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- EMDB-5188: tmRNA-SmpB: a journey to the center of the bacterial ribosome -

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Basic information

Entry
Database: EMDB / ID: EMD-5188
TitletmRNA-SmpB: a journey to the center of the bacterial ribosome
Map dataStructure of tmRNA-SMPB complex accomodated into a stalled bacterial ribosome
Sample
  • Sample: Thermus thermophilus 70S ribosome
  • Complex: 70S ribosomeRibosome
KeywordsTMRNA / SMPB / RIBOSOME / TRANS-TRANSLATION
Function / homologytrans-translation / SsrA-binding protein / SsrA-binding protein, conserved site / Small protein B / SmpB protein / SsrA-binding protein. / rRNA binding / cytoplasm / SsrA-binding protein
Function and homology information
Biological speciesThermus thermophilus (bacteria)
Methodsingle particle reconstruction / cryo EM / Resolution: 13.0 Å
AuthorsWeis F / Bron P / Giudice E / Rolland JP / Thomas D / Felden B / Gillet R
CitationJournal: EMBO J / Year: 2010
Title: tmRNA-SmpB: a journey to the centre of the bacterial ribosome.
Authors: Félix Weis / Patrick Bron / Emmanuel Giudice / Jean-Paul Rolland / Daniel Thomas / Brice Felden / Reynald Gillet /
Abstract: Ribosomes mediate protein synthesis by decoding the information carried by messenger RNAs (mRNAs) and catalysing peptide bond formation between amino acids. When bacterial ribosomes stall on ...Ribosomes mediate protein synthesis by decoding the information carried by messenger RNAs (mRNAs) and catalysing peptide bond formation between amino acids. When bacterial ribosomes stall on incomplete messages, the trans-translation quality control mechanism is activated by the transfer-messenger RNA bound to small protein B (tmRNA-SmpB ribonucleoprotein complex). Trans-translation liberates the stalled ribosomes and triggers degradation of the incomplete proteins. Here, we present the cryo-electron microscopy structures of tmRNA-SmpB accommodated or translocated into stalled ribosomes. Two atomic models for each state are proposed. This study reveals how tmRNA-SmpB crosses the ribosome and how, as the problematic mRNA is ejected, the tmRNA resume codon is placed onto the ribosomal decoding site by new contacts between SmpB and the nucleotides upstream of the tag-encoding sequence. This provides a structural basis for the transit of the large tmRNA-SmpB complex through the ribosome and for the means by which the tmRNA internal frame is set for translation to resume.
History
DepositionApr 16, 2010-
Header (metadata) releaseAug 23, 2010-
Map releaseOct 13, 2010-
UpdateMar 6, 2013-
Current statusMar 6, 2013Processing site: RCSB / Status: Released

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Structure visualization

Movie
  • Surface view with section colored by density value
  • Surface level: 0.01
  • Imaged by UCSF Chimera
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  • Surface view colored by height
  • Surface level: 0.01
  • Imaged by UCSF Chimera
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  • Surface view with fitted model
  • Atomic models: PDB-3iyq
  • Surface level: 0.01
  • Imaged by UCSF Chimera
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  • Simplified surface model + fitted atomic model
  • Atomic modelsPDB-3iyq
  • 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_5188.map.gz / Format: CCP4 / Size: 7.8 MB / Type: IMAGE STORED AS FLOATING POINT NUMBER (4 BYTES)
AnnotationStructure of tmRNA-SMPB complex accomodated into a stalled bacterial ribosome
Voxel sizeX=Y=Z: 3.28 Å
Density
Contour LevelBy AUTHOR: 0.01 / Movie #1: 0.01
Minimum - Maximum-0.05623283 - 0.12125266
Average (Standard dev.)0.00177611 (±0.01004907)
SymmetrySpace group: 1
Details

EMDB XML:

Map geometry
Axis orderXYZ
Origin-64-64-64
Dimensions128128128
Spacing128128128
CellA=B=C: 419.84 Å
α=β=γ: 90.0 °

