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- EMDB-10837: Rix1-Rea1 pre-60S particle - Rea1, body 3 (rigid body refinement,... -

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Basic information

Entry
Database: EMDB / ID: EMD-10837
TitleRix1-Rea1 pre-60S particle - Rea1, body 3 (rigid body refinement, composite structure of Rea1 ring and tail)
Map data
SampleRix1-Rea1 pre-60S assembly particle - Rea1 with the Rsa4-UBL domain:
Midasin / Ribosome assembly protein 4
Function / homology
Function and homology information


regulation of ribosomal subunit export from nucleus / protein-RNA complex remodeling / preribosome, large subunit precursor / rRNA processing / ribosomal large subunit assembly / ATPase activity / nucleolus / mitochondrion / nucleoplasm / ATP binding / nucleus
ATPase, dynein-related, AAA domain / WD40 repeat / WD40-repeat-containing domain / WD40/YVTN repeat-like-containing domain superfamily / NLE / Midasin / Sigma-54 interaction domain, ATP-binding site 1 / AAA+ ATPase domain / von Willebrand factor, type A / WD40 repeat, conserved site ...ATPase, dynein-related, AAA domain / WD40 repeat / WD40-repeat-containing domain / WD40/YVTN repeat-like-containing domain superfamily / NLE / Midasin / Sigma-54 interaction domain, ATP-binding site 1 / AAA+ ATPase domain / von Willebrand factor, type A / WD40 repeat, conserved site / G-protein beta WD-40 repeat / P-loop containing nucleoside triphosphate hydrolase / WD40-repeat-containing domain superfamily / von Willebrand factor A-like domain superfamily / Midasin, AAA lid domain 7 / Midasin AAA lid domain 5
Ribosome assembly protein 4 / Midasin
Biological speciesSaccharomyces cerevisiae (baker's yeast) / Baker's yeast (baker's yeast)
Methodsingle particle reconstruction / cryo EM / Resolution: 4.2 Å
AuthorsKater L / Beckmann R
CitationJournal: Mol. Cell / Year: 2020
Title: Construction of the Central Protuberance and L1 Stalk during 60S Subunit Biogenesis.
Authors: Lukas Kater / Valentin Mitterer / Matthias Thoms / Jingdong Cheng / Otto Berninghausen / Roland Beckmann / Ed Hurt /
Abstract: Ribosome assembly is driven by numerous assembly factors, including the Rix1 complex and the AAA ATPase Rea1. These two assembly factors catalyze 60S maturation at two distinct states, triggering ...Ribosome assembly is driven by numerous assembly factors, including the Rix1 complex and the AAA ATPase Rea1. These two assembly factors catalyze 60S maturation at two distinct states, triggering poorly understood large-scale structural transitions that we analyzed by cryo-electron microscopy. Two nuclear pre-60S intermediates were discovered that represent previously unknown states after Rea1-mediated removal of the Ytm1-Erb1 complex and reveal how the L1 stalk develops from a pre-mature nucleolar to a mature-like nucleoplasmic state. A later pre-60S intermediate shows how the central protuberance arises, assisted by the nearby Rix1-Rea1 machinery, which was solved in its pre-ribosomal context to molecular resolution. This revealed a Rix1-Ipi3 tetramer anchored to the pre-60S via Ipi1, strategically positioned to monitor this decisive remodeling. These results are consistent with a general underlying principle that temporarily stabilized immature RNA domains are successively remodeled by assembly factors, thereby ensuring failsafe assembly progression.
Validation ReportPDB-ID: 6ylf

SummaryFull reportAbout validation report
History
DepositionApr 7, 2020-
Header (metadata) releaseJul 29, 2020-
Map releaseJul 29, 2020-
UpdateJul 29, 2020-
Current statusJul 29, 2020Processing site: PDBe / Status: Released

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Structure visualization

Movie
  • Surface view with section colored by density value
  • Surface level: 0.03
  • Imaged by UCSF Chimera
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  • Surface view colored by height
  • Surface level: 0.03
  • Imaged by UCSF Chimera
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  • Surface view with fitted model
  • Atomic models: PDB-6ylf
  • Surface level: 0.03
  • Imaged by UCSF Chimera
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Movie viewer
Structure viewerEM map:
SurfViewMolmilJmol/JSmol
Supplemental images

Downloads & links

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Map

FileDownload / File: emd_10837.map.gz / Format: CCP4 / Size: 163.6 MB / Type: IMAGE STORED AS FLOATING POINT NUMBER (4 BYTES)
Projections & slices

