EMD-5503 Experiments and Validation

Dissecting the in vivo assembly of the 30S ribosomal subunit reveals the role of RimM

Single particle reconstruction
Overview of EMD-5503
Sample name: Immature ribosomal small subunit from rimm gene deleted E.coli strain
Organism: Escherichia coli
Fitted atomic model: 3j2b

Map parameters

Minimum density: -7.752
Maximum density: 5.345
Average density: -4.176
Standard deviation: 0.605
Recommended contour level: -2.80 (author)

Sample information

Sample name: Immature ribosomal small subunit from rimm gene deleted E.coli strain
Ribosome: Small subunit from rimm gene deleted E.coli strain

Validation information

Experimental information

 

Image processing

Applied symmetry: C1
Software: Spider
Number of particles: 30892
Reconstruction protocol: Reference Projections
CTF correction: Weiner filter

Fitting

Initial atomic model: 3OFA
Used chain: A
Fitting software: MDFF
Fitting protocol: flexible
Target criteria: Cross-correlation
Refinement space: REAL
Details: Protocol: Initial local fitting was done using Chimera and then MDFF was used for flexible fitting. ref: Trabuco, L.G., Villa, E., Mitra, K., Frank, J. and Schulten, K. (2008) Flexible fitting of atomic structures into electron microscopy maps using molecular dynamics. Structure, 16, 673-683

Imaging

Session
Microscope model: FEI TECNAI F20
Electron source: FIELD EMISSION GUN
Electron dose (e/A**2): 20.0
Nominal magnification: 80000.0
Defocus max (nm): 4,000.00
Defocus min (nm): 1,300.00
Holder model: GATAN LIQUID NITROGEN

Specimen

Preparation
Specimen state: particle
Buffer: 150mM NH4Cl,10mM Tris-HCL,10mM MgCl2
Vitrification
Apparatus: FEI VITROBOT MARK IV
Cryogen: ETHANE
Humidity: 100
Protocol: Blot for 1 seconds before plunging