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- EMDB-14508: 90S particle/90S-pre-60S bipartite particle (Urb2-Noc1 mak5 D333A) -

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Basic information

Entry
Database: EMDB / ID: EMD-14508
Title90S particle/90S-pre-60S bipartite particle (Urb2-Noc1 mak5 D333A)
Map data90S particle Urb2-Noc1 mak5 D333A purification (Local filtered map)
Sample
  • Complex: 90S particle Urb2-Noc1 mak5 D333A purification
Biological speciesSaccharomyces cerevisiae (brewer's yeast)
Methodsingle particle reconstruction / cryo EM / Resolution: 7.25 Å
AuthorsIsmail S / Flemming D / Thoms M / Beckmann R / Hurt E
Funding support Germany, European Union, 3 items
OrganizationGrant numberCountry
German Research Foundation (DFG)HU 363/15-2 Germany
German Research Foundation (DFG)GRK1721 Germany
European Research Council (ERC)ADG 741781European Union
CitationJournal: Cell Rep / Year: 2022
Title: Emergence of the primordial pre-60S from the 90S pre-ribosome.
Authors: Sherif Ismail / Dirk Flemming / Matthias Thoms / José Vicente Gomes-Filho / Lennart Randau / Roland Beckmann / Ed Hurt /
Abstract: Synthesis of ribosomes begins in the nucleolus with formation of the 90S pre-ribosome, during which the pre-40S and pre-60S pathways diverge by pre-rRNA cleavage. However, it remains unclear how, ...Synthesis of ribosomes begins in the nucleolus with formation of the 90S pre-ribosome, during which the pre-40S and pre-60S pathways diverge by pre-rRNA cleavage. However, it remains unclear how, after this uncoupling, the earliest pre-60S subunit continues to develop. Here, we reveal a large-subunit intermediate at the beginning of its construction when still linked to the 90S, the precursor to the 40S subunit. This primordial pre-60S is characterized by the SPOUT domain methyltransferase Upa1-Upa2, large α-solenoid scaffolds, Mak5, one of several RNA helicases, and two small nucleolar RNA (snoRNAs), C/D box snR190 and H/ACA box snR37. The emerging pre-60S does not efficiently disconnect from the 90S pre-ribosome in a dominant mak5 helicase mutant, allowing a 70-nm 90S-pre-60S bipartite particle to be visualized by electron microscopy. Our study provides insight into the assembly pathway when the still-connected nascent 40S and 60S subunits are beginning to separate.
History
DepositionMar 4, 2022-
Header (metadata) releaseMay 11, 2022-
Map releaseMay 11, 2022-
UpdateMay 11, 2022-
Current statusMay 11, 2022Processing site: PDBe / Status: Released

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Structure visualization

Supplemental images

emd_14508.png

Downloads & links

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Map

FileDownload / File: emd_14508.map.gz / Format: CCP4 / Size: 30.5 MB / Type: IMAGE STORED AS FLOATING POINT NUMBER (4 BYTES)
Annotation90S particle Urb2-Noc1 mak5 D333A purification (Local filtered map)
Projections & slices

Image control

Size
Brightness
Contrast
Others
AxesZ (Sec.)Y (Row.)X (Col.)
3.17 Å/pix.
x 200 pix.
= 634. Å		3.17 Å/pix.
x 200 pix.
= 634. Å		3.17 Å/pix.
x 200 pix.
= 634. Å

Surface

Projections

Slices (1/3)

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Images are generated by Spider.

Voxel sizeX=Y=Z: 3.17 Å
Density

Histogram

Histogram (log scale)
Contour LevelBy AUTHOR: 1.15
Minimum - Maximum-1.3196378 - 4.933927
Average (Standard dev.)0.069078535 (±0.3743329)
SymmetrySpace group: 1
Details

EMDB XML:

Map geometry
Axis orderXYZ
Origin000
Dimensions200200200
Spacing200200200
CellA=B=C: 634.0 Å
α=β=γ: 90.0 °

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Supplemental data

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Additional map: 90S-pre-60S bipartite particle Urb2-Noc1 mak5 D333A purification (90S...

Fileemd_14508_additional_1.map
Annotation90S-pre-60S bipartite particle Urb2-Noc1 mak5 D333A purification (90S particles re-extracted with a box size of 450x450 pixel) (Local filtered map)
Projections & Slices
AxesZYX

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Additional map: 90S-pre-60S bipartite particle Urb2-Noc1 mak5 D333A purification (90S...

Fileemd_14508_additional_2.map
Annotation90S-pre-60S bipartite particle Urb2-Noc1 mak5 D333A purification (90S particles re-extracted with a box size of 450x450 pixel) (Homogeneous Refinement)
Projections & Slices
AxesZYX

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Density Histograms

Histogram

Histogram (log scale)

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Half map: 90S particle Urb2-Noc1 mak5 D333A purification (Half map A)

Fileemd_14508_half_map_1.map
Annotation90S particle Urb2-Noc1 mak5 D333A purification (Half map A)
Projections & Slices
AxesZYX

Projections

Slices (1/2)
Density Histograms

Histogram

Histogram (log scale)

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Half map: 90S particle Urb2-Noc1 mak5 D333A purification (Half map B)

Fileemd_14508_half_map_2.map
Annotation90S particle Urb2-Noc1 mak5 D333A purification (Half map B)
Projections & Slices
AxesZYX

Projections

Slices (1/2)
Density Histograms

Histogram

Histogram (log scale)

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Sample components

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Entire : 90S particle Urb2-Noc1 mak5 D333A purification

EntireName: 90S particle Urb2-Noc1 mak5 D333A purification
Components
  • Complex: 90S particle Urb2-Noc1 mak5 D333A purification

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Supramolecule #1: 90S particle Urb2-Noc1 mak5 D333A purification

SupramoleculeName: 90S particle Urb2-Noc1 mak5 D333A purification / type: complex / Chimera: Yes / ID: 1 / Parent: 0
Details: The homogeneous refined and local filtered maps of the 90S-pre-60S bipartite particle (enlarged box size 450 x 450 pixel) are attached as additional maps.
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

MicroscopeTFS GLACIOS
Image recordingFilm or detector model: FEI FALCON III (4k x 4k) / Average electron dose: 60.0 e/Å2
Electron beamAcceleration voltage: 200 kV / Electron source: FIELD EMISSION GUN
Electron opticsIllumination mode: SPOT SCAN / Imaging mode: BRIGHT FIELD / Nominal defocus max: 2.6 µm / Nominal defocus min: 1.2 µm

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Image processing

Final reconstructionResolution.type: BY AUTHOR / Resolution: 7.25 Å / Resolution method: FSC 0.143 CUT-OFF / Number images used: 24935
Initial angle assignmentType: MAXIMUM LIKELIHOOD
Final angle assignmentType: MAXIMUM LIKELIHOOD

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