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New mRNA-tRNA translocation intermediates revealed by cryoEM: Class 6 (rotated 70S ribosome with A-site and P-site tRNAs)
by single particle reconstruction, at 11.5 A resolution
#1: Surface view with section colored by density value, Surface level: 0.15, Made by UCSF CHIMERA
#2: Surface view colored by height, Surface level: 0.15, Made by UCSF CHIMERA
#3: Surface view with fitted model, atomic models: PDB-3j10, PDB-3j14, Surface level: 0.15, Made by UCSF CHIMERA
#4: Simplified surface model with fitted atomic model: PDB-3j10, Made by Jmol
#5: Simplified surface model with fitted atomic model: PDB-3j14, Made by Jmol
 Entry | |
| Summary | |
| Database / ID | EM DATA BANK (EMDB) / 5362 |
|---|---|
| Authors | Agirrezabala X, Liao HY, Schreiner E, Fu J, Ortiz-Meoz RF, Schulten K, Green R, Frank J |
| EMDB Sites |  EMDB @PDBe (EU), EMDB @RCSB (USA) |
| Structure Visualization | |
| Movies |  Movie Page#1: Surface view with section colored by density value, Surface level: 0.15, Made by UCSF CHIMERA #2: Surface view colored by height, Surface level: 0.15, Made by UCSF CHIMERA #3: Surface view with fitted model, atomic models: PDB-3j10, PDB-3j14, Surface level: 0.15, Made by UCSF CHIMERA #4: Simplified surface model with fitted atomic model: PDB-3j10, Made by Jmol #5: Simplified surface model with fitted atomic model: PDB-3j14, Made by Jmol |
| Supplemental images | |
| Structure viewers |  Yorodumi,  Launch PeppeR (About PeppeR), Volume viewer (RCSB, PDBe) |
| Related Structure Data | |
| Related Entries |
Cite: data citing same article Fit: output model of fitting |
| Similar strucutres (beta) |
 List of similar structure data  about Omokage system |
 Article | |
| Citation - Primary | |
| Article | Proc. Natl. Acad. Sci. U.S.A., Vol. 109, Issue 16, Page 6094-9, Year 2012 |
|---|---|
| Title | Structural characterization of mRNA-tRNA translocation intermediates. |
| Authors | Xabier Agirrezabala, Hstau Y Liao, Eduard Schreiner, Jie Fu, Rodrigo F Ortiz-Meoz, Klaus Schulten, Rachel Green, Joachim Frank Structural Biology Unit, CIC-bioGUNE, Bizkaia Technology Park, Derio 48160, Basque Country, Spain. |
| Keywords |  Binding Sites, Cryoelectron Microscopy, Crystallography, X-Ray, Escherichia coli (genetics), Macromolecular Substances (chemistry), Models, Molecular, Molecular Conformation, Protein Biosynthesis, RNA, Messenger (genetics), RNA, Transfer (genetics, 9014-25-9), Ribosomes (metabolism), Thermodynamics |
| Links | DOI: 10.1073/pnas.1201288109, PubMed: 22467828, PMC: PMC3340995 |
 Map | |||||||||||||||||||||||||
| File |  EMD-5362.map ( map file in CCP4 format, 62502 KB ) | ||||||||||||||||||||||||
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| Projections & Slices | Size of images:
Images are generated by Spider package. | ||||||||||||||||||||||||
| Density |
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| Data Type | Image stored as Reals | ||||||||||||||||||||||||
| Space Group Number | 1 | ||||||||||||||||||||||||
| Map Geometry |
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| Unit Cell | A = 375.0 A , B = 375.0 A , C = 375.0 A , alpha = 90.0 degrees , beta = 90.0 degrees , gamma = 90.0 degrees | ||||||||||||||||||||||||
| Pixel Spacing | X = 1.5 A , Y = 1.5 A , Z = 1.5 A | ||||||||||||||||||||||||
| CCP4 map header info | |||||||||||||||||||||||||
| Annotation Details | Cryo-EM-revealed rotated 70S ribosome with A and P site tRNAs (class 6) | ||||||||||||||||||||||||
 Supplement |
 Sample | |
| Name | Trp-tRNA-EFTu-GDP-kir-70S ribosome |
|---|---|
| Number of Components | 3 |
| Oligomeric State | monomeric |
| Theoretical Mass | 2.8 MDa |
| Experimental Mass | 2.8 MDa |
| Component #1: ribosome-prokaryote - 70S ribosome | |
| Scientific name | 70S ribosome |
| Scientific Name of Species | Escherichia coli (NCBI Taxonomy: 562)
|
| Prokaryote | ALL |
| Recombinant expression | No |
 Experiment | |
| Sample Preparation | |
| Specimen State | particle |
|---|---|
| Specimen Conc | 0.03 mg/ml |
| Specimen Support Details | 200 mesh |
| Buffer | Details: HiFi buffer (50 mM Tris-HCl, pH 7.5, 70mM NH4Cl, 30 mM KCl, 3.5 mM MgCl2, 0.5 mM spermidine, 8mM putrescine, 2 mM DTT, 3.5 mM MgCl2) pH: 7.5 |
| Vitrification | |
| Method | blot for 3 seconds |
| Cryogen Name | NITROGEN |
| Details | Vitrification instrument: Vitrobot |
| Humidity | 90 |
| Instrument | FEI VITROBOT |
| Temperature | 80 Kelvin |
| Imaging | |
| Microscope | FEI TECNAI F30 |
| Date | 01-MAY-2007 |
| Details | automated data collection system AutoEMation (CCD mag. 100000x) |
| Electron Gun | |
| Electron Source | FIELD EMISSION GUN |
| Accelerating Voltage | 300 kV |
| Electron Dose | 25 e/A**2 |
| Electron Beam Tilt Params | no tilt |
| Illumination Mode | FLOOD BEAM |
| Lens | |
| Magnification | Nominal: 59000 X, Calibrated: 58269 X |
| Astigmatism | objective corrected at 100,000 times magnification |
| Nominal Cs | 2.26 mm |
| Imaging Mode | BRIGHT FIELD |
| Defocus | 1200 nm - 4000 nm |
| Energy Filter | Type: no filter , Window: |
| Specimen Holder | |
| Holder | cartridge ( OTHER ) |
| Tilt Angle | 0 degrees - 0 degrees |
| Temperature | 80.7 Kelvin ( 80.7 Kelvin - 80.7 Kelvin ) |
| Camera | |
| Detector | TVIPS TemCam-F415 (CCD) |
| Image Acquisition | |
 Processing | |
| Method | single particle reconstruction |
|---|---|
| 3 D reconstruction | |
| Software | XMIPP, SPIDER |
| CTF Correction | defocus groups, Wiener filter |
| Details | Classification using ML3D |
| Resolution By Author | 11.5 |
| Resolution Method | FSC at 0.5 cut-off |
| Single Particle | |
| Applied Symmetry | C1 (asymmetric) |
| Number of Projections | 37000 |
| Atomic Model Fitting | |
| Model #0 | |
| Details | Protocol: Molecular Dynamics based flexible fitting |
| Refinement Protocol | flexible |
| Refinement Space | REAL |
| Target Criteria | RMSD, cross correlation |
| Software | MDFF |
| Model #1 | |
| Refinement Space | REAL |
| Target Criteria | RMSD, cross correlation |
| Refinement Protocol | flexible |
| Details | Protocol: Molecular Dynamics based flexible fitting |
| Software | MDFF |
| Fitted Coordinate | |
| PDB entry ID | |
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