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- EMDB-2683: Mammalian 80S-HCV-IRES initiation complex with eIF5B POST-like state -
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Open data
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Basic information
Entry | Database: EMDB / ID: EMD-2683 | |||||||||
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Title | Mammalian 80S-HCV-IRES initiation complex with eIF5B POST-like state | |||||||||
![]() | Mammalian translation initiation 80S HCV-IRES complex with eIF5B POST-like state | |||||||||
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![]() | ribosome / translation initiation / Hepatitis C Virus internal ribosome entry site / eukaryotic initiation factor 5B | |||||||||
Function / homology | ![]() regulation of translational initiation / translation initiation factor activity / GTPase activity / GTP binding / mitochondrion / cytoplasm Similarity search - Function | |||||||||
Biological species | ![]() ![]() ![]() | |||||||||
Method | single particle reconstruction / cryo EM / Resolution: 9.5 Å | |||||||||
![]() | Yamamoto H / Unbehaun A / Loerke J / Behrmann E / Collier M / Marianne C / Burger J / Mielke T / Spahn CMT | |||||||||
![]() | ![]() Title: Structure of the mammalian 80S initiation complex with initiation factor 5B on HCV-IRES RNA. Authors: Hiroshi Yamamoto / Anett Unbehaun / Justus Loerke / Elmar Behrmann / Marianne Collier / Jörg Bürger / Thorsten Mielke / Christian M T Spahn / ![]() Abstract: The universally conserved eukaryotic initiation factor (eIF) 5B, a translational GTPase, is essential for canonical translation initiation. It is also required for initiation facilitated by the ...The universally conserved eukaryotic initiation factor (eIF) 5B, a translational GTPase, is essential for canonical translation initiation. It is also required for initiation facilitated by the internal ribosomal entry site (IRES) of hepatitis C virus (HCV) RNA. eIF5B promotes joining of 60S ribosomal subunits to 40S ribosomal subunits bound by initiator tRNA (Met-tRNAi(Met)). However, the exact molecular mechanism by which eIF5B acts has not been established. Here we present cryo-EM reconstructions of the mammalian 80S-HCV-IRES-Met-tRNAi(Met)-eIF5B-GMPPNP complex. We obtained two substates distinguished by the rotational state of the ribosomal subunits and the configuration of initiator tRNA in the peptidyl (P) site. Accordingly, a combination of conformational changes in the 80S ribosome and in initiator tRNA facilitates binding of the Met-tRNAi(Met) to the 60S P site and redefines the role of eIF5B as a tRNA-reorientation factor. | |||||||||
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 | ![]() | 9.1 MB | ![]() | |
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Header (meta data) | ![]() ![]() | 11.8 KB 11.8 KB | Display Display | ![]() |
Images | ![]() | 150.1 KB | ||
Archive directory | ![]() ![]() | HTTPS FTP |
-Related structure data
Related structure data | ![]() 4ujcMC ![]() 2682C ![]() 4ujdC M: atomic model generated by this map C: citing same article ( |
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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 | Mammalian translation initiation 80S HCV-IRES complex with eIF5B POST-like state | ||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||
Voxel size | X=Y=Z: 1.26 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 : Mammalian translation initiation 80S HCV-IRES complex with eIF5B;...
Entire | Name: Mammalian translation initiation 80S HCV-IRES complex with eIF5B; POST-like state |
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Components |
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-Supramolecule #1000: Mammalian translation initiation 80S HCV-IRES complex with eIF5B;...
