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Yorodumi- EMDB-5658: Cryo-electron microscopy structure of the mammalian 43S preinitia... -
+Open data
-Basic information
Entry | Database: EMDB / ID: EMD-5658 | |||||||||
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Title | Cryo-electron microscopy structure of the mammalian 43S preinitiation complex bound to DHX29 | |||||||||
Map data | Reconstruction of the mammalian 43S preinitiation complex bound to DHX29 | |||||||||
Sample |
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Keywords | 43S / preinitiation complex / eIF3 / eIF2 / Met-tRNAiMet / DHX29 / 40S | |||||||||
Function / homology | Function and homology information positive regulation of mRNA binding / viral translational termination-reinitiation / eukaryotic translation initiation factor 3 complex, eIF3e / eukaryotic translation initiation factor 3 complex, eIF3m / translation reinitiation / IRES-dependent viral translational initiation / multi-eIF complex / formation of cytoplasmic translation initiation complex / eukaryotic translation initiation factor 3 complex / eukaryotic 43S preinitiation complex ...positive regulation of mRNA binding / viral translational termination-reinitiation / eukaryotic translation initiation factor 3 complex, eIF3e / eukaryotic translation initiation factor 3 complex, eIF3m / translation reinitiation / IRES-dependent viral translational initiation / multi-eIF complex / formation of cytoplasmic translation initiation complex / eukaryotic translation initiation factor 3 complex / eukaryotic 43S preinitiation complex / cytoplasmic translational initiation / eukaryotic 48S preinitiation complex / metal-dependent deubiquitinase activity / regulation of translational initiation / Formation of the ternary complex, and subsequently, the 43S complex / nuclear-transcribed mRNA catabolic process, nonsense-mediated decay / Ribosomal scanning and start codon recognition / Translation initiation complex formation / Formation of a pool of free 40S subunits / GTP hydrolysis and joining of the 60S ribosomal subunit / L13a-mediated translational silencing of Ceruloplasmin expression / negative regulation of translational initiation / negative regulation of proteasomal ubiquitin-dependent protein catabolic process / translation initiation factor binding / translation initiation factor activity / positive regulation of translation / translational initiation / PML body / negative regulation of ERK1 and ERK2 cascade / receptor tyrosine kinase binding / fibrillar center / metallopeptidase activity / ribosome binding / ubiquitinyl hydrolase 1 / microtubule / cysteine-type deubiquitinase activity / postsynaptic density / cadherin binding / mRNA binding / synapse / chromatin / nucleolus / structural molecule activity / proteolysis / RNA binding / extracellular exosome / nucleoplasm / identical protein binding / membrane / nucleus / cytoplasm / cytosol Similarity search - Function | |||||||||
Biological species | Oryctolagus cuniculus (rabbit) | |||||||||
Method | single particle reconstruction / cryo EM / Resolution: 11.6 Å | |||||||||
Authors | Hashem Y / des Georges A / Dhote V / Langlois R / Liao HY / Grassucci RA / Hellen CUT / Pestova TV / Frank J | |||||||||
Citation | Journal: Cell / Year: 2013 Title: Structure of the mammalian ribosomal 43S preinitiation complex bound to the scanning factor DHX29. Authors: Yaser Hashem / Amedee des Georges / Vidya Dhote / Robert Langlois / Hstau Y Liao / Robert A Grassucci / Christopher U T Hellen / Tatyana V Pestova / Joachim Frank / Abstract: Eukaryotic translation initiation begins with assembly of a 43S preinitiation complex. First, methionylated initiator methionine transfer RNA (Met-tRNAi(Met)), eukaryotic initiation factor (eIF) 2, ...Eukaryotic translation initiation begins with assembly of a 43S preinitiation complex. First, methionylated initiator methionine transfer RNA (Met-tRNAi(Met)), eukaryotic initiation factor (eIF) 2, and guanosine triphosphate form a ternary complex (TC). The TC, eIF3, eIF1, and eIF1A cooperatively bind to the 40S subunit, yielding the 43S preinitiation complex, which is ready to attach to messenger RNA (mRNA) and start scanning to the initiation codon. Scanning on structured mRNAs additionally requires DHX29, a DExH-box protein that also binds directly to the 40S subunit. Here, we present a cryo-electron microscopy structure of the mammalian DHX29-bound 43S complex at 11.6 Å resolution. It reveals that eIF2 interacts with the 40S subunit via its α subunit and supports Met-tRNAi(Met) in an unexpected P/I orientation (eP/I). The structural core of eIF3 resides on the back of the 40S subunit, establishing two principal points of contact, whereas DHX29 binds around helix 16. The structure provides insights into eukaryote-specific aspects of translation, including the mechanism of action of DHX29. | |||||||||
