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- EMDB-3890: State D (Nsa1-TAP Flag-Ytm1) - Visualizing the assembly pathway o... -

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Entry
Database: EMDB / ID: 3890
TitleState D (Nsa1-TAP Flag-Ytm1) - Visualizing the assembly pathway of nucleolar pre-60S ribosomes
Map dataState D (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 / homologyDomain of unknown function DUF4217 / 60S ribosomal protein L18a/ L20, eukaryotes / Ribosomal protein L37e, conserved site / Bacterial Fmu (Sun)/eukaryotic nucleolar NOL1/Nop2p, conserved site / Ribosomal protein L7Ae/L8/Nhp2 family / Ribosome biogenesis protein Nop16 / Uncharacterised protein family UPF0642 / WD40 repeat, conserved site / Ribosomal protein L3, conserved site / Ribosomal protein L14P, conserved site ...Domain of unknown function DUF4217 / 60S ribosomal protein L18a/ L20, eukaryotes / Ribosomal protein L37e, conserved site / Bacterial Fmu (Sun)/eukaryotic nucleolar NOL1/Nop2p, conserved site / Ribosomal protein L7Ae/L8/Nhp2 family / Ribosome biogenesis protein Nop16 / Uncharacterised protein family UPF0642 / WD40 repeat, conserved site / Ribosomal protein L3, conserved site / Ribosomal protein L14P, conserved site / Ribosomal protein L6, alpha-beta domain / Ribosomal protein L31e, conserved site / G-protein beta WD-40 repeat / Ribosomal protein L15e, conserved site / Ribosomal protein L18e/L15P / Ribosomal protein L18e, conserved site / Ribosomal protein S8e/ribosomal biogenesis NSA2 / Ribosomal protein L32e, conserved site / Ribosomal protein L31e domain superfamily / Ribosomal proteins 50S-L18Ae/60S-L20/60S-L18A / Nucleolar GTP-binding protein 1 / Ribosomal protein L15e core domain superfamily / Ribosomal RNA methyltransferase Spb1, domain of unknown function DUF3381 / Ribosomal protein L1-like / Ribosomal protein L19/L19e conserved site / Ribosomal protein L4 domain superfamily / Ribosomal protein L30e, conserved site / Ribosomal protein 50S-L18Ae/60S-L20/60S-L18A / Ribosomal protein L13, conserved site / RNA (C5-cytosine) methyltransferase, NOP2 / RNA (C5-cytosine) methyltransferase / GTP-binding protein, orthogonal bundle domain superfamily / Ribosomal protein L30, central domain, archaeal type / Ribosomal protein L35Ae, conserved site / Ribosomal protein L27e, conserved site / Ribosome biogenesis protein BRX1 / Ribosomal protein L2, domain 2 / Zinc-binding ribosomal protein / Nucleolar complex-associated protein 3, N-terminal / DEAD/DEAH box helicase domain / Nucleotide-binding alpha-beta plait domain superfamily / Ribosomal protein L23/L15e core domain superfamily / Ribosomal RNA methyltransferase, Spb1, C-terminal / BOP1, N-terminal domain / NLE / NOG, C-terminal / Ribosomal protein L30, N-terminal / Ribosomal protein L4/L1e, eukaryotic/archaeal, conserved site / Ribosomal protein L25/L23 / Helicase superfamily 1/2, ATP-binding domain / RNA helicase, DEAD-box type, Q motif / Guanine nucleotide-binding protein-like 3, N-terminal domain / Ribosomal protein L22/L17, conserved site / Nucleolar complex-associated protein 3 / Ribosomal protein L21e, conserved site / Ribosomal protein L13e, conserved site / Ribosomal protein L29, conserved site / Ribosomal protein L34e, conserved site / Ribosomal protein L30, conserved site / WD40-repeat-containing domain / Ribosome biogenesis factor, NIP7 / Ribosomal RNA large subunit methyltransferase E / Ribosomal protein L1, 3-layer alpha/beta-sandwich / Ribosomal protein L30, ferredoxin-like fold domain / Armadillo-type fold / Ribosomal protein L19/L19e, domain 1 / PUA-like superfamily / WD40/YVTN repeat-like-containing domain superfamily / 60S ribosomal protein L4, C-terminal domain / P-loop containing nucleoside triphosphate hydrolase / Nop2p / Ribosomal protein L13 / Ribosomal protein L27e superfamily / Ribosomal protein L35A superfamily / RNA recognition motif. (a.k.a. RRM, RBD, or RNP domain) / Ribosomal protein L22p/L17e / Ribosomal protein L14p/L23e / DEAD/DEAH box helicase / Helicase conserved C-terminal domain / Ribosomal protein L23 / Ribosomal protein L3 / Ribosomal protein L30p/L7e / Ribosomal protein L6 / WD domain, G-beta repeat / Ribosomal protein L10 / KOW motif / Ribosomal protein L4/L1 family / Ribosomal protein L34Ae superfamily / Ribosomal protein L34e / Ribosomal protein L13e / Ribosomal protein L19e / Ribosomal protein L7Ae/L30e/S12e/Gadd45 family / Ribosomal protein L35Ae / Ribosomal protein L24e / Ribosomal protein S8e / Ribosomal protein L31e / Ribosomal protein L1p/L10e family / 16S rRNA methyltransferase RsmB/F / Ribosomal protein L6e / Ribosomal protein L36e
Function and homology information
SourceSaccharomyces cerevisiae (baker's yeast)
Methodsingle particle reconstruction / cryo EM / 4.2 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: 6em5

SummaryFull reportAbout validation report
DateDeposition: Sep 29, 2017 / Header (metadata) release: Nov 15, 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: 0.025
  • Imaged by UCSF Chimera
  • Download
  • Surface view colored by radius
  • Surface level: 0.025
  • Imaged by UCSF Chimera
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  • Surface view with fitted model
  • Atomic models: : PDB-6em5
  • Surface level: 0.025
  • Imaged by UCSF Chimera
  • Download
Movie viewer
Structure viewerEM map:
SurfViewMolmilJmol/JSmol
Supplemental images

Downloads & links

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Map

Fileemd_3890.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:0.025 (by author), 0.025 (movie #1):
Minimum - Maximum-0.041138835 - 0.12395228
Average (Standard dev.)0.00065085466 (0.007468635)
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-0.0410.1240.001

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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: 34453
3D reconstructionAlgorithm: FOURIER SPACE / Software: RELION / Resolution: 4.2 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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