 CUT-OFF / Number images used: 39892

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