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- EMDB-8195: m48S late-stage initiation complex, purified from rabbit reticulo... -

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Basic information

Entry
Database: EMDB / ID: EMD-8195
Titlem48S late-stage initiation complex, purified from rabbit reticulocytes lysates, displaying eIF2 ternary complex and eIF3 i and g subunits relocated to the intersubunit face
Map data
Sample
  • Complex: m48S late-stage initiation complex, purified from rabbit reticulocytes lysates, displaying eIF2 ternary complex and eIF3 i and g subunits relocated to the intersubunit face
Function / homology
Function and homology information


viral translational termination-reinitiation / eukaryotic translation initiation factor 3 complex, eIF3m / IRES-dependent viral translational initiation / eukaryotic translation initiation factor 3 complex / eukaryotic 43S preinitiation complex / formation of cytoplasmic translation initiation complex / eukaryotic 48S preinitiation complex / Formation of the ternary complex, and subsequently, the 43S complex / regulation of translational initiation / Ribosomal scanning and start codon recognition ...viral translational termination-reinitiation / eukaryotic translation initiation factor 3 complex, eIF3m / IRES-dependent viral translational initiation / eukaryotic translation initiation factor 3 complex / eukaryotic 43S preinitiation complex / formation of cytoplasmic translation initiation complex / eukaryotic 48S preinitiation complex / Formation of the ternary complex, and subsequently, the 43S complex / regulation of translational initiation / Ribosomal scanning and start codon recognition / Translation initiation complex formation / Formation of a pool of free 40S subunits / GTP hydrolysis and joining of the 60S ribosomal subunit / L13a-mediated translational silencing of Ceruloplasmin expression / translation initiation factor binding / translational initiation / translation initiation factor activity / cytoplasmic stress granule / molecular adaptor activity / synapse / RNA binding / extracellular exosome / cytosol
Similarity search - Function
Eukaryotic translation initiation factor 3 subunit B / eIF3B, RNA recognition motif / Translation initiation factor, beta propellor-like domain / Eukaryotic translation initiation factor eIF2A / RNA recognition motif / RNA recognition motif / Eukaryotic RNA Recognition Motif (RRM) profile. / RNA recognition motif domain / RNA-binding domain superfamily / Nucleotide-binding alpha-beta plait domain superfamily / WD40/YVTN repeat-like-containing domain superfamily
Similarity search - Domain/homology
Eukaryotic translation initiation factor 3 subunit B
Similarity search - Component
Biological speciesOryctolagus cuniculus (rabbit)
Methodsingle particle reconstruction / cryo EM / Resolution: 5.8 Å
AuthorsSimonetti A / Brito Querido J / Myasnikov AG / Mancera-Martinez E / Renaud A / Kuhn L / Hashem Y
CitationJournal: Mol Cell / Year: 2016
Title: eIF3 Peripheral Subunits Rearrangement after mRNA Binding and Start-Codon Recognition.
Authors: Angelita Simonetti / Jailson Brito Querido / Alexander G Myasnikov / Eder Mancera-Martinez / Adeline Renaud / Lauriane Kuhn / Yaser Hashem /
Abstract: mRNA translation initiation in eukaryotes requires the cooperation of a dozen eukaryotic initiation factors (eIFs) forming several complexes, which leads to mRNA attachment to the small ribosomal ...mRNA translation initiation in eukaryotes requires the cooperation of a dozen eukaryotic initiation factors (eIFs) forming several complexes, which leads to mRNA attachment to the small ribosomal 40S subunit, mRNA scanning for start codon, and accommodation of initiator tRNA at the 40S P site. eIF3, composed of 13 subunits, 8 core (a, c, e, f, h, l, k, and m) and 5 peripheral (b, d, g, i, and j), plays a central role during this process. Here we report a cryo-electron microscopy structure of a mammalian 48S initiation complex at 5.8 Å resolution. It shows the relocation of subunits eIF3i and eIF3g to the 40S intersubunit face on the GTPase binding site, at a late stage in initiation. On the basis of a previous study, we demonstrate the relocation of eIF3b to the 40S intersubunit face, binding below the eIF2-Met-tRNAi(Met) ternary complex upon mRNA attachment. Our analysis reveals the deep rearrangement of eIF3 and unravels the molecular mechanism underlying eIF3 function in mRNA scanning and timing of ribosomal subunit joining.
History
DepositionMay 17, 2016-
Header (metadata) releaseJul 13, 2016-
Map releaseJul 13, 2016-
UpdateAug 2, 2017-
Current statusAug 2, 2017Processing site: PDBe / Status: Released

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Structure visualization

Movie
  • Surface view with section colored by density value
  • Surface level: 0.0143
  • Imaged by UCSF Chimera
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  • Surface view colored by height
  • Surface level: 0.0143
  • Imaged by UCSF Chimera
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  • Surface view with fitted model
  • Atomic models: PDB-5k1h
  • Surface level: 0.0143
  • Imaged by UCSF Chimera
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  • Simplified surface model + fitted atomic model
  • Atomic modelsPDB-5k1h
  • Imaged by Jmol
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Movie viewer
Structure viewerEM map:
SurfViewMolmilJmol/JSmol
Supplemental images

