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- EMDB-19352: Cryo-EM structure of the 60S subunit with the RQC complex contain... -

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Basic information

Entry
Database: EMDB / ID: EMD-19352
TitleCryo-EM structure of the 60S subunit with the RQC complex containing the mutant F340I Rqc2 protein and the Flag tagged Ltn1-Delta_ring
Map dataSaccharomyces cerevisiae 60S subunit with the RQC complex containing the mutant F340I Rqc2 protein and the Flag tagged Ltn1-Delta-ring
Sample
  • Complex: Cryo-EM structure of the 60S subunit with the RQC complex containing the mutant F340I Rqc2 protein and the Flag tagged Ltn1-Delta_ring
KeywordsRibosome quality complex / RQC2 / peptidyl-tRNA hydrolase / No-go decay / Non-stop decay / RIBOSOME
Biological speciesSaccharomyces cerevisiae (brewer's yeast)
Methodsingle particle reconstruction / cryo EM / Resolution: 2.9 Å
AuthorsFabret C / Giudice E / Chat S / Gillet R / Namy O
Funding support France, 2 items
OrganizationGrant numberCountry
Agence Nationale de la Recherche (ANR)ANR-17-CE12-0024-01 France
Fondation pour la Recherche Medicale (FRM)EQU202203014663 France
CitationJournal: Structure / Year: 2025
Title: RQC2 is a major player in peptide release from stalled ribosomes.
Authors: Céline Fabret / Emmanuel Giudice / Sophie Chat / Reynald Gillet / Olivier Namy /
Abstract: Eukaryotic cells prevent the accumulation of potentially toxic aberrant polypeptides and maintain ribosome availability through surveillance and clearance mechanisms, including the evolutionarily ...Eukaryotic cells prevent the accumulation of potentially toxic aberrant polypeptides and maintain ribosome availability through surveillance and clearance mechanisms, including the evolutionarily conserved ribosome-associated quality control complex (RQC). RQC pathways have been widely investigated, with the identification of several factors ANKZF1/Vms1p, Ptrh1, and Arb1p involved in release/cleavage of the peptide-tRNA from 60S subunits. We aimed here to identify the genes involved in peptide release from stalled ribosomes. Using a genetic screen, we identified a mutant allele of RQC2 as involved in this process. We present the cryoelectron microscopy (cryo-EM) structure of RQC, which reveals how the F340I mutation affects mutant binding. This altered binding, in turn, disrupts the A-site's ability to bind the tRNA in the presence of Ltn1. These data account for the limitation of C-terminal alanine and threonine (CAT) tailing by the F340I mutation and suggest a model explaining the role of the Rqc2 protein in peptide release.
History
DepositionJan 5, 2024-
Header (metadata) releaseApr 16, 2025-
Map releaseApr 16, 2025-
UpdateJun 18, 2025-
Current statusJun 18, 2025Processing site: PDBe / Status: Released

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Structure visualization

Supplemental images

emd_19352.png

Downloads & links

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Map

FileDownload / File: emd_19352.map.gz / Format: CCP4 / Size: 669.9 MB / Type: IMAGE STORED AS FLOATING POINT NUMBER (4 BYTES)
AnnotationSaccharomyces cerevisiae 60S subunit with the RQC complex containing the mutant F340I Rqc2 protein and the Flag tagged Ltn1-Delta-ring
Projections & slices

Image control

Size
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Others
AxesZ (Sec.)Y (Row.)X (Col.)
0.73 Å/pix.
x 560 pix.
= 409.36 Å		0.73 Å/pix.
x 560 pix.
= 409.36 Å		0.73 Å/pix.
x 560 pix.
= 409.36 Å

Surface

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Images are generated by Spider.

Voxel sizeX=Y=Z: 0.731 Å
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Histogram

Histogram (log scale)
Contour LevelBy AUTHOR: 0.003
Minimum - Maximum-0.009857988 - 0.030789714
Average (Standard dev.)0.0001234862 (±0.0014152003)
SymmetrySpace group: 1
Details

EMDB XML:

Map geometry
Axis orderXYZ
Origin000
Dimensions560560560
Spacing560560560
CellA=B=C: 409.36002 Å
α=β=γ: 90.0 °

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Supplemental data

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Mask #1

Fileemd_19352_msk_1.map
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Additional map: Saccharomyces cerevisiae 60S subunit with the RQC complex...

