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Title:Visualizing tmRNA entry into a stalled ribosome.
Authors:M.Valle, R.Gillet, S.Kaur, A.Henne, V.Ramakrishnan, and J.Frank
Sample:70S-Psite tRNA - mRNA - EF-Tu.GDP - alanylated tmRNA - SmpB - Kirromycin - S1
Aggregation state:Single particle (13 angstroms resolution)
Red flagLatest update:2011-05-26
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bullet_white.gif  Summary
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bullet.png Experimental details
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bullet_white.gif  Visualization
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Sample
Sample name: 70S-Psite tRNA - mRNA - EF-Tu.GDP - alanylated tmRNA - SmpB - Kirromycin - S1
Components:
ID Type Name Exp. MW (MDa) Oligomeric details Recombinant expression Synthetic Organism GO identifier InterPro identifier Virus identifier Details
1ribosome-prokaryote70S RibosomeThermus thermophilus
2nucleic-acidtRNAfalseP site
3proteinEF-Tufalseunidentified
4ligandtmRNAfalse
5proteinSmpBfalseunidentified
Experiment
Sample preparation:
pHSample conc.DetailsStainingSample support details
7.5 mg/mL25 mM Hepes-KOH pH 7.5, 30 mM NH4Cl, 7mM MgCl2, 2 mM ATP, 6 mM PEP, 10g/ml PK
Vitrification:
Cryogen nameHumidityTemp.Instr.MethodTime resolvedDetails
ETHANE90%93 KHOMEMADE PLUNGERBlot for 2 seconds before plunging msVitrification instrument: two side blotting plunger
Imaging:
MicroscopeVoltageIllumination modeImaging modeCs:Defocus min.Defocus max.Nominal mag.Calibrated mag.Electron sourceDetectorDetector distanceAstigmatism
FEI TECNAI F20200 kVFLOOD BEAMBRIGHT FIELD2.0 mm1450 nm3700 nm4900052000FIELD EMISSION GUNKodak SO163 film mm

Specimen holderHolder modelTilt min.Tilt max.Energy filterEnergy windowTemp.Temp. min.Temp. max.Beam tiltElectron doseOther detailsDate
Cryo transferGATAN LIQUID NITROGEN°° eV93 K K K mrad15 e/Å22002-8-1
Processing
Software:SPIDER
CTF correction:defocus groups
Resolution by author:13 Å
Resolution method:FSC at 0.15 cut-off
Unit cell:
Scanned images:
Num. imagesSampling sizeOD rangeQuant. bit numberOther detailsScanner
4414 μm/pixel1.212linkZEISS SCAI
Fitting:
PDBProtocolTarget crit.SoftwareB valueFitting spacePDB chainDetails
Rigid Bodycross correlation coefficientmanualThe domains were separately fitted by manual docking using program O
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