Downloads & links

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Map

FileDownload / File: emd_8195.map.gz / Format: CCP4 / Size: 30.5 MB / Type: IMAGE STORED AS FLOATING POINT NUMBER (4 BYTES)
Voxel sizeX=Y=Z: 2.2 Å
Density
Contour LevelBy AUTHOR: 0.0143 / Movie #1: 0.0143
Minimum - Maximum-0.050907068 - 0.17095527
Average (Standard dev.)0.0010064766 (±0.013673666)
SymmetrySpace group: 1
Details

EMDB XML:

Map geometry
Axis orderXYZ
Origin000
Dimensions200200200
Spacing200200200
CellA=B=C: 440.0 Å
α=β=γ: 90.0 °

CCP4 map header:

modeImage stored as Reals
Å/pix. X/Y/Z2.22.22.2
M x/y/z200200200
origin x/y/z0.0000.0000.000
length x/y/z440.000440.000440.000
α/β/γ90.00090.00090.000
MAP C/R/S123
start NC/NR/NS000
NC/NR/NS200200200
D min/max/mean-0.0510.1710.001

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Supplemental data

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Sample components

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Entire : m48S late-stage initiation complex, purified from rabbit reticulo...

EntireName: m48S late-stage initiation complex, purified from rabbit reticulocytes lysates, displaying eIF2 ternary complex and eIF3 i and g subunits relocated to the intersubunit face
Components
  • Complex: m48S late-stage initiation complex, purified from rabbit reticulocytes lysates, displaying eIF2 ternary complex and eIF3 i and g subunits relocated to the intersubunit face

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Supramolecule #1: m48S late-stage initiation complex, purified from rabbit reticulo...

SupramoleculeName: m48S late-stage initiation complex, purified from rabbit reticulocytes lysates, displaying eIF2 ternary complex and eIF3 i and g subunits relocated to the intersubunit face
type: complex / ID: 1 / Parent: 0 / Macromolecule list: #1-#42
Source (natural)Organism: Oryctolagus cuniculus (rabbit)
Molecular weightTheoretical: 2 MDa

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Experimental details

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Structure determination

Methodcryo EM
Processingsingle particle reconstruction
Aggregation stateparticle

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Sample preparation

Concentration0.5 mg/mL
BufferpH: 7.6
GridModel: Quantifoil / Material: COPPER / Mesh: 400 / Support film - Material: CARBON / Support film - topology: HOLEY / Pretreatment - Type: GLOW DISCHARGE / Pretreatment - Atmosphere: AIR
VitrificationCryogen name: ETHANE / Chamber humidity: 100 % / Chamber temperature: 277 K / Instrument: FEI VITROBOT MARK IV

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Electron microscopy

MicroscopeFEI TITAN KRIOS
Electron beamAcceleration voltage: 300 kV / Electron source: FIELD EMISSION GUN
Electron opticsC2 aperture diameter: 100.0 µm / Illumination mode: FLOOD BEAM / Imaging mode: BRIGHT FIELDBright-field microscopy / Cs: 0.01 mm / Nominal defocus max: 4.5 µm / Nominal defocus min: 0.8 µm / Nominal magnification: 59000
Sample stageSpecimen holder model: FEI TITAN KRIOS AUTOGRID HOLDER / Cooling holder cryogen: NITROGEN
Image recordingFilm or detector model: FEI FALCON II (4k x 4k) / Detector mode: INTEGRATING / Digitization - Sampling interval: 14.0 µm / Digitization - Frames/image: 2-8 / Number grids imaged: 1 / Number real images: 5700 / Average exposure time: 1.5 sec. / Average electron dose: 24.0 e/Å2
Experimental equipment
Model: Titan Krios / Image courtesy: FEI Company

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Image processing

CTF correctionSoftware - Name: CTFFIND (ver. 4)
Startup modelType of model: PDB ENTRY
PDB model - PDB ID:
Initial angle assignmentType: PROJECTION MATCHING / Software - Name: RELION (ver. 1.3)
Final angle assignmentType: PROJECTION MATCHING / Software - Name: RELION (ver. 1.3)
Final reconstructionNumber classes used: 10 / Algorithm: FOURIER SPACE / Resolution.type: BY AUTHOR / Resolution: 5.8 Å / Resolution method: FSC 0.143 CUT-OFF / Software - Name: RELION (ver. 1.3) / Number images used: 475000

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Atomic model buiding 1

Initial modelPDB ID:
RefinementProtocol: FLEXIBLE FIT
Output model

PDB-5k1h:
eIF3b relocated to the intersubunit face to interact with eIF1 and below the eIF2 ternary-complex. from the structure of a partial yeast 48S preinitiation complex in closed conformation.

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