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- EMDB-3891: State E (TAP-Flag-Ytm1 E80A) - Visualizing the assembly pathway o... -

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Database: EMDB / ID: 3891
TitleState E (TAP-Flag-Ytm1 E80A) - Visualizing the assembly pathway of nucleolar pre-60S ribosomes
Map dataState E (TAP-Flag-Ytm1 E80A) - Visualizing the assembly pathway of nucleolar pre-60S ribosmes
SampleLarge subunit biogenesis particle purified via TAP-Flag-Ytm1 E80A
Function/homology27S pre-rRNA (guanosine2922-2'-O)-methyltransferase / AdoMet-dependent rRNA methyltransferase, Spb1 / Nucleolar complex-associated protein 3, N-terminal / Ribosomal RNA methyltransferase, Spb1, C-terminal / Nucleolar complex-associated protein 3 / RNA (C5-cytosine) methyltransferase, NOP2 / ec:2.1.1.310: / Ribosomal RNA methyltransferase Spb1, domain of unknown function DUF3381 / Noc2p-Noc3p complex / Domain of unknown function (DUF3381) ...27S pre-rRNA (guanosine2922-2'-O)-methyltransferase / AdoMet-dependent rRNA methyltransferase, Spb1 / Nucleolar complex-associated protein 3, N-terminal / Ribosomal RNA methyltransferase, Spb1, C-terminal / Nucleolar complex-associated protein 3 / RNA (C5-cytosine) methyltransferase, NOP2 / ec:2.1.1.310: / Ribosomal RNA methyltransferase Spb1, domain of unknown function DUF3381 / Noc2p-Noc3p complex / Domain of unknown function (DUF3381) / Nucleolar complex-associated protein / Ribosome biogenesis factor, NIP7 / Spb1 C-terminal domain / CCAAT-binding factor / rRNA (guanine) methyltransferase activity / Type II pantothenate kinase, bacterial / Ribosome biogenesis factor NIP7-like / WD repeat WDR12/Ytm1 / Ribosomal RNA large subunit methyltransferase E / CBF/Mak21 family / rRNA (cytosine-C5-)-methyltransferase activity / rRNA (uridine-2'-O-)-methyltransferase activity / Guanine nucleotide-binding protein-like 3, N-terminal domain / Uncharacterised protein family UPF0642 / Nop2p / Ribosomal RNA small subunit methyltransferase F, N-terminal / Uncharacterised protein family (UPF0113) / Eukaryotic rRNA processing / NOL1/NOP2/sun family signature. / snoRNA release from pre-rRNA / GTP-binding protein, orthogonal bundle domain superfamily / Ribosome biogenesis protein Nop16 / Bacterial Fmu (Sun)/eukaryotic nucleolar NOL1/Nop2p, conserved site / GNL3L/Grn1 putative GTPase / Ribosome biogenesis protein BRX1 / Protein of unknown function (DUF2423) / Ribosome assembly factor Mrt4 / BOP1, N-terminal domain / Nucleolar GTP-binding protein 1 / NOG, C-terminal / WD repeat BOP1/Erb1 / Type II pantothenate kinase / Domain of unknown function DUF4217 / Nucleolar GTP-binding protein 1, Rossman-fold domain / nuclear division / Eukaryotic rRNA processing protein EBP2 / Pescadillo / NLE / Ribosome biogenesis protein 15, RNA recognition motif / 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 Nop16 / N-terminal domain of 16S rRNA methyltransferase RsmF / SAM-dependent methyltransferase RsmB/NOP2-type / RNA (C5-cytosine) methyltransferase / SAM-dependent MTase RsmB/NOP-type domain profile. / BOP1NT (NUC169) domain / Domain of unknown function (DUF4217) / NOGCT (NUC087) domain / PeBoW complex / Nucleolar GTP-binding protein 1 (NOG1) / Pescadillo N-terminus / rRNA primary transcript binding / NLE (NUC135) domain / Fumble / Circularly permuted (CP)-type guanine nucleotide-binding (G) domain profile. / Circularly permuted (CP)-type guanine nucleotide-binding (G) domain / Brix domain profile. / Brix domain / Translation initiation factor IF6 / OBG-type guanine nucleotide-binding (G) domain profile. / OBG-type guanine nucleotide-binding (G) domain / 16S rRNA methyltransferase RsmB/F / maturation of 5.8S rRNA from tricistronic rRNA transcript (SSU-rRNA, 5.8S rRNA, LSU-rRNA) / PUA domain superfamily / nuclear pre-replicative complex / pre-replicative complex assembly involved in nuclear cell cycle DNA replication / actomyosin contractile ring assembly / PUA domain profile. / PUA domain / eIF-6 family / Brix domain / mature ribosome assembly / regulation of cell division / pantothenate kinase / pantothenate kinase activity / rRNA base methylation / RNA-dependent ATPase activity / ribosomal subunit export from nucleus / BRCT domain, a BRCA1 C-terminus domain / nuclear-transcribed mRNA catabolic process / pre-mRNA 5'-splice site binding / cleavage in ITS2 between 5.8S rRNA and LSU-rRNA of tricistronic rRNA transcript (SSU-rRNA, 5.8S rRNA, LSU-rRNA) / GTP binding domain / ATPase activator activity / ribosomal large subunit binding / DEAD-box subfamily ATP-dependent helicases signature. / ribonucleoprotein complex assembly / preribosome, large subunit precursor / ATP-dependent RNA helicase DEAD-box, conserved site / Ribosomal protein L22e
Function and homology information
SourceSaccharomyces cerevisiae / / yeast /
MethodCryo EM / single particle reconstruction / 3.3 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: 5elz

SummaryFull report
PDB-ID: 6elz

SummaryFull report
About 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: 0.07
  • Imaged by UCSF CHIMERA
  • Download
  • Surface view colored by radius
  • Surface level: 0.07
  • Imaged by UCSF CHIMERA
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  • Surface view with fitted model
  • Atomic models: : PDB-6elz
  • Surface level: 0.07
  • Imaged by UCSF CHIMERA
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3D viewer
Supplemental images

Downloads & links

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Map

Fileemd_3891.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:0.07 (by author), 0.07 (movie #1):
Minimum - Maximum-0.2904615 - 0.5212299
Average (Standard dev.)-0.00013189367 (0.018902814)
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-0.2900.521-0.000

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Supplemental data

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Sample components

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Entire Large subunit biogenesis particle purified via TAP-Flag-Ytm1 E80A

EntireName: Large subunit biogenesis particle purified via TAP-Flag-Ytm1 E80A
Number of components: 1

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

ProteinName: Large subunit biogenesis particle purified via TAP-Flag-Ytm1 E80A
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: 112099
3D reconstructionAlgorithm: FOURIER SPACE / Software: RELION / Resolution: 3.3 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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