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- EMDB-3893: State C (Nsa1-TAP Flag-Ytm1) - Visualizing the assembly pathway o... -

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Entry
Database: EMDB / ID: 3893
TitleState C (Nsa1-TAP Flag-Ytm1) - Visualizing the assembly pathway of nucleolar pre-60S ribosomes
Map dataState C (Nsa1-TAP Flag-Ytm1) - Visualizing the assembly pathway of nucleolar pre-60S ribosmes
SampleLarge subunit biogenesis particle purified via Nsa1-TAP and Flag-Ytm1
Function/homologyGuanine nucleotide-binding protein-like 3, N-terminal domain / Uncharacterised protein family UPF0642 / Eukaryotic rRNA processing / Mak16 protein / snoRNA release from pre-rRNA / Ribosome biogenesis protein Nop16 / GTP-binding protein, orthogonal bundle domain superfamily / Ribosome biogenesis protein BRX1 / WDR74/Nsa1 / GNL3L/Grn1 putative GTPase ...Guanine nucleotide-binding protein-like 3, N-terminal domain / Uncharacterised protein family UPF0642 / Eukaryotic rRNA processing / Mak16 protein / snoRNA release from pre-rRNA / Ribosome biogenesis protein Nop16 / GTP-binding protein, orthogonal bundle domain superfamily / Ribosome biogenesis protein BRX1 / WDR74/Nsa1 / GNL3L/Grn1 putative GTPase / Protein of unknown function (DUF2423) / Ribosome assembly factor Mrt4 / Nucleolar, Nop52 / Nucleolar GTP-binding protein 1 / WD repeat BOP1/Erb1 / NOG, C-terminal / BOP1, N-terminal domain / Nucleolar GTP-binding protein 1, Rossman-fold domain / Domain of unknown function DUF4217 / nuclear division / Pescadillo / Eukaryotic rRNA processing protein EBP2 / Mak16 protein C-terminal region / exonucleolytic trimming to generate mature 5'-end of 5.8S rRNA from tricistronic rRNA transcript (SSU-rRNA, 5.8S rRNA, LSU-rRNA) / Ribosome biogenesis protein 15, RNA recognition motif / Ribosome biogenesis protein Nop16 / BOP1NT (NUC169) domain / Nucleolar protein,Nop52 / Domain of unknown function (DUF4217) / PeBoW complex / NOGCT (NUC087) domain / Pescadillo N-terminus / Nucleolar GTP-binding protein 1 (NOG1) / rRNA primary transcript binding / Circularly permuted (CP)-type guanine nucleotide-binding (G) domain profile. / Circularly permuted (CP)-type guanine nucleotide-binding (G) domain / Brix domain / Brix domain profile. / Translation initiation factor IF6 / OBG-type guanine nucleotide-binding (G) domain profile. / OBG-type guanine nucleotide-binding (G) domain / maturation of 5.8S rRNA from tricistronic rRNA transcript (SSU-rRNA, 5.8S rRNA, LSU-rRNA) / actomyosin contractile ring assembly / eIF-6 family / Brix domain / mature ribosome assembly / regulation of cell division / RNA-dependent ATPase activity / ribosomal subunit export from nucleus / Ribosomal L28e/Mak16 / BRCT domain, a BRCA1 C-terminus domain / nuclear-transcribed mRNA catabolic process / cleavage in ITS2 between 5.8S rRNA and LSU-rRNA of tricistronic rRNA transcript (SSU-rRNA, 5.8S rRNA, LSU-rRNA) / GTP binding domain / Ribosomal L28e protein family / ATPase activator activity / ribosomal large subunit binding / preribosome, small subunit precursor / DEAD-box subfamily ATP-dependent helicases signature. / ATP-dependent RNA helicase DEAD-box, conserved site / Ribosomal protein L13e, conserved site / preribosome, large subunit precursor / Ribosomal protein L13e signature. / TRASH domain / Ribosomal protein L13e / 60S ribosomal protein L18a/ L20, eukaryotes / Ribosomal protein 50S-L18Ae/60S-L20/60S-L18A / 50S ribosomal protein L18Ae/60S ribosomal protein L20 and L18a / Ribosomal protein L6e signature. / DEAD-box RNA helicase Q motif profile. / RNA helicase, DEAD-box type, Q motif / proteasome binding / 50S ribosome-binding GTPase / SRP-dependent cotranslational protein targeting to membrane / GTP hydrolysis and joining of the 60S ribosomal subunit / Ribosomal protein L35Ae signature. / Ribosomal protein L35Ae, conserved site / RNA secondary structure unwinding / Ribosomal protein L36e signature. / maturation of LSU-rRNA from tricistronic rRNA transcript (SSU-rRNA, 5.8S rRNA, LSU-rRNA) / Formation of a pool of free 40S subunits / Ribosomal protein L7, eukaryotic / 60S ribosomal protein L6E / Ribosomal protein L30, N-terminal / Ribosomal protein L7A/L8 / Nonsense Mediated Decay (NMD) independent of the Exon Junction Complex (EJC) / Ribosomal protein L36e / Nonsense Mediated Decay (NMD) enhanced by the Exon Junction Complex (EJC) / 60S ribosomal protein L4, C-terminal domain / Ribosomal protein L35A / Ribosomal protein L14e domain / Ribosomal protein S8e/ribosomal biogenesis NSA2 / Ribosomal protein L18e signature. / Ribosomal protein L18e, conserved site / ribosomal large subunit export from nucleus / Ribosomal protein L13, eukaryotic/archaeal / Ribosomal protein L18e / Ribosomal protein L13e / ribosomal large subunit biogenesis / L13a-mediated translational silencing of Ceruloplasmin expression
Function and homology information
SourceSaccharomyces cerevisiae / / yeast /
MethodCryo EM / single particle reconstruction / 3.6 Å resolution
AuthorsKater L / Thoms M / Barrio-Garcia C / Cheng J / Ismail S / Ahmed YL / Bange G / Kressler D / Berninghausen O / Sinning I / Hurt E / Beckmann R
CitationJournal: Cell / Year: 2017
Title: Visualizing the Assembly Pathway of Nucleolar Pre-60S Ribosomes.
Authors: Lukas Kater / Matthias Thoms / Clara Barrio-Garcia / Jingdong Cheng / Sherif Ismail / Yasar Luqman Ahmed / Gert Bange / Dieter Kressler / Otto Berninghausen / Irmgard Sinning / Ed Hurt / Roland Beckmann
Abstract: Eukaryotic 60S ribosomal subunits are comprised of three rRNAs and ∼50 ribosomal proteins. The initial steps of their formation take place in the nucleolus, but, owing to a lack of structural ...Eukaryotic 60S ribosomal subunits are comprised of three rRNAs and ∼50 ribosomal proteins. The initial steps of their formation take place in the nucleolus, but, owing to a lack of structural information, this process is poorly understood. Using cryo-EM, we solved structures of early 60S biogenesis intermediates at 3.3 Å to 4.5 Å resolution, thereby providing insights into their sequential folding and assembly pathway. Besides revealing distinct immature rRNA conformations, we map 25 assembly factors in six different assembly states. Notably, the Nsa1-Rrp1-Rpf1-Mak16 module stabilizes the solvent side of the 60S subunit, and the Erb1-Ytm1-Nop7 complex organizes and connects through Erb1's meandering N-terminal extension, eight assembly factors, three ribosomal proteins, and three 25S rRNA domains. Our structural snapshots reveal the order of integration and compaction of the six major 60S domains within early nucleolar 60S particles developing stepwise from the solvent side around the exit tunnel to the central protuberance.
Copyright: 2017 The Authors. Published by Elsevier Inc. All rights reserved.
Validation ReportPDB-ID: 6em1