Image control

Size
Brightness
Contrast
Others
AxesZ (Sec.)Y (Row.)X (Col.)
1.06 Å/pix.
x 350 pix.
= 370.65 Å
1.06 Å/pix.
x 350 pix.
= 370.65 Å
1.06 Å/pix.
x 350 pix.
= 370.65 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
Contour LevelBy AUTHOR: 0.03 / Movie #1: 0.03
Minimum - Maximum-0.0015304224 - 0.14313743
Average (Standard dev.)0.00037300837 (±0.0035066353)
SymmetrySpace group: 1
Details

EMDB XML:

Map geometry
Axis orderXYZ
Origin000
Dimensions350350350
Spacing350350350
CellA=B=C: 370.65 Å
α=β=γ: 90.0 °

CCP4 map header:

modeImage stored as Reals
Å/pix. X/Y/Z1.0591.0591.059
M x/y/z350350350
origin x/y/z0.0000.0000.000
length x/y/z370.650370.650370.650
α/β/γ90.00090.00090.000
MAP C/R/S123
start NC/NR/NS000
NC/NR/NS350350350
D min/max/mean-0.0020.1430.000

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Supplemental data

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Additional map: Local resolution filtered map of the Tail of...

Fileemd_10837_additional_1.map
AnnotationLocal resolution filtered map of the Tail of Rea1 of the nucleoplasmic Rix1-Rea1 pre-60S particle
Projections & Slices
AxesZYX

Projections

Slices (1/2)
Density Histograms

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Additional map: Local resolution filtered map of the Ring of...

Fileemd_10837_additional_2.map
AnnotationLocal resolution filtered map of the Ring of Rea1 of the nucleoplasmic Rix1-Rea1 pre-60S particle
Projections & Slices
AxesZYX

Projections

Slices (1/2)
Density Histograms

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Sample components

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Entire Rix1-Rea1 pre-60S assembly particle - Rea1 with the Rsa4-UBL domain

EntireName: Rix1-Rea1 pre-60S assembly particle - Rea1 with the Rsa4-UBL domain
Details: Rix1-TAP Flag-Rea1 derived pre-60S assembly complex
Number of components: 3

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Component #1: protein, Rix1-Rea1 pre-60S assembly particle - Rea1 with the Rsa4...

ProteinName: Rix1-Rea1 pre-60S assembly particle - Rea1 with the Rsa4-UBL domain
Details: Rix1-TAP Flag-Rea1 derived pre-60S assembly complex
Recombinant expression: No
SourceSpecies: Saccharomyces cerevisiae (baker's yeast)

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Component #2: protein, Midasin

ProteinName: Midasin / Number of Copies: 1 / Recombinant expression: No
MassTheoretical: 559.951438 kDa
SourceSpecies: Baker's yeast (baker's yeast)

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Component #3: protein, Ribosome assembly protein 4

ProteinName: Ribosome assembly protein 4 / Number of Copies: 1 / Recombinant expression: No
MassTheoretical: 57.106781 kDa
SourceSpecies: Baker's yeast (baker's yeast)

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Experimental details

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Sample preparation

SpecimenSpecimen state: Particle / Method: cryo EM
Sample solutionpH: 7.5
VitrificationInstrument: FEI VITROBOT MARK IV / Cryogen name: ETHANE / Temperature: 277 K / Humidity: 90 %

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Electron microscopy imaging

Experimental equipment
Model: Titan Krios / Image courtesy: FEI Company
ImagingMicroscope: FEI TITAN KRIOS
Electron gunElectron source: FIELD EMISSION GUN / Accelerating voltage: 300 kV / Electron dose: 75 e/Å2 / Illumination mode: FLOOD BEAM
LensImaging mode: BRIGHT FIELD
Specimen HolderModel: FEI TITAN KRIOS AUTOGRID HOLDER
CameraDetector: GATAN K2 QUANTUM (4k x 4k)

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Image acquisition

Image acquisitionSampling size: 5 µm

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Image processing

ProcessingMethod: single particle reconstruction / Applied symmetry: C1 (asymmetric) / Number of projections: 55397
3D reconstructionAlgorithm: FOURIER SPACE / Software: RELION / Resolution: 4.2 Å / Resolution method: FSC 0.143 CUT-OFF

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Atomic model buiding

Modeling #1Refinement protocol: rigid body / Refinement space: REAL
Input PDB model: 6HYP, 6HYD, 6OR5, 6QTA
Output model

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