Fileemd_19352_additional_1.map
AnnotationSaccharomyces cerevisiae 60S subunit with the RQC complex - Postprocessed & Masked map
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Additional map: Saccharomyces cerevisiae 60S subunit with the RQC complex...

Fileemd_19352_additional_2.map
AnnotationSaccharomyces cerevisiae 60S subunit with the RQC complex - map filtered according to local resolution
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Half map: Saccharomyces cerevisiae 60S subunit with the RQC complex...

Fileemd_19352_half_map_1.map
AnnotationSaccharomyces cerevisiae 60S subunit with the RQC complex - Half Map 1
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Half map: Saccharomyces cerevisiae 60S subunit with the RQC complex...

Fileemd_19352_half_map_2.map
AnnotationSaccharomyces cerevisiae 60S subunit with the RQC complex - Half Map 2
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Sample components

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Entire : Cryo-EM structure of the 60S subunit with the RQC complex contain...

EntireName: Cryo-EM structure of the 60S subunit with the RQC complex containing the mutant F340I Rqc2 protein and the Flag tagged Ltn1-Delta_ring
Components
  • Complex: Cryo-EM structure of the 60S subunit with the RQC complex containing the mutant F340I Rqc2 protein and the Flag tagged Ltn1-Delta_ring

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Supramolecule #1: Cryo-EM structure of the 60S subunit with the RQC complex contain...

SupramoleculeName: Cryo-EM structure of the 60S subunit with the RQC complex containing the mutant F340I Rqc2 protein and the Flag tagged Ltn1-Delta_ring
type: complex / ID: 1 / Parent: 0 / Macromolecule list: #1
Source (natural)Organism: Saccharomyces cerevisiae (brewer's yeast)

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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: 7.4
Component:
ConcentrationName
20.0 mMHepes-KOH
100.0 mMKoAc
10.0 mMMgoAc
0.5 mMDTT
2.0 mg/mL3x Flag peptide
0.02 %Glutaraldehyde

Details: + complete protease inhibitor cocktail
GridModel: Quantifoil R2/1 / Material: COPPER / Support film - Material: CARBON / Support film - topology: CONTINUOUS / Support film - Film thickness: 0.3 / Pretreatment - Type: GLOW DISCHARGE
VitrificationCryogen name: ETHANE / Chamber humidity: 90 % / Chamber temperature: 293 K / Instrument: FEI VITROBOT MARK III
DetailsImmunoaffinity purify Saccharomyces cerevisiae 60S ribosomal subunit with the RQC complex containing the mutant Rqc2-F340I protein and the Flag tagged Ltn1-Delta_ring

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Electron microscopy

MicroscopeFEI TITAN KRIOS
Specialist opticsSpherical aberration corrector: Cs-corrected / Energy filter - Name: TFS Selectris X
Image recordingFilm or detector model: FEI FALCON IV (4k x 4k) / Number real images: 49229 / Average exposure time: 6.0 sec. / Average electron dose: 33.0 e/Å2
Electron beamAcceleration voltage: 300 kV / Electron source: FIELD EMISSION GUN
Electron opticsIllumination mode: FLOOD BEAM / Imaging mode: BRIGHT FIELD / Nominal defocus max: 1.8 µm / Nominal defocus min: 0.9 µm / Nominal magnification: 165000
Experimental equipment
Model: Titan Krios / Image courtesy: FEI Company

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Image processing

Particle selectionNumber selected: 946082
CTF correctionSoftware - Name: CTFFIND (ver. 4.1.13) / Type: PHASE FLIPPING AND AMPLITUDE CORRECTION
Startup modelType of model: NONE
Final reconstructionNumber classes used: 1 / Algorithm: BACK PROJECTION / Resolution.type: BY AUTHOR / Resolution: 2.9 Å / Resolution method: FSC 0.143 CUT-OFF / Software - Name: RELION (ver. 4.0) / Number images used: 63756
Initial angle assignmentType: MAXIMUM LIKELIHOOD / Software - Name: cryoSPARC (ver. 4.2)
Final angle assignmentType: MAXIMUM LIKELIHOOD / Software - Name: RELION (ver. 4.0)
Final 3D classificationSoftware - Name: RELION (ver. 4.0)
FSC plot (resolution estimation)

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