EMD-5527 Experiments and Validation

Reconstruction of the deltaGCC mutant HCV IRES bound to rabbit 40S ribosomal subunit

Single particle reconstruction
Overview of EMD-5527
Sample name: mutant HCV IRES bound to 40S
Organisms: Oryctolagus cuniculus, Hepatitis C virus

Map parameters

Recommended contour level: 2.25 (author)
Number of grid points: 150 × 150 × 150
Voxel size: 2.66 × 2.66 × 2.66 Å
Minimum density: -6.434
Maximum density: 9.64
Average density: 0.00
Standard deviation: 1.00

Sample information

Sample name: mutant HCV IRES bound to 40S
Nucleic acid: hepatitis C virus internal ribosome entry site
Ribosome: Rabbit 40S ribosomal subunit

Validation information

Experimental information

 

Image processing

Applied symmetry: C1
Software: FREALIGN
Number of particles: 29920
CTF correction: CTFFIND3

Imaging

Session
Microscope model: FEI TECNAI F20
Electron source: FIELD EMISSION GUN
Electron dose (e/A**2): 15.0
Nominal CS (mm): 2.00
Nominal magnification: 62000.0
Defocus max (nm): 5,000.00
Defocus min (nm): 1,250.00
Tilt max (degrees): 0.00
Tilt min (degrees): 0.00
Holder model: GATAN LIQUID NITROGEN
Acquisition
Number of images: 1790
Details: Used 2x binned images yielding a pixel size of 2.66 A/pixel.

Specimen

Preparation
Specimen state: particle
Buffer: 20 mM Tris-HCl, pH 7.4, 200 mM KOAc, 2.5 mM MgCl2, 2 mM DTT, 40 mM KCl
Specimen support: Quantifoil R1.2/1.3 400 mesh copper grids with holey carbon
Vitrification
Apparatus: FEI VITROBOT MARK IV
Cryogen: ETHANE
Humidity: 100
Protocol: Blot and plunge into liquid ethane