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Title:3D cryo-EM Structure of Archaeal 20S Proteasome in Complex with the C-terminus of PAN
Authors:Yu Y, Smith DM, Kim H, Rodriguez V, Goldberg AL, Cheng Y
Sample:Archaeal 20S proteasome in complex with a hybrid of PA26 and PAN
Method:Single particle reconstruction (7.5 angstroms resolution)
Red flagLatest update:2011-09-23
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Sample
Sample name: Archaeal 20S proteasome in complex with a hybrid of PA26 and PAN
Oligomeric state: 20S proteasome is composed of 4 heptamer rings coaxially stacked to which heptamers of the PA26 PAN hybrid cap on both ends
Theoretical molecular weight of the sample: 1.1
Components:
ID Type Name Exp. MW (MDa) Oligomeric details Recombinant expression Synthetic Organism UniProt identifier GO identifier InterPro identifier Virus identifier Details
1protein20S proteasome, PA26-PAN2 heptamers of 20S alpha subunit, 2 heptamers of 20S beta subunit, and 2 heptamers of PA26-PANtrueThermoplasma acidophilum
Experiment
Specimen state: Particle
Specimen preparation:
pHSpecimen conc.DetailsStainingSpecimen support details
8.50.3 mg/mL50 mM Tris pH 8.5, 1mM DTT, 10 mM MgCl2, 5% Glycerol400 mesh Cu grids
Vitrification:
Cryogen nameHumidityTemp.Instr.MethodTime resolvedDetails
ETHANE100%90 KFEI VITROBOTblot for 4 seconds before plunging msVitrification instrument: Vitrobot
Imaging:
MicroscopeVoltageIllumination modeImaging modeCsDefocus min.Defocus max.Nominal mag.Calibrated mag.Electron sourceDetectorDetector distanceAstigmatism
FEI TECNAI F20200 kVFLOOD BEAMBRIGHT FIELD2.0 mm1000 nm5000 nm62000FIELD EMISSION GUNGENERIC GATAN (4k x 4k) mm

Specimen holderHolder modelTilt min.Tilt max.Energy filterEnergy windowTemp.Temp. min.Temp. max.Beam tiltElectron doseOther detailsDate
Gatan CT-3500GATAN LIQUID NITROGEN eV90 K90 K90 K mrad20 e/Å2data was collected using Leginon17-Sep-2007
Processing
Protocol:Fourier space 3D reconstruction
Software:FREALIGN
CTF correction:each particle
Number of particles:13020
Imposed symmetry:D7
Resolution by author:7.5 Å
Resolution method:FSC 0.143
Processing details:D7 symmetry was applied. The particles were manually selected.
Scanned images:
Num. imagesSampling sizeOD rangeQuant. bit numberOther detailsScanner
μm/pixelimages were recorded on CCD camera
Fitting:
PDBProtocolTarget crit.SoftwareB valueFitting spacePDB chainDetails