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Yorodumi- PDB-7syw: Structure of the wt IRES eIF5B-containing 48S initiation complex,... -
+Open data
-Basic information
Entry | Database: PDB / ID: 7syw | |||||||||||||||
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Title | Structure of the wt IRES eIF5B-containing 48S initiation complex, closed conformation. Structure 15(wt) | |||||||||||||||
Components |
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Keywords | RIBOSOME / HCV / IRES / 40S | |||||||||||||||
Function / homology | Function and homology information multi-eIF complex / eukaryotic 43S preinitiation complex / translation factor activity, RNA binding / eukaryotic 48S preinitiation complex / ribosomal subunit / Formation of the ternary complex, and subsequently, the 43S complex / 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 ...multi-eIF complex / eukaryotic 43S preinitiation complex / translation factor activity, RNA binding / eukaryotic 48S preinitiation complex / ribosomal subunit / Formation of the ternary complex, and subsequently, the 43S complex / 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 / translational initiation / translation initiation factor activity / ribosome assembly / DNA-(apurinic or apyrimidinic site) lyase / tRNA binding / structural constituent of ribosome / translation / GTPase activity / synapse / GTP binding / RNA binding / cytosol / cytoplasm Similarity search - Function | |||||||||||||||
Biological species | Homo sapiens (human) Hepatitis C virus Oryctolagus cuniculus (rabbit) | |||||||||||||||
Method | ELECTRON MICROSCOPY / single particle reconstruction / cryo EM / Resolution: 3.7 Å | |||||||||||||||
Authors | Brown, Z.P. / Abaeva, I.S. / De, S. / Hellen, C.U.T. / Pestova, T.V. / Frank, J. | |||||||||||||||
Funding support | United States, 4items
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Citation | Journal: EMBO J / Year: 2022 Title: Molecular architecture of 40S translation initiation complexes on the hepatitis C virus IRES. Authors: Zuben P Brown / Irina S Abaeva / Swastik De / Christopher U T Hellen / Tatyana V Pestova / Joachim Frank / Abstract: Hepatitis C virus mRNA contains an internal ribosome entry site (IRES) that mediates end-independent translation initiation, requiring a subset of eukaryotic initiation factors (eIFs). Biochemical ...Hepatitis C virus mRNA contains an internal ribosome entry site (IRES) that mediates end-independent translation initiation, requiring a subset of eukaryotic initiation factors (eIFs). Biochemical studies revealed that direct binding of the IRES to the 40S ribosomal subunit places the initiation codon into the P site, where it base pairs with eIF2-bound Met-tRNAiMet forming a 48S initiation complex. Subsequently, eIF5 and eIF5B mediate subunit joining, yielding an elongation-competent 80S ribosome. Initiation can also proceed without eIF2, in which case Met-tRNAiMet is recruited directly by eIF5B. However, the structures of initiation complexes assembled on the HCV IRES, the transitions between different states, and the accompanying conformational changes have remained unknown. To fill these gaps, we now obtained cryo-EM structures of IRES initiation complexes, at resolutions up to 3.5 Å, that cover all major stages from the initial ribosomal association, through eIF2-containing 48S initiation complexes, to eIF5B-containing complexes immediately prior to subunit joining. These structures provide insights into the dynamic network of 40S/IRES contacts, highlight the role of IRES domain II, and reveal conformational changes that occur during the transition from eIF2- to eIF5B-containing 48S complexes and prepare them for subunit joining. | |||||||||||||||
History |
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-Structure visualization
Structure viewer | Molecule: MolmilJmol/JSmol |
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-Downloads & links
-Download
PDBx/mmCIF format | 7syw.cif.gz | 1.9 MB | Display | PDBx/mmCIF format |
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PDB format | pdb7syw.ent.gz | 1.5 MB | Display | PDB format |
PDBx/mmJSON format | 7syw.json.gz | Tree view | PDBx/mmJSON format | |
Others | Other downloads |
-Validation report
Arichive directory | https://data.pdbj.org/pub/pdb/validation_reports/sy/7syw ftp://data.pdbj.org/pub/pdb/validation_reports/sy/7syw | HTTPS FTP |
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-Related structure data
Related structure data | 25543MC 7syiC 7syjC 7sykC 7sylC 7syoC 7sypC 7syqC 7syrC 7sysC 7sytC 7syuC 7syvC 7syxC M: map data used to model this data C: citing same article (ref.) |
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Similar structure data | Similarity search - Function & homologyF&H Search |
-Links
-Assembly
Deposited unit |
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1 |
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-Components
-RNA chain , 3 types, 3 molecules 2iz
#1: RNA chain | Mass: 603100.938 Da / Num. of mol.: 1 / Source method: isolated from a natural source / Source: (natural) Oryctolagus cuniculus (rabbit) / Plasmid details: rabbit reticulocyte lysate |
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#36: RNA chain | Mass: 24231.510 Da / Num. of mol.: 1 / Source method: isolated from a natural source / Source: (natural) Oryctolagus cuniculus (rabbit) / References: GenBank: 174924 |
