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- EMDB-29145: Human nucleolar pre-60S ribosomal subunit (State C2) - Domain I f... -

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Basic information

Entry
Database: EMDB / ID: EMD-29145
TitleHuman nucleolar pre-60S ribosomal subunit (State C2) - Domain I focused map
Map data
Sample
  • Complex: Human nucleolar pre-60S ribosomal subunit
KeywordsPre-60S ribosomal subunit / Assembly intermediate / Ribosome / Nucleoprotein complex
Biological speciesHomo sapiens (human)
Methodsingle particle reconstruction / cryo EM / Resolution: 2.82 Å
AuthorsVanden Broeck A / Klinge S
Funding supportEuropean Union, United States, 2 items
OrganizationGrant numberCountry
European Molecular Biology Organization (EMBO)ALTF 711-2019European Union
The G. Harold and Leila Y. Mathers FoundationMF-2104-01554 United States
CitationJournal: Science / Year: 2023
Title: Principles of human pre-60 biogenesis.
Authors: Arnaud Vanden Broeck / Sebastian Klinge /
Abstract: During the early stages of human large ribosomal subunit (60) biogenesis, an ensemble of assembly factors establishes and fine-tunes the essential RNA functional centers of pre-60 particles by an ...During the early stages of human large ribosomal subunit (60) biogenesis, an ensemble of assembly factors establishes and fine-tunes the essential RNA functional centers of pre-60 particles by an unknown mechanism. Here, we report a series of cryo-electron microscopy structures of human nucleolar and nuclear pre-60 assembly intermediates at resolutions of 2.5 to 3.2 angstroms. These structures show how protein interaction hubs tether assembly factor complexes to nucleolar particles and how guanosine triphosphatases and adenosine triphosphatase couple irreversible nucleotide hydrolysis steps to the installation of functional centers. Nuclear stages highlight how a conserved RNA-processing complex, the rixosome, couples large-scale RNA conformational changes with pre-ribosomal RNA processing by the RNA degradation machinery. Our ensemble of human pre-60 particles provides a rich foundation with which to elucidate the molecular principles of ribosome formation.
History
DepositionDec 15, 2022-
Header (metadata) releaseJul 12, 2023-
Map releaseJul 12, 2023-
UpdateJul 19, 2023-
Current statusJul 19, 2023Processing site: RCSB / Status: Released

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Structure visualization

Supplemental images

emd_29145.png

Downloads & links

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Map

FileDownload / File: emd_29145.map.gz / Format: CCP4 / Size: 421.9 MB / Type: IMAGE STORED AS FLOATING POINT NUMBER (4 BYTES)
Voxel sizeX=Y=Z: 1.072 Å
Density
Contour LevelBy AUTHOR: 0.65
Minimum - Maximum-1.8361639 - 3.7782671
Average (Standard dev.)0.002938118 (±0.052548192)
SymmetrySpace group: 1
Details

EMDB XML:

Map geometry
Axis orderXYZ
Origin000
Dimensions480480480
Spacing480480480
CellA=B=C: 514.56 Å
α=β=γ: 90.0 °

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Supplemental data

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Mask #1

Fileemd_29145_msk_1.map
Projections & Slices
AxesZYX

Projections

Slices (1/2)
Density Histograms

Histogram

Histogram (log scale)

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Half map: #1

Fileemd_29145_half_map_1.map
Projections & Slices
AxesZYX

Projections

Slices (1/2)
Density Histograms

Histogram

Histogram (log scale)

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Half map: #2

Fileemd_29145_half_map_2.map
Projections & Slices
AxesZYX

Projections

Slices (1/2)
Density Histograms

Histogram

Histogram (log scale)

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Sample components

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Entire : Human nucleolar pre-60S ribosomal subunit

EntireName: Human nucleolar pre-60S ribosomal subunit
Components
  • Complex: Human nucleolar pre-60S ribosomal subunit

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Supramolecule #1: Human nucleolar pre-60S ribosomal subunit

SupramoleculeName: Human nucleolar pre-60S ribosomal subunit / type: complex / ID: 1 / Parent: 0
Source (natural)Organism: Homo sapiens (human) / Strain: HEK293F

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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: 7.6
GridModel: Quantifoil R3.5/1 / Material: GOLD / Mesh: 400 / Support film - Material: CARBON / Support film - topology: CONTINUOUS / Support film - Film thickness: 2 / Pretreatment - Type: GLOW DISCHARGE / Pretreatment - Time: 30 sec.
VitrificationCryogen name: ETHANE / Chamber humidity: 95 % / Chamber temperature: 283 K / Instrument: FEI VITROBOT MARK IV
Details: Four applications with manual blotting before last blotting with the vitrobot..

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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 / Nominal defocus max: 2.5 µm / Nominal defocus min: 0.5 µm / Nominal magnification: 64000
Specialist opticsEnergy filter - Slit width: 20 eV
Image recordingFilm or detector model: GATAN K3 (6k x 4k) / Number grids imaged: 4 / Number real images: 172699 / Average exposure time: 2.0 sec. / Average electron dose: 60.0 e/Å2
Experimental equipment
Model: Titan Krios / Image courtesy: FEI Company

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Image processing

Particle selectionNumber selected: 15679142
Startup modelType of model: INSILICO MODEL
Initial angle assignmentType: MAXIMUM LIKELIHOOD / Software - Name: RELION (ver. 4.0)
Final 3D classificationSoftware: (Name: RELION (ver. 4.0), cryoSPARC (ver. 3.3.2))
Final angle assignmentType: MAXIMUM LIKELIHOOD / Software - Name: cryoSPARC (ver. 3.3.2)
Final reconstructionApplied symmetry - Point group: C1 (asymmetric) / Resolution.type: BY AUTHOR / Resolution: 2.82 Å / Resolution method: FSC 0.143 CUT-OFF / Software - Name: cryoSPARC (ver. 3.3.2) / Number images used: 48037
FSC plot (resolution estimation)

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