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- EMDB-1302: RF3 induces ribosomal conformational changes responsible for diss... -

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Basic information

Entry
Database: EMDB / ID: EMD-1302
TitleRF3 induces ribosomal conformational changes responsible for dissociation of class I release factors.
Map dataThis is the map of a ribosome.
Sample
  • Sample: E.coli 70s ribosomal release complex bound with RF3
  • Complex: 70s
  • Protein or peptide: RF3
  • RNA: tRNATransfer RNA
Function / homology
Function and homology information


regulation of translational termination / translation release factor activity, codon nonspecific / translation release factor activity, codon specific / guanosine tetraphosphate binding / translational termination / maintenance of translational fidelity / GDP binding / GTPase activity / GTP binding / cytosol / cytoplasm
Similarity search - Function
Peptide chain release factor 3, C-terminal / Peptide chain release factor 3, domain III superfamily / Peptide chain release factor 3, GTP-binding domain / Class II release factor RF3, C-terminal domain / Peptide chain release factor 3 / EF-G domain III/V-like / Tr-type G domain, conserved site / Translational (tr)-type guanine nucleotide-binding (G) domain signature. / Translation elongation factor EFTu-like, domain 2 / Elongation factor Tu domain 2 ...Peptide chain release factor 3, C-terminal / Peptide chain release factor 3, domain III superfamily / Peptide chain release factor 3, GTP-binding domain / Class II release factor RF3, C-terminal domain / Peptide chain release factor 3 / EF-G domain III/V-like / Tr-type G domain, conserved site / Translational (tr)-type guanine nucleotide-binding (G) domain signature. / Translation elongation factor EFTu-like, domain 2 / Elongation factor Tu domain 2 / Translational (tr)-type GTP-binding domain / Elongation factor Tu GTP binding domain / Translational (tr)-type guanine nucleotide-binding (G) domain profile. / Small GTP-binding protein domain / Translation protein, beta-barrel domain superfamily / P-loop containing nucleoside triphosphate hydrolase
Similarity search - Domain/homology
Peptide chain release factor RF3 / Peptide chain release factor 3
Similarity search - Component
Biological speciesEscherichia coli (E. coli)
Methodsingle particle reconstruction / cryo EM / negative staining / Resolution: 15.5 Å
AuthorsGao H / Zhou Z / Rawat U / Huang C / Bouakaz L / Wang C / Liu Y / Zavialov A / Gursky R / Sanyal S ...Gao H / Zhou Z / Rawat U / Huang C / Bouakaz L / Wang C / Liu Y / Zavialov A / Gursky R / Sanyal S / Ehrenberg M / Frank J / Song H
CitationJournal: Cell / Year: 2007
Title: RF3 induces ribosomal conformational changes responsible for dissociation of class I release factors.
Authors: Haixiao Gao / Zhihong Zhou / Urmila Rawat / Chenhui Huang / Lamine Bouakaz / Chernhoe Wang / Zhihong Cheng / Yuying Liu / Andrey Zavialov / Richard Gursky / Suparna Sanyal / Måns Ehrenberg ...Authors: Haixiao Gao / Zhihong Zhou / Urmila Rawat / Chenhui Huang / Lamine Bouakaz / Chernhoe Wang / Zhihong Cheng / Yuying Liu / Andrey Zavialov / Richard Gursky / Suparna Sanyal / Måns Ehrenberg / Joachim Frank / Haiwei Song /
Abstract: During translation termination, class II release factor RF3 binds to the ribosome to promote rapid dissociation of a class I release factor (RF) in a GTP-dependent manner. We present the crystal ...During translation termination, class II release factor RF3 binds to the ribosome to promote rapid dissociation of a class I release factor (RF) in a GTP-dependent manner. We present the crystal structure of E. coli RF3*GDP, which has a three-domain architecture strikingly similar to the structure of EF-Tu*GTP. Biochemical data on RF3 mutants show that a surface region involving domains II and III is important for distinct steps in the action cycle of RF3. Furthermore, we present a cryo-electron microscopy (cryo-EM) structure of the posttermination ribosome bound with RF3 in the GTP form. Our data show that RF3*GTP binding induces large conformational changes in the ribosome, which break the interactions of the class I RF with both the decoding center and the GTPase-associated center of the ribosome, apparently leading to the release of the class I RF.
History
DepositionNov 29, 2006-
Header (metadata) releaseNov 29, 2006-
Map releaseJun 22, 2007-
UpdateJun 29, 2016-
Current statusJun 29, 2016Processing site: PDBe / Status: Released

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Structure visualization

Movie
  • Surface view with section colored by density value
  • Surface level: 25
  • Imaged by UCSF Chimera
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  • Surface view colored by height
  • Surface level: 25
  • Imaged by UCSF Chimera
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  • Surface view with fitted model
  • Atomic models: PDB-2o0f, PDB-3dg5
  • Surface level: 25
  • Imaged by UCSF Chimera
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  • Simplified surface model + fitted atomic model
  • Atomic modelsPDB-2o0f
  • Imaged by Jmol
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Movie viewer
Structure viewerEM map:
SurfViewMolmilJmol/JSmol
Supplemental images

1302.gif

Downloads & links

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Map

FileDownload / File: emd_1302.map.gz / Format: CCP4 / Size: 8.2 MB / Type: IMAGE STORED AS FLOATING POINT NUMBER (4 BYTES)
AnnotationThis is the map of a ribosome.
Voxel sizeX=Y=Z: 2.82 Å
Density
Contour Level1: 57.200000000000003 / Movie #1: 25
Minimum - Maximum-80.433800000000005 - 242.787000000000006
Average (Standard dev.)5.95889 (±23.236000000000001)
SymmetrySpace group: 1
Details

