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- EMDB-16276: Release state - ES27up (pre-60S) -

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Basic information

Entry
Database: EMDB / ID: EMD-16276
TitleRelease state - ES27up (pre-60S)
Map dataComposite map
Sample
  • Complex: Release state - ES27up (pre-60S)
Biological speciesSaccharomyces cerevisiae (brewer's yeast)
Methodsingle particle reconstruction / cryo EM / Resolution: 3.0 Å
AuthorsThoms M / Mitterer V / Buschauer R / Berninghausen O / Hurt E / Beckmann R
Funding support Germany, European Union, 3 items
OrganizationGrant numberCountry
German Research Foundation (DFG)HU363/15-2 Germany
European Research Council (ERC)885711European Union
European Research Council (ERC)741781European Union
CitationJournal: Elife / Year: 2023
Title: Concurrent remodelling of nucleolar 60S subunit precursors by the Rea1 ATPase and Spb4 RNA helicase.
Authors: Valentin Mitterer / Matthias Thoms / Robert Buschauer / Otto Berninghausen / Ed Hurt / Roland Beckmann /
Abstract: Biogenesis intermediates of nucleolar ribosomal 60S precursor particles undergo a number of structural maturation steps before they transit to the nucleoplasm and are finally exported into the ...Biogenesis intermediates of nucleolar ribosomal 60S precursor particles undergo a number of structural maturation steps before they transit to the nucleoplasm and are finally exported into the cytoplasm. The AAA-ATPase Rea1 participates in the nucleolar exit by releasing the Ytm1-Erb1 heterodimer from the evolving pre-60S particle. Here, we show that the DEAD-box RNA helicase Spb4 with its interacting partner Rrp17 is further integrated into this maturation event. Spb4 binds to a specific class of late nucleolar pre-60S intermediates, whose cryo-EM structure revealed how its helicase activity facilitates melting and restructuring of 25S rRNA helices H62 and H63/H63a prior to Ytm1-Erb1 release. In vitro maturation of such Spb4-enriched pre-60S particles, incubated with purified Rea1 and its associated pentameric Rix1-complex in the presence of ATP, combined with cryo-EM analysis depicted the details of the Rea1-dependent large-scale pre-ribosomal remodeling. Our structural insights unveil how the Rea1 ATPase and Spb4 helicase remodel late nucleolar pre-60S particles by rRNA restructuring and dismantling of a network of several ribosomal assembly factors.
History
DepositionDec 5, 2022-
Header (metadata) releaseMar 29, 2023-
Map releaseMar 29, 2023-
UpdateMar 29, 2023-
Current statusMar 29, 2023Processing site: PDBe / Status: Released

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Structure visualization

Supplemental images

emd_16276.png

Downloads & links

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Map

FileDownload / File: emd_16276.map.gz / Format: CCP4 / Size: 476.8 MB / Type: IMAGE STORED AS FLOATING POINT NUMBER (4 BYTES)
AnnotationComposite map
Projections & slices

Image control

Size
Brightness
Contrast
Others
AxesZ (Sec.)Y (Row.)X (Col.)
1.06 Å/pix.
x 500 pix.
= 529.5 Å		1.06 Å/pix.
x 500 pix.
= 529.5 Å		1.06 Å/pix.
x 500 pix.
= 529.5 Å

Surface

Projections

Slices (1/3)

Slices (1/2)

Slices (2/3)

Images are generated by Spider.

Voxel sizeX=Y=Z: 1.059 Å
Density

Histogram

Histogram (log scale)
Contour LevelBy AUTHOR: 0.5
Minimum - Maximum0.0 - 6.2436132
Average (Standard dev.)0.015541363 (±0.12726326)
SymmetrySpace group: 1
Details

EMDB XML:

Map geometry
Axis orderXYZ
Origin000
Dimensions500500500
Spacing500500500
CellA=B=C: 529.5 Å
α=β=γ: 90.0 °

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Supplemental data

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Additional map: Foot - Local Refinement - Local filtered map

Fileemd_16276_additional_1.map
AnnotationFoot - Local Refinement - Local filtered map
Projections & Slices
AxesZYX

Projections

Slices (1/2)
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Histogram

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Additional map: ES27 - Local Refinement - Local filtered map

Fileemd_16276_additional_2.map
AnnotationES27 - Local Refinement - Local filtered map
Projections & Slices
AxesZYX

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Additional map: Nog1 - Local Refinement - Local filtered map

Fileemd_16276_additional_3.map
AnnotationNog1 - Local Refinement - Local filtered map
Projections & Slices
AxesZYX

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Additional map: Spb1-MTD - Local Refinement - Local filtered map

Fileemd_16276_additional_4.map
AnnotationSpb1-MTD - Local Refinement - Local filtered map
Projections & Slices
AxesZYX

Projections

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Additional map: Consensus cryo-EM map - Local filtered map

Fileemd_16276_additional_5.map
AnnotationConsensus cryo-EM map - Local filtered map
Projections & Slices
AxesZYX

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Additional map: Consensus cryo-EM map - Homogeneous Refinement

Fileemd_16276_additional_6.map
AnnotationConsensus cryo-EM map - Homogeneous Refinement
Projections & Slices
AxesZYX

Projections

Slices (1/2)
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Additional map: Consensus cryo-EM map - Homogeneous Refinement - sharpened map

Fileemd_16276_additional_7.map
AnnotationConsensus cryo-EM map - Homogeneous Refinement - sharpened map
Projections & Slices
AxesZYX

Projections

Slices (1/2)
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Histogram

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Half map: Consensus cryo-EM map - half map A

Fileemd_16276_half_map_1.map
AnnotationConsensus cryo-EM map - half map A
Projections & Slices
AxesZYX

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Slices (1/2)
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Histogram

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Half map: Consensus cryo-EM map - half map B

Fileemd_16276_half_map_2.map
AnnotationConsensus cryo-EM map - half map B
Projections & Slices
AxesZYX

Projections

Slices (1/2)
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Histogram

Histogram (log scale)

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Sample components

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Entire : Release state - ES27up (pre-60S)

EntireName: Release state - ES27up (pre-60S)
Components
  • Complex: Release state - ES27up (pre-60S)

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Supramolecule #1: Release state - ES27up (pre-60S)

SupramoleculeName: Release state - ES27up (pre-60S) / type: complex / ID: 1 / Chimera: Yes / Parent: 0 / Macromolecule list: #1-#60
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: 7.5
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: SPOT SCAN / Imaging mode: BRIGHT FIELDBright-field microscopy / Nominal defocus max: 3.5 µm / Nominal defocus min: 0.5 µm
Image recordingFilm or detector model: GATAN K2 SUMMIT (4k x 4k) / Average electron dose: 46.8 e/Å2
Experimental equipment
Model: Titan Krios / Image courtesy: FEI Company

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Image processing

Startup modelType of model: EMDB MAP
EMDB ID:

3891

Initial angle assignmentType: MAXIMUM LIKELIHOOD
Final angle assignmentType: MAXIMUM LIKELIHOOD
Final reconstructionResolution.type: BY AUTHOR / Resolution: 3.0 Å / Resolution method: FSC 0.143 CUT-OFF / Number images used: 29118

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