5LZB
Structure of SelB-Sec-tRNASec bound to the 70S ribosome in the initial binding state (IB)
ELECTRON MICROSCOPY
| Sample |
|---|
| Initial binding state of E. coli ribosome 70S-SelB-GDPNP-Sec-tRNASec-fMet-tRNAfmet-SECIS mRNA complex (IB) |
| Specimen Preparation | |
|---|---|
| Sample Aggregation State | PARTICLE |
| Vitrification Instrument | FEI VITROBOT MARK IV |
| Cryogen Name | ETHANE |
| Sample Vitrification Details | |
| 3D Reconstruction | |
|---|---|
| Reconstruction Method | SINGLE PARTICLE |
| Number of Particles | 8002 |
| Reported Resolution (Å) | 5.3 |
| Resolution Method | FSC 0.143 CUT-OFF |
| Other Details | Gold-standard refinement of independent half maps in Relion 1.3 |
| Refinement Type | |
| Symmetry Type | POINT |
| Map-Model Fitting and Refinement | |||||
|---|---|---|---|---|---|
| Id | 1 | ||||
| Refinement Space | |||||
| Refinement Protocol | FLEXIBLE FIT | ||||
| Refinement Target | Maximum likelihood | ||||
| Overall B Value | |||||
| Fitting Procedure | |||||
| Details | For parts of the model exhibiting larger conformational differences and/or lower local map resolution, additional cycles of real space refinement and ... | ||||
| Data Acquisition | |||||||||
|---|---|---|---|---|---|---|---|---|---|
| Detector Type | FEI FALCON II (4k x 4k) | ||||||||
| Electron Dose (electrons/Å**2) | 30 | ||||||||
| Imaging Experiment | 1 |
|---|---|
| Date of Experiment | |
| Temperature (Kelvin) | |
| Microscope Model | FEI TITAN KRIOS |
| Minimum Defocus (nm) | 700 |
| Maximum Defocus (nm) | 2600 |
| Minimum Tilt Angle (degrees) | |
| Maximum Tilt Angle (degrees) | |
| Nominal CS | 0.001 |
| Imaging Mode | BRIGHT FIELD |
| Specimen Holder Model | FEI TITAN KRIOS AUTOGRID HOLDER |
| Nominal Magnification | 59000 |
| Calibrated Magnification | |
| Source | FIELD EMISSION GUN |
| Acceleration Voltage (kV) | 300 |
| Imaging Details | Using a Cs-corrector from CEOS electron optical aberrations were corrected to residual phase errors of 45degree at scattering angles of >12 to 15 mrad. |
| EM Software | ||
|---|---|---|
| Task | Software Package | Version |
| MODEL REFINEMENT | Rosetta | 3.5 |
| MODEL REFINEMENT | Coot | 0.8.3 |
| MODEL REFINEMENT | RCrane | 1.1 |
| MODEL REFINEMENT | PHENIX | 1.10.1 & developmental |
| MODEL REFINEMENT | PHENIX | 1.10.1 |
| RECONSTRUCTION | RELION | 1.3 |
| MODEL REFINEMENT | ERRASER | |
| Image Processing | ||||
|---|---|---|---|---|
| CTF Correction Type | CTF Correction Details | Number of Particles Selected | Particle Selection Details | |
| PHASE FLIPPING ONLY | Local CTF correction, after MSA based classification and averaging of local power spectra | 969526 | 969526 ribosome particles showing Thon rings better than 3.5A (see CTF correction) | |
