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Title:Ribosome structures to near-atomic resolution from thirty thousand cryo-EM particles
Authors:Bai XC, Fernandez IS, McMullan G, Scheres SHW
Sample:T. thermophilus 70S ribosome
Method:Single particle reconstruction (5.1 angstroms resolution)
Red flagLatest update:2013-07-03
Sample
Sample name: T. thermophilus 70S ribosome
Theoretical molecular weight of the sample: 3
Components:
ID Type Name Exp. MW (MDa) Oligomeric details Recombinant expression Synthetic Organism UniProt identifier GO identifier InterPro identifier Virus identifier Details
1ribosome-prokaryote70S ribosome3falseThermus thermophilus
Experiment
Specimen state: Particle
Specimen preparation:
pHSpecimen conc.DetailsStainingSpecimen support details
7.450.2 mg/mL3mM Hepes-KOH, 6.6 mM Tris-acetate pH 7.2, 3 mM NH4Cl, 6.6 mM NH4-acetate, 48 mM K-acetate, 4 mM Mg-acetate, 2.4 mM DTTQuantifoil grids (2/2) with 3 nm thin carbon on top
Vitrification:
Cryogen nameHumidityTemp.Instr.MethodTime resolvedDetails
ETHANE100%90 KFEI VITROBOT MARK IIBlot 2.5 seconds before plunging ms
Imaging:
MicroscopeVoltageIllumination modeImaging modeCsDefocus min.Defocus max.Nominal mag.Calibrated mag.Electron sourceDetectorDetector distanceAstigmatism
FEI POLARA 300300 kVFLOOD BEAMBRIGHT FIELD2 mm1178 nm3597 nm5900079096FIELD EMISSION GUNFEI FALCON I (4k x 4k) mmObjective lens astigmatism was corrected at 59,000 times magnification

Specimen holderHolder modelTilt min.Tilt max.Energy filterEnergy windowTemp.Temp. min.Temp. max.Beam tiltElectron doseOther detailsDate
GATAN LIQUID NITROGEN°° eV85 K80 K90 K mrad16 e/Å201-JUL-2012
Processing
Software:CTFFIND3, RELION
CTF correction:each image
Number of particles:35404
Resolution by author:5.1 Å
Resolution method:FSC 0.143, gold-standard
Processing details:Use a newly developed statistical movie processing approach to compensate for beam-induced movement. Use a newly developed statistical movie processing to compensate for beam-induced movement.
Scanned images:
Num. imagesSampling sizeOD rangeQuant. bit numberOther detailsScanner
36014 μm/pixelEvery image is the average of sixteen frames recorded by the direct electron detector
Fitting:
PDBProtocolTarget crit.SoftwareB valueFitting spacePDB chainDetails
2WH1 rigid bodyChimeraREALProtocol: Rigid body. The domains were separately fitted by manual docking using program Chimera
2WH2 rigid bodyChimeraREALProtocol: Rigid body. The domains were separately fitted by manual docking using program Chimera