EMD-5410 Experiments and Validation

Cryo-electron microscopic reconstruction of wild rabbit hemorrhagic disease virus

Single particle reconstruction
Overview of EMD-5410
Sample name: Wild Rabbit Hemorrhagic Disease Virus (strain HYD)
Organism: Rabbit hemorrhagic disease virus
Fitted atomic model: 3j1p

Map parameters

Minimum density: -8.733
Maximum density: 16.738
Average density: 0.12
Standard deviation: 1.836
Recommended contour level: 2.60 (author)

Sample information

Sample name: Wild Rabbit Hemorrhagic Disease Virus (strain HYD)
Virus: Rabbit hemorrhagic disease virus

Validation information

Experimental information

 

Image processing

Applied symmetry: I
Software: EMAN1.9, Spider
Number of particles: 26000
Number of class averages: 475
Reconstruction protocol: Projection matching and Fourier reconstruction
CTF correction: CTF correction of each whole micrograph
Details: The final map was sharpened to 4.5 Angstrom with B factor -300 and then filtered to 5.0 Angstrom. The particles were selected using an automatic selection program FindEM.

Fitting

Fitting 1
Initial atomic model: 4EGT
Used chain: A
Fitting software: Chimera
Fitting protocol: rigid body
Refinement space: REAL
Details: Protocol: Rigid body. The P domains were separately fitted into the map by automatic rigid body fitting using Chimera (FIT IN MAP).
Fitting 2
Initial atomic model: 4EJR
Used chain: A
Fitting software: Chimera
Fitting protocol: rigid body
Refinement space: REAL
Details: Protocol: Rigid body. The S domains were separately fitted into the map by automatic rigid body fitting using Chimera (FIT IN MAP).

Imaging

Session
Microscope model: FEI TITAN KRIOS
Electron source: FIELD EMISSION GUN
Electron dose (e/A**2): 20.0
Nominal CS (mm): 2.70
Nominal magnification: 96000.0
Defocus max (nm): 2,500.00
Defocus min (nm): 1,500.00
Tilt max (degrees): 0.00
Tilt min (degrees): 0.00
Holder model: FEI TITAN KRIOS AUTOGRID HOLDER
Acquisition
Number of images: 1100
Sampling interval (μm): 15.0

Specimen

Preparation
Specimen state: particle
Buffer: 50mM Tris, 50mM NaCl, 5mM EDTA
Specimen support: 200 mesh copper grid with holey array carbon support (GiG), glow discharged
Vitrification
Apparatus: FEI VITROBOT MARK IV
Cryogen: ETHANE
Humidity: 100
Protocol: Blot for 3 seconds before plunging