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Title:Electron cryo-microscopy of microtubule-bound human kinesin-5 motor domain in AMPPNP state.
Authors:Goulet A, Behnke-Parks WM, Sindelar C, Rosenfeld S, Moores C
Sample:13-protofilament microtubule-bound human kinesin-5 motor domain with AMPPNP
Method:Single particle reconstruction (9.7 angstroms resolution)
Red flagLatest update:2013-01-09
Sample name: 13-protofilament microtubule-bound human kinesin-5 motor domain with AMPPNP
Oligomeric state: 13-protofilament microtubule with one kineisn-5 motor domain bound every tubulin heterodimers
ID Type Name Exp. MW (MDa) Theo. MW (MDa) Oligomeric details Recombinant expression Synthetic Organism UniProt identifier GO identifier InterPro identifier Virus identifier Details
1proteinalpha-beta tubulin dimerheterodimerfalseBos taurus
2proteinKinesin-5 motor domainmonomertrueHomo sapiens
Specimen state: Particle
Specimen preparation:
pHSpecimen conc.DetailsStainingSpecimen support details
6.8 mg/mL80 mM PIPES, 5 mM MgCl2, 1 mM EGTA, 5mM AMPPNP400 mesh holey carbon grids
Cryogen nameHumidityTemp.Instr.MethodTime resolvedDetails
ETHANE100% KFEI VITROBOT MARK Ichamber at 24 degrees C, blot 2.5 sec ms
MicroscopeVoltageIllumination modeImaging modeCsDefocus min.Defocus max.Nominal mag.Calibrated mag.Electron sourceDetectorDetector distanceAstigmatism
FEI TECNAI F20200 kVFLOOD BEAMBRIGHT FIELD2.0 mm700 nm2200 nm50000FIELD EMISSION GUNKODAK SO-163 FILM mmObjective lens astigmatism was corrected at 150,000 times magnification

Specimen holderHolder modelTilt min.Tilt max.Energy filterEnergy windowTemp.Temp. min.Temp. max.Beam tiltElectron doseOther detailsDate
GATAN LIQUID NITROGEN°° eV90 K K K mrad18 e/Å210-JAN-2011
CTF correction:FREALIGN
Number of particles:3587
Resolution by author:9.7 Å
Resolution method:FSC 0.5
Processing details:Approximately 50,000 asymmetric units were averaged in the final reconstruction. The particles were selected along individual microtubules.
Scanned images:
Num. imagesSampling sizeOD rangeQuant. bit numberOther detailsScanner
467 μm/pixel8ZEISS SCAI
PDBProtocolTarget crit.SoftwareB valueFitting spacePDB chainDetails
3HQD flexiblecross-correlationChimera, FlexEMREALProtocol: rigid body then flexible fitting. The domain was fitted as a rigid body. The N-terminal residues 6 to 16 were built in the EM map and the final model was refined by flexible fitting.
1JFF rigid bodycross-correlationChimeraREALProtocol: rigid body. alpha- and b-tubulin were separately fitted.