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- EMDB-14992: FAP-80S Complex - Non-rotated state with empty A site -

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Basic information

Entry
Database: EMDB / ID: EMD-14992
TitleFAP-80S Complex - Non-rotated state with empty A site
Map dataFAP-80S - Non-rotated state - Empty A site
Sample
  • Complex: Fap1-bound yeast 80S ribosome in non-rotated state with empty A site
Biological speciesSaccharomyces cerevisiae (brewer's yeast)
Methodsingle particle reconstruction / cryo EM / Resolution: 3.0 Å
AuthorsIkeuchi K / Buschauer R / Berninghausen O / Becker T / Beckmann R
Funding support Germany, European Union, 3 items
OrganizationGrant numberCountry
German Research Foundation (DFG)BE 1814/15-1 Germany
German Research Foundation (DFG)RTG1721 Germany
European Research Council (ERC)885711European Union
CitationJournal: Mol Cell / Year: 2022
Title: Sensing of individual stalled 80S ribosomes by Fap1 for nonfunctional rRNA turnover.
Authors: Sihan Li / Ken Ikeuchi / Misaki Kato / Robert Buschauer / Takato Sugiyama / Shungo Adachi / Hideo Kusano / Tohru Natsume / Otto Berninghausen / Yoshitaka Matsuo / Thomas Becker / Roland ...Authors: Sihan Li / Ken Ikeuchi / Misaki Kato / Robert Buschauer / Takato Sugiyama / Shungo Adachi / Hideo Kusano / Tohru Natsume / Otto Berninghausen / Yoshitaka Matsuo / Thomas Becker / Roland Beckmann / Toshifumi Inada /
Abstract: Cells can respond to stalled ribosomes by sensing ribosome collisions and employing quality control pathways. How ribosome stalling is resolved without collisions, however, has remained elusive. ...Cells can respond to stalled ribosomes by sensing ribosome collisions and employing quality control pathways. How ribosome stalling is resolved without collisions, however, has remained elusive. Here, focusing on noncolliding stalling exhibited by decoding-defective ribosomes, we identified Fap1 as a stalling sensor triggering 18S nonfunctional rRNA decay via polyubiquitination of uS3. Ribosome profiling revealed an enrichment of Fap1 at the translation initiation site but also an association with elongating individual ribosomes. Cryo-EM structures of Fap1-bound ribosomes elucidated Fap1 probing the mRNA simultaneously at both the entry and exit channels suggesting an mRNA stasis sensing activity, and Fap1 sterically hinders the formation of canonical collided di-ribosomes. Our findings indicate that individual stalled ribosomes are the potential signal for ribosome dysfunction, leading to accelerated turnover of the ribosome itself.
History
DepositionMay 18, 2022-
Header (metadata) releaseOct 5, 2022-
Map releaseOct 5, 2022-
UpdateOct 5, 2022-
Current statusOct 5, 2022Processing site: PDBe / Status: Released

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Structure visualization

Supplemental images

Downloads & links

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Map

FileDownload / File: emd_14992.map.gz / Format: CCP4 / Size: 421.9 MB / Type: IMAGE STORED AS FLOATING POINT NUMBER (4 BYTES)
AnnotationFAP-80S - Non-rotated state - Empty A site
Projections & slices

Image control

Size
Brightness
Contrast
Others
AxesZ (Sec.)Y (Row.)X (Col.)
1.05 Å/pix.
x 480 pix.
= 501.6 Å
1.05 Å/pix.
x 480 pix.
= 501.6 Å
1.05 Å/pix.
x 480 pix.
= 501.6 Å

Surface

Projections

Slices (1/3)

Slices (1/2)

Slices (2/3)

Images are generated by Spider.

Voxel sizeX=Y=Z: 1.045 Å
Density
Contour LevelBy AUTHOR: 0.025
Minimum - Maximum-0.21827902 - 0.5205617
Average (Standard dev.)0.00016413766 (±0.013185275)
SymmetrySpace group: 1
Details

EMDB XML:

Map geometry
Axis orderXYZ
Origin000
Dimensions480480480
Spacing480480480
CellA=B=C: 501.59998 Å
α=β=γ: 90.0 °

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Supplemental data

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Half map: Half map 1

Fileemd_14992_half_map_1.map
AnnotationHalf map 1
Projections & Slices
AxesZYX

Projections

Slices (1/2)
Density Histograms

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Half map: Half map 2

Fileemd_14992_half_map_2.map
AnnotationHalf map 2
Projections & Slices
AxesZYX

Projections

Slices (1/2)
Density Histograms

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Sample components

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Entire : Fap1-bound yeast 80S ribosome in non-rotated state with empty A site

EntireName: Fap1-bound yeast 80S ribosome in non-rotated state with empty A site
Components
  • Complex: Fap1-bound yeast 80S ribosome in non-rotated state with empty A site

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Supramolecule #1: Fap1-bound yeast 80S ribosome in non-rotated state with empty A site

SupramoleculeName: Fap1-bound yeast 80S ribosome in non-rotated state with empty A site
type: complex / Chimera: Yes / ID: 1 / Parent: 0 / Macromolecule list: #1-#85
Source (natural)Organism: Saccharomyces cerevisiae (brewer's yeast) / Strain: W303

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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
Image recordingFilm or detector model: GATAN K2 SUMMIT (4k x 4k) / Average electron dose: 43.4 e/Å2
Electron beamAcceleration voltage: 300 kV / Electron source: FIELD EMISSION GUN
Electron opticsIllumination mode: SPOT SCAN / Imaging mode: BRIGHT FIELD / Nominal defocus max: 3.0 µm / Nominal defocus min: 0.5 µm
Experimental equipment
Model: Titan Krios / Image courtesy: FEI Company

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Image processing

Final reconstructionResolution.type: BY AUTHOR / Resolution: 3.0 Å / Resolution method: FSC 0.143 CUT-OFF / Number images used: 18607
Initial angle assignmentType: NOT APPLICABLE
Final angle assignmentType: NOT APPLICABLE
FSC plot (resolution estimation)

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