EMDB XML:

Map geometry
Axis orderXYZ
Origin-65-65-65
Dimensions130130130
Spacing130130130
CellA=B=C: 366.6 Å
α=β=γ: 90 °

CCP4 map header:

modeImage stored as Reals
Å/pix. X/Y/Z2.822.822.82
M x/y/z130130130
origin x/y/z0.0000.0000.000
length x/y/z366.600366.600366.600
α/β/γ90.00090.00090.000
MAP C/R/S123
start NC/NR/NS-65-65-65
NC/NR/NS130130130
D min/max/mean-80.434242.7875.959

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Supplemental data

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Sample components

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Entire : E.coli 70s ribosomal release complex bound with RF3

EntireName: E.coli 70s ribosomal release complex bound with RF3
Components
  • Sample: E.coli 70s ribosomal release complex bound with RF3
  • Complex: 70s
  • Protein or peptide: RF3
  • RNA: tRNATransfer RNA

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Supramolecule #1000: E.coli 70s ribosomal release complex bound with RF3

SupramoleculeName: E.coli 70s ribosomal release complex bound with RF3 / type: sample / ID: 1000 / Number unique components: 3

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Supramolecule #1: 70s

SupramoleculeName: 70s / type: complex / ID: 1 / Details: E.coli / Ribosome-details: ribosome-prokaryote: ALL

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Macromolecule #1: RF3

MacromoleculeName: RF3 / type: protein_or_peptide / ID: 1 / Details: E.coli / Number of copies: 1 / Oligomeric state: Monomer / Recombinant expression: No
Source (natural)Organism: Escherichia coli (E. coli) / synonym: E.coli

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Macromolecule #2: tRNA

MacromoleculeName: tRNA / type: rna / ID: 2 / Details: Hybrid P/E / Classification: OTHER / Structure: OTHER / Synthetic?: No
Source (natural)Organism: Escherichia coli (E. coli) / synonym: E.coli

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Experimental details

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Structure determination

Methodnegative staining, cryo EM
Processingsingle particle reconstruction
Aggregation stateparticle

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Sample preparation

BufferpH: 7.5 / Details: Polymix
StainingType: NEGATIVE / Details: Cryo
GridDetails: Quanti-foil grids coated with a thin carbon layer
VitrificationCryogen name: ETHANE / Instrument: OTHER
Details: Vitrification instrument: Vitrobot. Rapid-freezing in liquid ethane

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Electron microscopy

MicroscopeFEI TECNAI F20
Electron beamAcceleration voltage: 200 kV / Electron source: FIELD EMISSION GUN
Electron opticsCalibrated magnification: 49696 / Illumination mode: FLOOD BEAM / Imaging mode: BRIGHT FIELDBright-field microscopy / Cs: 2.0 mm / Nominal defocus max: 4.0 µm / Nominal defocus min: 2.0 µm / Nominal magnification: 50000
Sample stageSpecimen holder: cryo transfer / Specimen holder model: GATAN LIQUID NITROGEN
TemperatureAverage: 93 K
Alignment procedureLegacy - Astigmatism: objective lens astigmatism was corrected at 100,000 times magnification
Legacy - Electron beam tilt params: 0
DateJan 30, 2002
Image recordingCategory: FILM / Film or detector model: KODAK SO-163 FILM / Digitization - Scanner: ZEISS SCAI / Digitization - Sampling interval: 14 µm / Average electron dose: 20 e/Å2
Tilt angle min0
Tilt angle max0
Experimental equipment
Model: Tecnai F20 / Image courtesy: FEI Company

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Image processing

CTF correctionDetails: CTF correctionn of 3D map
Final reconstructionApplied symmetry - Point group: C1 (asymmetric) / Algorithm: OTHER / Resolution.type: BY AUTHOR / Resolution: 15.5 Å / Resolution method: FSC 0.5 CUT-OFF / Software - Name: SPIDER package / Number images used: 45000

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Atomic model buiding 1

Initial modelPDB ID:

2avy
PDB Unreleased entry


Chain - Chain ID: A
SoftwareName: RSRef, TNT
DetailsProtocol: Rigid Body. Real-space refinement using RSRef.
RefinementSpace: REAL / Protocol: RIGID BODY FIT / Target criteria: cross correlation coefficient
Output model

PDB-2o0f:
Docking of the modified RF3 X-ray structure into cryo-EM map of E.coli 70S ribosome bound with RF3

PDB-3dg5:
Coordinates of 16S and 23S rRNAs fitted into the cryo-EM map of RF3-bound termination complex

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Atomic model buiding 2

Initial modelPDB ID:

2aw4
PDB Unreleased entry


Chain - Chain ID: A
SoftwareName: RSRef, TNT
DetailsProtocol: Rigid Body. Real-space refinement using RSRef.
RefinementSpace: REAL / Protocol: RIGID BODY FIT / Target criteria: cross correlation coefficient
Output model

PDB-2o0f:
Docking of the modified RF3 X-ray structure into cryo-EM map of E.coli 70S ribosome bound with RF3

PDB-3dg5:
Coordinates of 16S and 23S rRNAs fitted into the cryo-EM map of RF3-bound termination complex

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