History |
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-Structure visualization
Movie |
Movie viewer |
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Structure viewer | EM map: SurfViewMolmilJmol/JSmol |
Supplemental images |
-Downloads & links
-EMDB archive
Map data | emd_5658.map.gz | 44.9 MB | EMDB map data format | |
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Header (meta data) | emd-5658-v30.xml emd-5658.xml | 16.6 KB 16.6 KB | Display Display | EMDB header |
Images | emd_5658_1.jpg | 79.9 KB | ||
Archive directory | http://ftp.pdbj.org/pub/emdb/structures/EMD-5658 ftp://ftp.pdbj.org/pub/emdb/structures/EMD-5658 | HTTPS FTP |
-Validation report
Summary document | emd_5658_validation.pdf.gz | 323.6 KB | Display | EMDB validaton report |
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Full document | emd_5658_full_validation.pdf.gz | 323.1 KB | Display | |
Data in XML | emd_5658_validation.xml.gz | 6 KB | Display | |
Arichive directory | https://ftp.pdbj.org/pub/emdb/validation_reports/EMD-5658 ftp://ftp.pdbj.org/pub/emdb/validation_reports/EMD-5658 | HTTPS FTP |
-Related structure data
Related structure data | 3j8cM M: atomic model generated by this map |
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Similar structure data |
-Links
EMDB pages | EMDB (EBI/PDBe) / EMDataResource |
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Related items in Molecule of the Month |
-Map
File | Download / File: emd_5658.map.gz / Format: CCP4 / Size: 47.7 MB / Type: IMAGE STORED AS FLOATING POINT NUMBER (4 BYTES) | ||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||
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Annotation | Reconstruction of the mammalian 43S preinitiation complex bound to DHX29 | ||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||
Voxel size | X=Y=Z: 2.245 Å | ||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||
Density |
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Symmetry | Space group: 1 | ||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||
Details | EMDB XML:
CCP4 map header:
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-Supplemental data
-Sample components
-Entire : Cryo-electron microscopy structure of the mammalian 43S preinitia...
Entire | Name: Cryo-electron microscopy structure of the mammalian 43S preinitiation complex bound to DHX29 |
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Components |
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-Supramolecule #1000: Cryo-electron microscopy structure of the mammalian 43S preinitia...
Supramolecule | Name: Cryo-electron microscopy structure of the mammalian 43S preinitiation complex bound to DHX29 type: sample / ID: 1000 Oligomeric state: One 40S binds one eIF3, one eIF2, one Met-tRNAiMet, and one DHX29 Number unique components: 5 |
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-Supramolecule #1: eukaryotic small ribosomal subunit
Supramolecule | Name: eukaryotic small ribosomal subunit / type: complex / ID: 1 / Name.synonym: 40S subunit / Recombinant expression: No / Database: NCBI / Ribosome-details: ribosome-eukaryote: SSU 40S |
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Source (natural) | Organism: Oryctolagus cuniculus (rabbit) / synonym: rabbit / Tissue: blood / Cell: reticulocytes |
Molecular weight | Theoretical: 1.5 MDa |
-Macromolecule #1: eukaryotic initiation factor 3
Macromolecule | Name: eukaryotic initiation factor 3 / type: protein_or_peptide / ID: 1 / Name.synonym: eIF3 / Number of copies: 1 / Oligomeric state: monomer / Recombinant expression: No / Database: NCBI |
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Source (natural) | Organism: Oryctolagus cuniculus (rabbit) / synonym: rabbit / Tissue: blood / Cell: reticulocytes |
Molecular weight | Theoretical: 800 KDa |
-Macromolecule #2: eukaryotic initiation factor 2
Macromolecule | Name: eukaryotic initiation factor 2 / type: protein_or_peptide / ID: 2 / Name.synonym: eIF2 / Number of copies: 1 / Oligomeric state: monomer / Recombinant expression: No / Database: NCBI |
