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- EMDB-3889: State B (Nsa1-TAP Flag-Ytm1) - Visualizing the assembly pathway o... -

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Entry
Database: EMDB / ID: 3889
TitleState B (Nsa1-TAP Flag-Ytm1) - Visualizing the assembly pathway of nucleolar pre-60S ribosomes
Map dataState B (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 / homologyRibosomal protein L1-like / WD40-repeat-containing domain superfamily / Ribosomal protein L1/ribosomal biogenesis protein / WD repeat BOP1/Erb1 / 50S ribosomal protein L18Ae/60S ribosomal protein L20 and L18a / Ribosomal L28e/Mak16 / 50S ribosomal protein L30e-like / Ribosome biogenesis protein 15, RNA recognition motif / Ribosomal protein L7, eukaryotic/archaeal / RNA-binding domain superfamily ...Ribosomal protein L1-like / WD40-repeat-containing domain superfamily / Ribosomal protein L1/ribosomal biogenesis protein / WD repeat BOP1/Erb1 / 50S ribosomal protein L18Ae/60S ribosomal protein L20 and L18a / Ribosomal L28e/Mak16 / 50S ribosomal protein L30e-like / Ribosome biogenesis protein 15, RNA recognition motif / Ribosomal protein L7, eukaryotic/archaeal / RNA-binding domain superfamily / Ribosomal protein L29/L35 superfamily / Ribosomal L18e/L15P superfamily / Ribosomal protein L32e superfamily / Ribosome biogenesis protein BRX1 / Ribosomal protein L22/L17 superfamily / BRCT domain superfamily / Ribosomal protein L14 superfamily / Ribosomal protein L13 superfamily / Ribosomal protein L30, ferredoxin-like fold domain superfamily / WDR74/Nsa1 / Ribosomal protein L36e domain superfamily / Ribosomal protein L7A/L8 superfamily / Ribosomal protein L24e/L24 superfamily / Ribosomal protein L35A superfamily / RNA recognition motif. (a.k.a. RRM, RBD, or RNP domain) / P-loop containing nucleoside triphosphate hydrolase / 60S ribosomal protein L4, C-terminal domain / Ribosomal protein L14p/L23e / Ribosome biogenesis protein Nop16 / Ribosomal protein L1, 3-layer alpha/beta-sandwich / WD40-repeat-containing domain / Ribosomal protein L30, conserved site / Ribosomal protein L29, conserved site / Ribosomal protein L13e, conserved site / Ribosomal protein L22/L17, conserved site / Ribosomal protein L32e, conserved site / Ribosomal protein L35Ae, conserved site / Ribosomal protein L37e, conserved site / Ribosomal protein L7Ae/L8/Nhp2 family / WD40 repeat, conserved site / Domain of unknown function DUF4217 / Ribosomal protein L3, conserved site / Ribosomal protein L14P, conserved site / Ribosomal protein L15e, conserved site / Ribosomal protein L18e/L15P / Ribosomal protein L18e, conserved site / 60S ribosomal protein L18a/ L20, eukaryotes / Ribosomal protein L30, central domain, archaeal type / Ribosomal protein L13, conserved site / Ribosomal protein 50S-L18Ae/60S-L20/60S-L18A / Ribosomal protein L4 domain superfamily / Ribosomal protein L15e core domain superfamily / Ribosomal protein L22p/L17e / DEAD/DEAH box helicase / Ribosomal protein L2, domain 2 / Ribosomal protein L3 signature. / BOP1NT (NUC169) domain / Ribosome biogenesis protein Nop16 / Domain of unknown function (DUF4217) / 60S ribosomal protein L4 C-terminal domain / BRCT domain, a BRCA1 C-terminus domain / Ribosomal proteins L26 eukaryotic, L24P archaeal / Ribosomal protein 60S L18 and 50S L18e / DEAD-box subfamily ATP-dependent helicases signature. / Ribosomal protein L14 signature. / Ribosomal protein L22 signature. / Ribosomal protein L29 signature. / Pescadillo N-terminus / Ribosomal protein L32e signature. / Ribosomal protein L30 signature. / Trp-Asp (WD) repeats signature. / Ribosomal protein L13 signature. / Ribosomal protein L1e signature. / Ribosomal protein L37e signature. / Ribosomal protein L7Ae signature. / Ribosomal protein L13e signature. / Ribosomal protein L35Ae signature. / Ribosomal protein L18e signature. / Ribosomal protein L24 signature. / Ribosomal L30 N-terminal domain / Nucleolar protein,Nop52 / Helicase conserved C-terminal domain / Ribosomal protein L6e / Ribosomal protein L3 / Ribosomal protein L30p/L7e / WD domain, G-beta repeat / KOW motif / Ribosomal protein L13 / Ribosomal protein L4/L1 family / Ribosomal protein L1p/L10e family / Ribosomal L15 / Ribosomal L29 protein / Ribosomal protein L36e / Ribosomal protein L24e / Eukaryotic rRNA processing protein EBP2 / Ribosomal protein L35Ae / Ribosomal protein L7Ae/L30e/S12e/Gadd45 family / Ribosomal protein L13e / Ribosomal protein L32 / Ribosomal proteins 50S-L18Ae/60S-L20/60S-L18A
Function and homology information
SourceSaccharomyces cerevisiae (baker's yeast)
Methodsingle particle reconstruction / cryo EM / 4.1 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.
Validation ReportPDB-ID: 6em4

SummaryFull reportAbout validation report
DateDeposition: Sep 29, 2017 / Header (metadata) release: Dec 27, 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: 0.03
  • Imaged by UCSF Chimera
  • Download
  • Surface view colored by radius
  • Surface level: 0.03
  • Imaged by UCSF Chimera
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  • Surface view with fitted model
  • Atomic models: : PDB-6em4
  • 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

Fileemd_3889.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.

Voxel sizeX=Y=Z: 1.084 Å
Density
Contour Level:0.03 (by author), 0.03 (movie #1):
Minimum - Maximum-0.036213778 - 0.13052347
Average (Standard dev.)0.00058982155 (0.006162797)
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-0.0360.1310.001

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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 (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

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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: 65539
3D reconstructionAlgorithm: FOURIER SPACE / Software: RELION / Resolution: 4.1 Å / Resolution method: FSC 0.143 CUT-OFF
FSC plot
(resolution estimation)

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Atomic model buiding

Output model

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