SummaryFull reportAbout validation report
DateDeposition: Sep 29, 2017 / Header (metadata) release: Dec 20, 2017 / Map release: Dec 27, 2017 / Last update: Dec 27, 2017

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Structure visualization

Movie
  • Surface view with section colored by density value
  • Surface level: 0.032
  • Imaged by UCSF CHIMERA
  • Download
  • Surface view colored by radius
  • Surface level: 0.032
  • Imaged by UCSF CHIMERA
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  • Surface view with fitted model
  • Atomic models: : PDB-6em1
  • Surface level: 0.032
  • Imaged by UCSF CHIMERA
  • Download
3D viewer
Supplemental images

Downloads & links

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Map

Fileemd_3893.map.gz (map file in CCP4 format, 296353 KB)
Projections & slices

Image control

Size
Brightness
Contrast
Others
AxesZ (Sec.)Y (Row.)X (Col.)
420 pix
1.08 Å/pix.
= 455.28 Å
420 pix
1.08 Å/pix.
= 455.28 Å
420 pix
1.08 Å/pix.
= 455.28 Å

Surface

Projections

Slices (1/3)

Slices (1/2)

Slices (2/3)

Images are generated by Spider package.

Voxel sizeX=Y=Z: 1.084 Å
Density
Contour Level:0.032 (by author), 0.032 (movie #1):
Minimum - Maximum-0.08158503 - 0.19416071
Average (Standard dev.)-0.00014478025 (0.0068711215)
Details

EMDB XML:

Space Group Number1
Map Geometry
Axis orderXYZ
Dimensions420420420
Origin000
Limit419419419
Spacing420420420
CellA=B=C: 455.28 Å
α=β=γ: 90 deg.

CCP4 map header:

modeImage stored as Reals
Å/pix. X/Y/Z1.0841.0841.084
M x/y/z420420420
origin x/y/z0.0000.0000.000
length x/y/z455.280455.280455.280
α/β/γ90.00090.00090.000
start NX/NY/NZ
NX/NY/NZ
MAP C/R/S123
start NC/NR/NS000
NC/NR/NS420420420
D min/max/mean-0.0820.194-0.000

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Supplemental data

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Sample components

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Entire Large subunit biogenesis particle purified via Nsa1-TAP and Flag-Ytm1

EntireName: Large subunit biogenesis particle purified via Nsa1-TAP and Flag-Ytm1
Number of components: 1

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Component #1: protein, Large subunit biogenesis particle purified via Nsa1-TAP ...

ProteinName: Large subunit biogenesis particle purified via Nsa1-TAP and Flag-Ytm1
Recombinant expression: No
SourceSpecies: Saccharomyces cerevisiae / / 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

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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: 27 e/Å2 / Illumination mode: FLOOD BEAM
LensCs: 2.7 mm / Imaging mode: BRIGHT FIELD
Specimen HolderModel: OTHER
CameraDetector: FEI FALCON II (4k x 4k)

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Image processing

ProcessingMethod: single particle reconstruction / Number of projections: 156937
3D reconstructionAlgorithm: FOURIER SPACE / Software: RELION / Resolution: 3.6 Å / Resolution method: FSC 0.143 CUT-OFF
FSC plot (resolution assessment)

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Atomic model buiding

Output model

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