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Source (natural) | Organism: Oryctolagus cuniculus (rabbit) / synonym: rabbit / Tissue: blood / Cell: reticulocytes |
Molecular weight | Theoretical: 150 KDa |
-Macromolecule #3: eukaryotic initiation factor 1
Macromolecule | Name: eukaryotic initiation factor 1 / type: protein_or_peptide / ID: 3 / Name.synonym: eIF1 / Number of copies: 1 / Oligomeric state: monomer / Recombinant expression: Yes |
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Source (natural) | Organism: Oryctolagus cuniculus (rabbit) / synonym: rabbit |
Molecular weight | Theoretical: 14 KDa |
Recombinant expression | Organism: Escherichia coli (E. coli) / Recombinant plasmid: pT7 |
-Macromolecule #4: eukaryotic initiation factor 1A
Macromolecule | Name: eukaryotic initiation factor 1A / type: protein_or_peptide / ID: 4 / Name.synonym: eIF1A / Number of copies: 1 / Oligomeric state: monomer / Recombinant expression: Yes |
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Source (natural) | Organism: Oryctolagus cuniculus (rabbit) / synonym: rabbit |
Molecular weight | Theoretical: 19 KDa |
Recombinant expression | Organism: Escherichia coli (E. coli) / Recombinant plasmid: pT7 |
-Macromolecule #5: DHX29
Macromolecule | Name: DHX29 / type: protein_or_peptide / ID: 5 / Number of copies: 1 / Oligomeric state: monomer / Recombinant expression: Yes |
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Source (natural) | Organism: Oryctolagus cuniculus (rabbit) / synonym: rabbit |
Molecular weight | Theoretical: 150 KDa |
Recombinant expression | Organism: Escherichia coli (E. coli) / Recombinant plasmid: pT7 |
-Macromolecule #6: Transfer RNA
Macromolecule | Name: Transfer RNA / type: rna / ID: 6 / Name.synonym: tRNA / Details: Met-tRNAiMet / Classification: TRANSFER / Structure: DOUBLE HELIX / Synthetic?: Yes |
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Source (natural) | Organism: Oryctolagus cuniculus (rabbit) / synonym: Rabbit |
Molecular weight | Theoretical: 25 KDa |
-Experimental details
-Structure determination
Method | cryo EM |
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Processing | single particle reconstruction |
Aggregation state | particle |
-Sample preparation
Concentration | 0.105 mg/mL |
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Buffer | pH: 7.5 Details: 20 mM Tris, 100 mM potassium acetate, 2 mM DTT, 2.5 mM magnesium chloride, 0.25 mM spermidine |
Grid | Details: 300 mesh copper/molybdenum holey carbon-coated Quantifoil 2/4 grid (Quantifoil Micro Tools GmbH) containing an additional continuous thin layer of carbon |
Vitrification | Cryogen name: ETHANE / Chamber humidity: 100 % / Chamber temperature: 120 K / Instrument: FEI VITROBOT MARK II / Method: Blot for seconds before plunging |
-Electron microscopy
Microscope | FEI TECNAI 20 |
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Temperature | Average: 110 K |
Date | Nov 1, 2012 |
Image recording | Category: CCD / Film or detector model: GATAN ULTRASCAN 4000 (4k x 4k) / Number real images: 8000 / Average electron dose: 12 e/Å2 / Bits/pixel: 32 |
Electron beam | Acceleration voltage: 110 kV / Electron source: FIELD EMISSION GUN |
Electron optics | Calibrated magnification: 51570 / Illumination mode: FLOOD BEAM / Imaging mode: BRIGHT FIELD / Cs: 2.26 mm / Nominal defocus max: -4.0 µm / Nominal defocus min: -1.0 µm |
Sample stage | Specimen holder model: GATAN LIQUID NITROGEN |
-Image processing
Details | The particles of this reconstruction were obtained after particle sorting using Relion |
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CTF correction | Details: Each particle |
Final reconstruction | Algorithm: OTHER / Resolution.type: BY AUTHOR / Resolution: 11.6 Å / Resolution method: OTHER / Software - Name: Spider, Relion / Number images used: 29000 |
-Atomic model buiding 1
Initial model | PDB ID: 2xzm |
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Software | Name: Chimera |
Refinement | Space: REAL / Protocol: RIGID BODY FIT |
Output model | PDB-3j8c: |
-Atomic model buiding 2
Initial model | PDB ID: |
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Software | Name: Chimera |
Details | Domains D1-D2 of eIF2-alpha were fitted manually in Chimera in order to accommodate them to their density |
Refinement | Space: REAL / Protocol: RIGID BODY FIT |
Output model | PDB-3j8c: |