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Open data
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Basic information
Entry | Database: EMDB / ID: EMD-5015 | |||||||||
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Title | A 12.6A cryo-EM map of the 80S.eEF2.AlF4-GDP complex | |||||||||
![]() | Cryo-EM reconstruction of 80S.eEF2 complex captured in transition state | |||||||||
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![]() | AlF4- / GDP / GTPase / Ribosome / Translocation | |||||||||
Function / homology | ![]() translation elongation factor activity / GTPase activity / GTP binding / cytosol Similarity search - Function | |||||||||
Biological species | ![]() ![]() | |||||||||
Method | single particle reconstruction / cryo EM / Resolution: 12.6 Å | |||||||||
![]() | Sengupta J / Nilsson J / Gursky R / Kjeldgaard M / Nissen P / Frank J | |||||||||
![]() | ![]() Title: Visualization of the eEF2-80S ribosome transition-state complex by cryo-electron microscopy. Authors: Jayati Sengupta / Jakob Nilsson / Richard Gursky / Morten Kjeldgaard / Poul Nissen / Joachim Frank / ![]() Abstract: In an attempt to understand ribosome-induced GTP hydrolysis on eEF2, we determined a 12.6-A cryo-electron microscopy reconstruction of the eEF2-bound 80S ribosome in the presence of aluminum ...In an attempt to understand ribosome-induced GTP hydrolysis on eEF2, we determined a 12.6-A cryo-electron microscopy reconstruction of the eEF2-bound 80S ribosome in the presence of aluminum tetrafluoride and GDP, with aluminum tetrafluoride mimicking the gamma-phosphate during hydrolysis. This is the first visualization of a structure representing a transition-state complex on the ribosome. Tight interactions are observed between the factor's G domain and the large ribosomal subunit, as well as between domain IV and an intersubunit bridge. In contrast, some of the domains of eEF2 implicated in small subunit binding display a large degree of flexibility. Furthermore, we find support for a transition-state model conformation of the switch I region in this complex where the reoriented switch I region interacts with a conserved rRNA region of the 40S subunit formed by loops of the 18S RNA helices 8 and 14. This complex is structurally distinct from the eEF2-bound 80S ribosome complexes previously reported, and analysis of this map sheds light on the GTPase-coupled translocation mechanism. | |||||||||
History |
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Structure visualization
Movie |
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Structure viewer | EM map: ![]() ![]() ![]() |
Supplemental images |
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Downloads & links
-EMDB archive
Map data | ![]() | 7.9 MB | ![]() | |
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Header (meta data) | ![]() ![]() | 9.3 KB 9.3 KB | Display Display | ![]() |
Images | ![]() | 327.6 KB | ||
Archive directory | ![]() ![]() | HTTPS FTP |
-Validation report
Summary document | ![]() | 78 KB | Display | ![]() |
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Full document | ![]() | 77.1 KB | Display | |
Data in XML | ![]() | 493 B | Display | |
Arichive directory | ![]() ![]() | HTTPS FTP |
-Related structure data
Related structure data | ![]() 3dwuMC ![]() 5016C ![]() 5017C ![]() 3dnyC C: citing same article ( M: atomic model generated by this map |
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Similar structure data |
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Links
EMDB pages | ![]() ![]() |
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Related items in Molecule of the Month |
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Map
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Annotation | Cryo-EM reconstruction of 80S.eEF2 complex captured in transition state | ||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||
Voxel size | X=Y=Z: 2.82 Å | ||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||
Density |
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Symmetry | Space group: 1 | ||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||
Details | EMDB XML:
CCP4 map header:
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-Supplemental data
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Sample components
-Entire : The eEF2-bound 80S ribosome in the presence of aluminum fluoride ...
Entire | Name: The eEF2-bound 80S ribosome in the presence of aluminum fluoride (AlF4-) and GDP |
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Components |
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-Supramolecule #1000: The eEF2-bound 80S ribosome in the presence of aluminum fluoride ...
Supramolecule | Name: The eEF2-bound 80S ribosome in the presence of aluminum fluoride (AlF4-) and GDP type: sample / ID: 1000 / Details: AlF4- mimicking the g-phosphate during hydrolysis / Number unique components: 4 |
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-Supramolecule #1: 80S ribosome
Supramolecule | Name: 80S ribosome / type: complex / ID: 1 / Recombinant expression: No / Database: NCBI / Ribosome-details: ribosome-eukaryote: ALL |
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Source (natural) | Organism: ![]() ![]() |
-Experimental details
-Structure determination
Method | cryo EM |
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![]() | single particle reconstruction |
Aggregation state | particle |
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Sample preparation
Buffer | pH: 7.2 / Details: 20 mM Hepes-NH3, 100 mM KCl, 20 mM MgCl2 |
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Grid | Details: Quantifoil grid |
Vitrification | Cryogen name: NITROGEN / Chamber humidity: 90 % / Chamber temperature: 93 K / Instrument: OTHER / Details: Vitrification instrument: Vitrobot |
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Electron microscopy
Microscope | FEI TECNAI F20 |
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Image recording | Digitization - Sampling interval: 2.82 µm / Average electron dose: 10 e/Å2 / Bits/pixel: 16 |
Electron beam | Acceleration voltage: 200 kV / Electron source: ![]() |
Electron optics | Illumination mode: FLOOD BEAM / Imaging mode: BRIGHT FIELD / Nominal defocus max: 4.5 µm / Nominal defocus min: 1.5 µm / Nominal magnification: 50000 |
Sample stage | Specimen holder: Single tilt cyro-holder / Specimen holder model: GATAN LIQUID NITROGEN |
Experimental equipment | ![]() Model: Tecnai F20 / Image courtesy: FEI Company |
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Image processing
Details | The image processing using SPIDER included a 3D projection alignment procedure with correction of the contrast transfer function and enhancement of the high-resolution Fourier amplitudes based on X-ray solution scattering data. |
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CTF correction | Details: Segregation in defocus groups and correction in volumes |
Final reconstruction | Algorithm: OTHER / Resolution.type: BY AUTHOR / Resolution: 12.6 Å / Resolution method: FSC 0.5 CUT-OFF / Software - Name: Spider / Details: Supervised Classification was used / Number images used: 28242 |
-Atomic model buiding 1
Initial model | PDB ID: |
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Details | The domains were separately fitted by manual docking using program O |
Refinement | Space: REAL / Protocol: RIGID BODY FIT / Target criteria: correlation coefficient |
Output model | ![]() PDB-3dwu: |