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Yorodumi- EMDB-3040: Cryo-EM structure of a mammalian ribosomal termination complex wi... -
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-Basic information
Entry | Database: EMDB / ID: EMD-3040 | |||||||||
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Title | Cryo-EM structure of a mammalian ribosomal termination complex with ABCE1, eRF1(AAQ) and the UGA stop codon | |||||||||
Map data | Reconstruction of a mammalian 80S ribosome-nascent chain complex containing the UGA stop codon bound to eRF1(AAQ) and ABCE1 | |||||||||
Sample |
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Keywords | ribosome / translation termination / release factor / stop codon / decoding | |||||||||
Function / homology | Function and homology information negative regulation of endoribonuclease activity / CTPase activity / translation termination factor activity / cytoplasmic translational termination / translation release factor complex / OAS antiviral response / regulation of translational termination / translation release factor activity / ribosome disassembly / protein methylation ...negative regulation of endoribonuclease activity / CTPase activity / translation termination factor activity / cytoplasmic translational termination / translation release factor complex / OAS antiviral response / regulation of translational termination / translation release factor activity / ribosome disassembly / protein methylation / translation release factor activity, codon specific / sequence-specific mRNA binding / ribosomal subunit / aminoacyl-tRNA hydrolase activity / regulation of G1 to G0 transition / oxidized pyrimidine DNA binding / response to TNF agonist / positive regulation of base-excision repair / nuclear-transcribed mRNA catabolic process, nonsense-mediated decay / protein tyrosine kinase inhibitor activity / positive regulation of intrinsic apoptotic signaling pathway in response to DNA damage by p53 class mediator / regulation of translation involved in cellular response to UV / positive regulation of intrinsic apoptotic signaling pathway in response to DNA damage / protein-DNA complex disassembly / positive regulation of DNA damage response, signal transduction by p53 class mediator resulting in transcription of p21 class mediator / IRE1-RACK1-PP2A complex / positive regulation of Golgi to plasma membrane protein transport / negative regulation of DNA repair / oxidized purine DNA binding / G1 to G0 transition / supercoiled DNA binding / negative regulation of intrinsic apoptotic signaling pathway in response to hydrogen peroxide / NF-kappaB complex / negative regulation of phagocytosis / ubiquitin-like protein conjugating enzyme binding / ribosomal subunit export from nucleus / Protein hydroxylation / protein kinase A binding / positive regulation of signal transduction by p53 class mediator / ubiquitin ligase inhibitor activity / ribosomal small subunit binding / Eukaryotic Translation Termination / phagocytic cup / positive regulation of mitochondrial depolarization / positive regulation of T cell receptor signaling pathway / positive regulation of activated T cell proliferation / Nonsense Mediated Decay (NMD) independent of the Exon Junction Complex (EJC) / BH3 domain binding / endoribonuclease inhibitor activity / cysteine-type endopeptidase activator activity involved in apoptotic process / ribosomal small subunit export from nucleus / Nonsense Mediated Decay (NMD) enhanced by the Exon Junction Complex (EJC) / translation regulator activity / cellular response to actinomycin D / translational termination / negative regulation of phosphatidylinositol 3-kinase/protein kinase B signal transduction / rough endoplasmic reticulum / gastrulation / spindle assembly / signaling adaptor activity / MDM2/MDM4 family protein binding / translational initiation / : / rescue of stalled ribosome / negative regulation of smoothened signaling pathway / maturation of LSU-rRNA from tricistronic rRNA transcript (SSU-rRNA, 5.8S rRNA, LSU-rRNA) / negative regulation of ubiquitin-dependent protein catabolic process / cytosolic ribosome / negative regulation of peptidyl-serine phosphorylation / DNA damage response, signal transduction by p53 class mediator resulting in cell cycle arrest / positive regulation of intrinsic apoptotic signaling pathway / class I DNA-(apurinic or apyrimidinic site) endonuclease activity / DNA-(apurinic or apyrimidinic site) lyase / positive regulation of microtubule polymerization / ribosomal large subunit biogenesis / ribonucleoside triphosphate phosphatase activity / negative regulation of protein ubiquitination / Hsp70 protein binding / positive regulation of interleukin-2 production / cellular response to leukemia inhibitory factor / small-subunit processome / SH2 domain binding / cyclin binding / DNA endonuclease activity / positive regulation of translation / protein kinase C binding / : / Hydrolases; Acting on acid anhydrides; Acting on GTP to facilitate cellular and subcellular movement / cellular response to glucose stimulus / positive regulation of protein-containing complex assembly / RNA polymerase II transcription regulatory region sequence-specific DNA binding / Hsp90 protein binding / base-excision repair / cellular response to gamma radiation / mRNA 5'-UTR binding / ribosomal small subunit biogenesis / Regulation of expression of SLITs and ROBOs / cytoplasmic ribonucleoprotein granule / transcription coactivator binding / negative regulation of cell growth Similarity search - Function | |||||||||
Biological species | Oryctolagus cuniculus (rabbit) / Homo sapiens (human) | |||||||||
Method | single particle reconstruction / cryo EM / Resolution: 3.83 Å | |||||||||
Authors | Brown A / Shao S / Murray J / Hegde RS / Ramakrishnan V | |||||||||
Citation | Journal: Nature / Year: 2015 Title: Structural basis for stop codon recognition in eukaryotes. Authors: Alan Brown / Sichen Shao / Jason Murray / Ramanujan S Hegde / V Ramakrishnan / Abstract: Termination of protein synthesis occurs when a translating ribosome encounters one of three universally conserved stop codons: UAA, UAG or UGA. Release factors recognize stop codons in the ribosomal ...Termination of protein synthesis occurs when a translating ribosome encounters one of three universally conserved stop codons: UAA, UAG or UGA. Release factors recognize stop codons in the ribosomal A-site to mediate release of the nascent chain and recycling of the ribosome. Bacteria decode stop codons using two separate release factors with differing specificities for the second and third bases. By contrast, eukaryotes rely on an evolutionarily unrelated omnipotent release factor (eRF1) to recognize all three stop codons. The molecular basis of eRF1 discrimination for stop codons over sense codons is not known. Here we present cryo-electron microscopy (cryo-EM) structures at 3.5-3.8 Å resolution of mammalian ribosomal complexes containing eRF1 interacting with each of the three stop codons in the A-site. Binding of eRF1 flips nucleotide A1825 of 18S ribosomal RNA so that it stacks on the second and third stop codon bases. This configuration pulls the fourth position base into the A-site, where it is stabilized by stacking against G626 of 18S rRNA. Thus, eRF1 exploits two rRNA nucleotides also used during transfer RNA selection to drive messenger RNA compaction. In this compacted mRNA conformation, stop codons are favoured by a hydrogen-bonding network formed between rRNA and essential eRF1 residues that constrains the identity of the bases. These results provide a molecular framework for eukaryotic stop codon recognition and have implications for future studies on the mechanisms of canonical and premature translation termination. | |||||||||
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_3040.map.gz | 12.8 MB | EMDB map data format | |
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Header (meta data) | emd-3040-v30.xml emd-3040.xml | 15.2 KB 15.2 KB | Display Display | EMDB header |
FSC (resolution estimation) | emd_3040_fsc.xml | 14.4 KB | Display | FSC data file |
Images | emd_3040.tif | 343.3 KB | ||
Archive directory | http://ftp.pdbj.org/pub/emdb/structures/EMD-3040 ftp://ftp.pdbj.org/pub/emdb/structures/EMD-3040 | HTTPS FTP |
-Related structure data
Related structure data | 3jaiMC 3038C 3039C 3jagC 3jahC C: citing same article (ref.) 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_3040.map.gz / Format: CCP4 / Size: 276 MB / Type: IMAGE STORED AS FLOATING POINT NUMBER (4 BYTES) | ||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||
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Annotation | Reconstruction of a mammalian 80S ribosome-nascent chain complex containing the UGA stop codon bound to eRF1(AAQ) and ABCE1 | ||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||
Voxel size | X=Y=Z: 1.34 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 : Affinity-purified 80S ribosome-nascent chain complex containing t...
Entire | Name: Affinity-purified 80S ribosome-nascent chain complex containing the UGA stop codon bound to eRF1(AAQ) and ABCE1 |
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Components |
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-Supramolecule #1000: Affinity-purified 80S ribosome-nascent chain complex containing t...
