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Title:Cryo EM map of the Gaalphaq-PLCbeta3 complex.
Authors:Dutta S, Lyon AM, Tesmer JJG, Skiniotis G
Sample:Reconstruction of the PLCbeta3 distal CTD in complex with the N-terminal helix of Galphaq
Method:Single particle reconstruction (19 angstroms resolution)
Red flagLatest update:2013-04-17
Sample
Sample name: Reconstruction of the PLCbeta3 distal CTD in complex with the N-terminal helix of Galphaq
Oligomeric state: Galphaq-PLCb3 dimer
Experimental molecular weight of the sample: 1.8
Components:
ID Type Name Exp. MW (MDa) Theo. MW (MDa) Oligomeric details Recombinant expression Synthetic Organism UniProt identifier GO identifier InterPro identifier Virus identifier Details
1proteinG alpha q0.0440.044monomertrueMus musculusGO:0007202IPR000654
2proteinphospholipase C beta 30.1390.139monomertrueHomo sapiensGO:0007186IPR016280
Experiment
Specimen state: Particle
Specimen preparation:
pHSpecimen conc.DetailsStainingSpecimen support details
8.01 mg/mL20 mM HEPES, pH 8, 200 mM NaCl, 2 mM DTT, 0.9 mM CaCl2, 50 uM GDP, 30 uM AlCl3, 10 mM NaF, 5 mM MgCl2glow-discharged Quantifoil R2/200 mesh grid
Vitrification:
Cryogen nameHumidityTemp.Instr.MethodTime resolvedDetails
ETHANE100% KFEI VITROBOT MARK IVBlot 3 microliter sample for 2 seconds. msVitrification carried out in liquid nitrogen atmosphere.
Imaging:
MicroscopeVoltageIllumination modeImaging modeCsDefocus min.Defocus max.Nominal mag.Calibrated mag.Electron sourceDetectorDetector distanceAstigmatism
FEI TECNAI F20120 kVFLOOD BEAMBRIGHT FIELD2 mm1500 nm3500 nm5000071138FIELD EMISSION GUNGATAN ULTRASCAN 4000 (4k x 4k) mmObjective lens astigmatism was corrected at 135,000 times magnification.

Specimen holderHolder modelTilt min.Tilt max.Energy filterEnergy windowTemp.Temp. min.Temp. max.Beam tiltElectron doseOther detailsDate
Used two holders.OTHER°° eV89 K89 K95 K mrad20 e/Å220-OCT-2011
Processing
Protocol:cross-common lines
Software:EMAN1
CTF correction:each particle
Number of particles:12692
Imposed symmetry:C1
Resolution by author:19 Å
Resolution method:FSC 0.5
Scanned images:
Num. imagesSampling sizeOD rangeQuant. bit numberOther detailsScanner
350 μm/pixel
Fitting:
PDBProtocolTarget crit.SoftwareB valueFitting spacePDB chainDetails
4GNK REAL