#39: RNA chain | Mass: 128746.109 Da / Num. of mol.: 1 Source method: isolated from a genetically manipulated source Source: (gene. exp.) Hepatitis C virus (isolate 1) / Production host: Escherichia coli (E. coli) / References: GenBank: 149384897 |
-Eukaryotic translation initiation factor ... , 2 types, 2 molecules Ax
#2: Protein | Mass: 16488.449 Da / Num. of mol.: 1 Source method: isolated from a genetically manipulated source Source: (gene. exp.) Homo sapiens (human) / Gene: EIF1AX, EIF1A, EIF4C / Production host: Escherichia coli (E. coli) / References: UniProt: P47813 |
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#38: Protein | Mass: 70788.000 Da / Num. of mol.: 1 / Source method: isolated from a natural source / Source: (natural) Oryctolagus cuniculus (rabbit) / References: UniProt: G1TRL5 |
-40S ribosomal protein ... , 9 types, 9 molecules BDFIWZabg
#3: Protein | Mass: 32958.016 Da / Num. of mol.: 1 / Source method: isolated from a natural source / Source: (natural) Oryctolagus cuniculus (rabbit) / Plasmid details: rabbit reticulocyte lysate / References: UniProt: G1TLT8 |
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#5: Protein | Mass: 24441.846 Da / Num. of mol.: 1 / Source method: isolated from a natural source / Source: (natural) Oryctolagus cuniculus (rabbit) / Plasmid details: rabbit reticulocyte lysate |
#7: Protein | Mass: 29654.869 Da / Num. of mol.: 1 / Source method: isolated from a natural source / Source: (natural) Oryctolagus cuniculus (rabbit) / References: UniProt: G1TK17 |
#10: Protein | Mass: 47356.172 Da / Num. of mol.: 1 / Source method: isolated from a natural source / Source: (natural) Oryctolagus cuniculus (rabbit) / References: UniProt: G1SVB0 |
#24: Protein | Mass: 9124.389 Da / Num. of mol.: 1 / Source method: isolated from a natural source / Source: (natural) Oryctolagus cuniculus (rabbit) / References: UniProt: G1TM82 |
#27: Protein | Mass: 15237.104 Da / Num. of mol.: 1 / Source method: isolated from a natural source / Source: (natural) Oryctolagus cuniculus (rabbit) / References: UniProt: A0A5F9CNN4 |
#28: Protein | Mass: 13645.028 Da / Num. of mol.: 1 / Source method: isolated from a natural source / Source: (natural) Oryctolagus cuniculus (rabbit) / References: UniProt: G1TDB3 |
#29: Protein | Mass: 11645.794 Da / Num. of mol.: 1 / Source method: isolated from a natural source / Source: (natural) Oryctolagus cuniculus (rabbit) |
#34: Protein | Mass: 21431.742 Da / Num. of mol.: 1 / Source method: isolated from a natural source / Source: (natural) Oryctolagus cuniculus (rabbit) / References: UniProt: G1SK22 |
+Protein , 24 types, 24 molecules CEGHJKLMNOPQRSTUVXYcdefh
-Protein/peptide , 1 types, 1 molecules n
#37: Protein/peptide | Mass: 3473.451 Da / Num. of mol.: 1 / Source method: isolated from a natural source / Source: (natural) Oryctolagus cuniculus (rabbit) / References: UniProt: A0A087WNH4 |
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-Non-polymers , 4 types, 4 molecules
#40: Chemical | ChemComp-ZN / |
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#41: Chemical | ChemComp-GTP / |
#42: Chemical | ChemComp-MG / |
#43: Chemical | ChemComp-NA / |
-Details
Has ligand of interest | N |
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-Experimental details
-Experiment
Experiment | Method: ELECTRON MICROSCOPY |
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EM experiment | Aggregation state: PARTICLE / 3D reconstruction method: single particle reconstruction |
-Sample preparation
Component | Name: 40S ribosomal small subunit with HCV IRES / Type: RIBOSOME / Entity ID: #1-#28, #30-#39 / Source: MULTIPLE SOURCES |
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Molecular weight | Value: 2 MDa / Experimental value: NO |
Source (natural) | Organism: Oryctolagus cuniculus (rabbit) |
Buffer solution | pH: 7.5 |
Specimen | Conc.: 7.5E-5 mg/ml / Embedding applied: NO / Shadowing applied: NO / Staining applied: NO / Vitrification applied: YES |
Specimen support | Details: H2/O2 mixture for 25 seconds at 25W power / Grid material: GOLD / Grid mesh size: 300 divisions/in. / Grid type: Quantifoil R0.6/1 |
Vitrification | Instrument: FEI VITROBOT MARK IV / Cryogen name: ETHANE-PROPANE / Humidity: 100 % / Chamber temperature: 277.15 K / Details: 4 second blot time, force 3 |
-Electron microscopy imaging
Experimental equipment | Model: Tecnai F30 / Image courtesy: FEI Company |
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Microscopy | Model: FEI TECNAI F30 |
Electron gun | Electron source: FIELD EMISSION GUN / Accelerating voltage: 300 kV / Illumination mode: FLOOD BEAM |
Electron lens | Mode: BRIGHT FIELDBright-field microscopy / Nominal magnification: 52000 X / Nominal defocus max: 2000 nm / Nominal defocus min: 1000 nm / Cs: 2.26 mm / Alignment procedure: COMA FREE |
Specimen holder | Cryogen: NITROGEN |
Image recording | Average exposure time: 4 sec. / Electron dose: 70.9 e/Å2 / Film or detector model: GATAN K3 (6k x 4k) |
Image scans | Sampling size: 5 µm |
-Processing
EM software |
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CTF correction | Type: PHASE FLIPPING AND AMPLITUDE CORRECTION | ||||||||||||||||||||||||||||||||
Symmetry | Point symmetry: C1 (asymmetric) | ||||||||||||||||||||||||||||||||
3D reconstruction | Resolution: 3.7 Å / Resolution method: FSC 0.143 CUT-OFF / Num. of particles: 133782 / Symmetry type: POINT | ||||||||||||||||||||||||||||||||
Atomic model building |
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