Supramolecule | Name: Affinity-purified 80S ribosome-nascent chain complex containing the UGA stop codon bound to eRF1(AAQ) and ABCE1 type: sample / ID: 1000 / Number unique components: 6 |
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Molecular weight | Theoretical: 2.1 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: Oryctolagus cuniculus (rabbit) / synonym: rabbit / Cell: reticulocyte / Location in cell: cytosol |
Molecular weight | Theoretical: 2 MDa |
-Macromolecule #1: eukaryotic release factor 1, G183A, G184A
Macromolecule | Name: eukaryotic release factor 1, G183A, G184A / type: protein_or_peptide / ID: 1 / Name.synonym: eRF1(AAQ) / Number of copies: 1 / Recombinant expression: Yes |
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Source (natural) | Organism: Homo sapiens (human) / synonym: Human |
Recombinant expression | Organism: Escherichia coli BL21(DE3) (bacteria) / Recombinant plasmid: pRSETA |
Sequence | UniProtKB: Eukaryotic peptide chain release factor subunit 1 |
-Macromolecule #2: ATP binding cassette E1
Macromolecule | Name: ATP binding cassette E1 / type: protein_or_peptide / ID: 2 / Name.synonym: ABCE1, Rli1 / Recombinant expression: No |
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Source (natural) | Organism: Oryctolagus cuniculus (rabbit) / synonym: rabbit / Location in cell: cytosol |
Sequence | UniProtKB: ATP-binding cassette sub-family E member 1 |
-Macromolecule #4: Sec61-beta
Macromolecule | Name: Sec61-beta / type: protein_or_peptide / ID: 4 / Details: in vitro translated peptide sequence / Number of copies: 1 / Recombinant expression: No |
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Source (natural) | Organism: Homo sapiens (human) / synonym: Human |
-Macromolecule #3: messenger RNA
Macromolecule | Name: messenger RNA / type: rna / ID: 3 / Name.synonym: mRNA Details: in vitro transcribed mRNA sequence containing UAA stop codon Classification: OTHER / Structure: SINGLE STRANDED / Synthetic?: No |
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Source (natural) | Organism: Homo sapiens (human) / synonym: Human |
Sequence | String: UCAAAGUUUG AG |
-Macromolecule #5: transfer RNA
Macromolecule | Name: transfer RNA / type: rna / ID: 5 / Name.synonym: tRNA / Classification: OTHER / Structure: OTHER / Synthetic?: No |
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Source (natural) | Organism: Oryctolagus cuniculus (rabbit) / synonym: rabbit |
Molecular weight | Theoretical: 20 KDa |
-Experimental details
-Structure determination
Method | cryo EM |
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Processing | single particle reconstruction |
Aggregation state | particle |
-Sample preparation
Buffer | pH: 7.4 / Details: 50 mM Hepes, 100 mM KAc, 5 mM MgAc2, 1 mM DTT |
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Grid | Details: R2/2 400 mesh Cu grids with thin continuous carbon support, glow discharged |
Vitrification | Cryogen name: ETHANE / Chamber humidity: 100 % / Instrument: FEI VITROBOT MARK III / Method: 30 sec wait time, blot for 3 sec before plunging |
-Electron microscopy
Microscope | FEI TITAN KRIOS |
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Electron beam | Acceleration voltage: 300 kV / Electron source: FIELD EMISSION GUN |
Electron optics | Calibrated magnification: 104478 / Illumination mode: FLOOD BEAM / Imaging mode: BRIGHT FIELDBright-field microscopy / Cs: 2.7 mm / Nominal defocus max: 3.6 µm / Nominal defocus min: 1.7 µm / Nominal magnification: 59000 |
Specialist optics | Energy filter - Name: FEI |
Sample stage | Specimen holder model: FEI TITAN KRIOS AUTOGRID HOLDER |
Details | Automated data acquisition using EPU (FEI) |
Date | Apr 17, 2015 |
Image recording | Category: CCD / Film or detector model: FEI FALCON II (4k x 4k) / Number real images: 1472 / Average electron dose: 30 e/Å2 Details: Every image is the average of 17 frames recorded by the direct electron detector |
Experimental equipment | Model: Titan Krios / Image courtesy: FEI Company |
-Image processing
-Atomic model buiding 1
Initial model | PDB ID: |
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Software | Name: Chimera, Coot |
Refinement | Space: RECIPROCAL / Protocol: FLEXIBLE FIT |
Output model | PDB-3jai: |
-Atomic model buiding 2
Initial model | PDB ID: |
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Software | Name: Chimera, Coot |
Refinement | Space: RECIPROCAL / Protocol: FLEXIBLE FIT |
Output model | PDB-3jai: |
-Atomic model buiding 3
Initial model | PDB ID: |
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Software | Name: Chimera, Coot |
Refinement | Space: RECIPROCAL / Protocol: FLEXIBLE FIT |
Output model | PDB-3jai: |
-Atomic model buiding 4
Initial model | PDB ID: |
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Software | Name: Chimera, Coot |
Details | Sequence was modified in Coot to agree with rabbit sequence |
Refinement | Space: RECIPROCAL / Protocol: FLEXIBLE FIT |
Output model | PDB-3jai: |
-Atomic model buiding 5
Initial model | PDB ID: |
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Software | Name: Chimera, Coot |
Details | Sequence was modified in Coot to agree with the most prevalent tRNA sequence for each particular codon |
Refinement | Space: RECIPROCAL / Protocol: FLEXIBLE FIT |
Output model | PDB-3jai: |