EMD-5896 Experiments and Validation

Cryo EM density of GDP-bound dynamic microtubules

Single particle reconstruction
Overview of EMD-5896
Sample name: Dynamic GDP-bound microtubule
Organisms: Sus scrofa, Homo sapiens
Fitted atomic model: 3j6f
Related EM entries by publication: EMD-5895, EMD-5897, EMD-5898, EMD-5899

Map parameters

Recommended contour level: 4.00 (emdb)
Number of grid points: 110 × 66 × 162
Voxel size: 1.74 × 1.74 × 1.74 Å
Minimum density: -4.672
Maximum density: 11.28
Average density: 0.528
Standard deviation: 1.926

Sample information

Sample name: Dynamic GDP-bound microtubule
Proteins: Alpha tubulin, Beta tubulin, kinesin

Validation information

Experimental information

 

Image processing

Applied symmetry: C1
Software: FREALIGN
Number of particles: 54445
Reconstruction protocol: projection matching
CTF correction: ctftilt

Fitting

Initial atomic model: 1JFF
Fitting software: ROSETTA
Fitting protocol: flexible
Refinement space: REAL
Details: Structure represents the minimized average structure of the 1% lowest energy structures from the refinement run.

Imaging

Session
Microscope model: FEI TITAN
Electron source: FIELD EMISSION GUN
Electron dose (e/A**2): 25.0
Nominal CS (mm): 2.70
Nominal magnification: 72000.0
Defocus max (nm): 3,500.00
Defocus min (nm): 1,400.00
Tilt max (degrees): 0.00
Tilt min (degrees): 0.00
Holder model: GATAN LIQUID NITROGEN
Acquisition
Number of images: 219
Scanner model: NIKON SUPER COOLSCAN 8000
Sampling interval (μm): 6.3499999999999996

Specimen

Preparation
Specimen state: helicalArray
Specimen concentration (mg/mL): 0.25
Buffer: 80mM PIPES, 1mM EGTA, 1mM MgCl2, 1mM DTT, 0.05% Nonidet P-40
Specimen support: 400 mesh C-flat 1.2/1.3, glow discharged in Edwards carbon evaporator
Aggregation parameters (helical parameters)
Handedness: LEFT HANDED
Phi increment (degrees): 25.760000000000002
Azimuthal (Z) increment (Å): 8.8200000000000003
Rotational symmetry: C1
Vitrification
Apparatus: FEI VITROBOT MARK II
Cryogen: ETHANE
Humidity: 90
Protocol: The grid was blotted for 2 seconds before plunging.