EMD-5529 Experiments and Validation

6.3 A Cryo-EM Structure of a Novel Calicivirus, Tulane Virus

Single particle reconstruction
Overview of EMD-5529
Sample name: Tulane virus
Organism: Tulane virus

Map parameters

Minimum density: -14.33
Maximum density: 20.86
Average density: 0.09
Standard deviation: 1.26
Recommended contour level: 4 (author)

Sample information

Sample name: Tulane virus
Virus: Tulane virus

Validation information

Experimental information

 

Image processing

Applied symmetry: I
Software: jspr.py, EMAN, EMAN2
Number of particles: 4338
Reconstruction protocol: projection matching
CTF correction: each particle

Fitting

Initial atomic model: 1IHM
Used chains: A, B, C
Fitting software: Chimera
Fitting protocol: rigid body
Target criteria: Correlation
Refinement space: REAL
Details: Protocol: rigid body. The three chains from 1IHM were first fitted into TV density as a whole rigid body and then divided into dimers, specific chains, domains, and subdomains and fitted.

Imaging

Session
Microscope model: FEI TITAN KRIOS
Electron source: FIELD EMISSION GUN
Electron dose (e/A**2): 25.0
Nominal CS (mm): 2.7
Nominal magnification: 37000.0
Defocus max (nm): 4,891
Defocus min (nm): 1,347
Holder model: FEI TITAN KRIOS AUTOGRID HOLDER
Acquisition
Number of images: 190
Scanner model: NIKON SUPER COOLSCAN 9000
Sampling interval (μm): 6.3499999999999996

Specimen

Preparation
Specimen state: particle
Buffer: 137mM NaCl, 2.7mM KCl, 10mM Na2HPO4, 2mM KH2PO4, pH 7.4
Staining procedure: Grids with sample floated on 2% uranyl acetate for 30 seconds.
Specimen support: 400 mesh copper grid with one lacy carbon layer and one layer of ultra thin carbon on top.
Vitrification
Apparatus: HOMEMADE PLUNGER
Cryogen: ETHANE