CCP4 map header:

modeImage stored as Reals
Å/pix. X/Y/Z3.283.283.28
M x/y/z128128128
origin x/y/z0.0000.0000.000
length x/y/z419.840419.840419.840
α/β/γ90.00090.00090.000
start NX/NY/NZ-34-26-72
NX/NY/NZ6953145
MAP C/R/S123
start NC/NR/NS-64-64-64
NC/NR/NS128128128
D min/max/mean-0.0560.1210.002

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Supplemental data

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Sample components

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Entire : Thermus thermophilus 70S ribosome

EntireName: Thermus thermophilus 70S ribosome
Components
  • Sample: Thermus thermophilus 70S ribosome
  • Complex: 70S ribosomeRibosome

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Supramolecule #1000: Thermus thermophilus 70S ribosome

SupramoleculeName: Thermus thermophilus 70S ribosome / type: sample / ID: 1000 / Number unique components: 5
Molecular weightTheoretical: 2.3 MDa

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Supramolecule #1: 70S ribosome

SupramoleculeName: 70S ribosome / type: complex / ID: 1 / Recombinant expression: No / Database: NCBI / Ribosome-details: ribosome-prokaryote: ALL
Source (natural)Organism: Thermus thermophilus (bacteria)
Molecular weightTheoretical: 2.3 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

BufferpH: 7.5
Details: 5mM Hepes-KOH (pH 7.5), 10mM NH4Cl, 10mM MgOAc, 50mM KCl, 0.1mM EDTA and 6mM BetaME
GridDetails: Quantifoil holey-carbon grids previously glow-discharged
VitrificationCryogen name: ETHANE / Chamber humidity: 100 % / Chamber temperature: 100 K / Instrument: OTHER / Details: Vitrification instrument: vitrobot / Method: Blot for 5 seconds before plunging

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Electron microscopy

MicroscopeJEOL 2200FS
Electron beamAcceleration voltage: 200 kV / Electron source: FIELD EMISSION GUN
Electron opticsCalibrated magnification: 45700 / Illumination mode: FLOOD BEAM / Imaging mode: BRIGHT FIELDBright-field microscopy / Cs: 2 mm / Nominal defocus max: 2.2 µm / Nominal defocus min: 0.9 µm / Nominal magnification: 50000
Specialist opticsEnergy filter - Name: JEOL / Energy filter - Lower energy threshold: 0.0 eV / Energy filter - Upper energy threshold: 20.0 eV
Sample stageSpecimen holder: Eucentric / Specimen holder model: GATAN LIQUID NITROGEN
TemperatureAverage: 95 K
DateJul 1, 2009
Image recordingCategory: FILM / Film or detector model: KODAK SO-163 FILM / Digitization - Scanner: NIKON SUPER COOLSCAN 9000 / Digitization - Sampling interval: 7.5 µm / Number real images: 70 / Bits/pixel: 8

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Image processing

CTF correctionDetails: Each particle
Final reconstructionResolution.type: BY AUTHOR / Resolution: 13.0 Å / Resolution method: FSC 0.5 CUT-OFF / Software - Name: IMAGIC-V / Number images used: 49061

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Atomic model buiding 1

Initial modelPDB ID:

Chain - #0 - Chain ID: A / Chain - #1 - Chain ID: B
SoftwareName: UCSF-Chimera, mdff
DetailsProtocol: Rigid Body and flexible fitting. The domains were initially separately fitted by manual docking and then further optimized using mdff.
RefinementSpace: REAL / Protocol: FLEXIBLE FIT / Target criteria: Cross-Correlation
Output model

PDB-3iyq:
tmRNA-SmpB: a journey to the center of the bacterial ribosome

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Atomic model buiding 2

Initial modelPDB ID:

Chain - #0 - Chain ID: A / Chain - #1 - Chain ID: B
SoftwareName: UCSF-Chimera, mdff
DetailsProtocol: Rigid Body and flexible fitting. each subdomain (H2b-c, H5, pk1, pk2, pk3, pk4) were initially separately fitted by manual docking and then further optimized using mdff.
RefinementSpace: REAL / Protocol: FLEXIBLE FIT / Target criteria: Cross-Correlation
Output model

PDB-3iyq:
tmRNA-SmpB: a journey to the center of the bacterial ribosome

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