Supramolecule | Name: Mammalian translation initiation 80S HCV-IRES complex with eIF5B; POST-like state type: sample / ID: 1000 / Number unique components: 4 |
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Molecular weight | Theoretical: 4.8 MDa |
-Supramolecule #1: 80S ribosome
Supramolecule | Name: 80S ribosome / type: complex / ID: 1 / Recombinant expression: No / Ribosome-details: ribosome-eukaryote: ALL |
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Source (natural) | Organism: ![]() ![]() |
Molecular weight | Theoretical: 4.5 MDa |
-Macromolecule #1: tRNA
Macromolecule | Name: tRNA / type: rna / ID: 1 / Classification: TRANSFER / Structure: DOUBLE HELIX / Synthetic?: No |
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Source (natural) | Organism: ![]() ![]() |
Molecular weight | Theoretical: 2.5 KDa |
-Macromolecule #3: HCV-IRES
Macromolecule | Name: HCV-IRES / type: rna / ID: 3 / Classification: OTHER / Structure: DOUBLE HELIX / Synthetic?: Yes |
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Source (natural) | Organism: ![]() |
Molecular weight | Theoretical: 162 KDa |
Sequence | String: GCCAGCCCCC UGAUGGGGGC GACACUCCAC CAUGAAUCAC UCCCCUGUGA GGAACUACUG UCUUCACGCA GAAAGCGUCU AGCCAUGGCG UUAGUAUGAG UGUCGUGCAG CCUCCAGGAC CCCCCCUCCC GGGAGAGCCA UAGUGGUCUG CGGAACCGGU GAGUACACCG ...String: GCCAGCCCCC UGAUGGGGGC GACACUCCAC CAUGAAUCAC UCCCCUGUGA GGAACUACUG UCUUCACGCA GAAAGCGUCU AGCCAUGGCG UUAGUAUGAG UGUCGUGCAG CCUCCAGGAC CCCCCCUCCC GGGAGAGCCA UAGUGGUCUG CGGAACCGGU GAGUACACCG GAAUUGCCAG GACGACCGGG UCCUUUCUUG GAUAAACCCG CUCAAUGCCU GGAGAUUUGG GCGUGCCCCC GCAAGACUGC UAGCCGAGUA GUGUUGGGUC GCGAAAGGCC UUGUGGUACU GCCUGAUAGG GUGCUUGCGA GUGCCCCGGG AGGUCUCGUA GACCGUGCAC CAUGAGCACG AAUCCUAAAC CUCAAAGAAA AACCAAACGU AACACCAACC GUCGCCCACA GGACGUCAAG UUCCCGGGUG GCGGUCUAGA CGCCGAGAUC AGAAAUCCCU CUCUCGGAUC GCAUUUGGAC UUCUGCCUUC GGGCACCACG GUCGGAUCCG AAUU |
-Macromolecule #2: eIF5B
Macromolecule | Name: eIF5B / type: protein_or_peptide / ID: 2 / Number of copies: 1 / Recombinant expression: No |
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Source (natural) | Organism: ![]() ![]() |
Molecular weight | Theoretical: 160 KDa |
Sequence | UniProtKB: Eukaryotic translation initiation factor 5B |
-Experimental details
-Structure determination
Method | cryo EM |
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![]() | single particle reconstruction |
Aggregation state | particle |
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Sample preparation
Concentration | 0.15 mg/mL |
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Buffer | pH: 7.6 Details: 20mM Tris-HCl, 5mM MgCl2, 100mM KCl, 0.2mM spermidine, 2mM DTT |
Grid | Details: Quantifoil R3-3 Cu 300 mesh with 2 nm carbon support film |
Vitrification | Cryogen name: ETHANE / Chamber humidity: 100 % / Chamber temperature: 93 K / Instrument: FEI VITROBOT MARK I / Method: blot for 2-4 seconds before plunging |
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Electron microscopy
Microscope | FEI POLARA 300 |
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Alignment procedure | Legacy - Astigmatism: Objective lens astigmatism was corrected at 115,000 times magnification |
Details | automated data collection using Leginon |
Date | Dec 22, 2012 |
Image recording | Category: CCD / Film or detector model: TVIPS TEMCAM-F416 (4k x 4k) / Number real images: 22287 / Average electron dose: 20 e/Å2 / Details: Automated data collection on using Leginon / Bits/pixel: 16 |
Electron beam | Acceleration voltage: 300 kV / Electron source: ![]() |
Electron optics | Calibrated magnification: 194805 / Illumination mode: FLOOD BEAM / Imaging mode: BRIGHT FIELD / Cs: 2.0 mm / Nominal defocus max: 4.5 µm / Nominal defocus min: 2.0 µm / Nominal magnification: 115000 |
Sample stage | Specimen holder: Cartridge system, LN2 cooled / Specimen holder model: GATAN HELIUM |
Experimental equipment | ![]() Model: Tecnai Polara / Image courtesy: FEI Company |
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Image processing
CTF correction | Details: CTFFIND3 |
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Final reconstruction | Applied symmetry - Point group: C1 (asymmetric) / Resolution.type: BY AUTHOR / Resolution: 9.5 Å / Resolution method: OTHER / Software - Name: spider, sparx / Number images used: 541570 |