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Yorodumi- EMDB-6667: Mechanistic insights into the alternative translation termination... -
+Open data
-Basic information
Entry | Database: EMDB / ID: EMD-6667 | |||||||||
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Title | Mechanistic insights into the alternative translation termination by ArfA and RF2 | |||||||||
Map data | ||||||||||
Sample |
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Keywords | ribosome / translation / RF2 / Arfa / non-stop mRNA / ribosome rescue | |||||||||
Function / homology | Function and homology information translation release factor activity, codon specific / stringent response / ribosomal large subunit binding / ornithine decarboxylase inhibitor activity / transcription antitermination factor activity, RNA binding / misfolded RNA binding / Group I intron splicing / RNA folding / transcriptional attenuation / endoribonuclease inhibitor activity ...translation release factor activity, codon specific / stringent response / ribosomal large subunit binding / ornithine decarboxylase inhibitor activity / transcription antitermination factor activity, RNA binding / misfolded RNA binding / Group I intron splicing / RNA folding / transcriptional attenuation / endoribonuclease inhibitor activity / RNA-binding transcription regulator activity / positive regulation of ribosome biogenesis / negative regulation of cytoplasmic translation / four-way junction DNA binding / translational termination / DnaA-L2 complex / translation repressor activity / negative regulation of DNA-templated DNA replication initiation / negative regulation of translational initiation / regulation of mRNA stability / mRNA regulatory element binding translation repressor activity / rescue of stalled ribosome / ribosome assembly / positive regulation of RNA splicing / assembly of large subunit precursor of preribosome / transcription elongation factor complex / regulation of DNA-templated transcription elongation / cytosolic ribosome assembly / DNA endonuclease activity / response to reactive oxygen species / transcription antitermination / regulation of cell growth / DNA-templated transcription termination / maintenance of translational fidelity / response to radiation / mRNA 5'-UTR binding / ribosomal small subunit biogenesis / large ribosomal subunit / small ribosomal subunit rRNA binding / ribosome biogenesis / ribosome binding / regulation of translation / ribosomal small subunit assembly / ribosomal large subunit assembly / small ribosomal subunit / transferase activity / large ribosomal subunit rRNA binding / 5S rRNA binding / cytosolic small ribosomal subunit / cytoplasmic translation / cytosolic large ribosomal subunit / tRNA binding / molecular adaptor activity / negative regulation of translation / rRNA binding / ribosome / structural constituent of ribosome / viral translational frameshifting / translation / response to antibiotic / negative regulation of DNA-templated transcription / mRNA binding / DNA binding / RNA binding / zinc ion binding / membrane / cytosol / cytoplasm Similarity search - Function | |||||||||
Biological species | Escherichia coli (E. coli) / Escherichia coli K-12 (bacteria) | |||||||||
Method | single particle reconstruction / cryo EM / Resolution: 3.01 Å | |||||||||
Authors | Ma C / Kurita D | |||||||||
Citation | Journal: Nature / Year: 2017 Title: Mechanistic insights into the alternative translation termination by ArfA and RF2. Authors: Chengying Ma / Daisuke Kurita / Ningning Li / Yan Chen / Hyouta Himeno / Ning Gao / Abstract: During cellular translation of messenger RNAs by ribosomes, the translation apparatus sometimes pauses or stalls at the elongation and termination steps. With the exception of programmed stalling, ...During cellular translation of messenger RNAs by ribosomes, the translation apparatus sometimes pauses or stalls at the elongation and termination steps. With the exception of programmed stalling, which is usually used by cells for regulatory purposes, ribosomes stalled on mRNAs need to be terminated and recycled to maintain adequate translation capacity. Much ribosome stalling originates in aberrant mRNAs that lack a stop codon. Transcriptional errors, misprocessing of primary transcripts, and undesired mRNA cleavage all contribute to the formation of non-stop mRNAs. Ribosomes stalled at the 3' end of non-stop mRNAs do not undergo normal termination owing to the lack of specific stop-codon recognition by canonical peptide release factors at the A-site decoding centre. In bacteria, the transfer-messenger RNA (tmRNA)-SmpB-mediated trans-translation rescue system reroutes stalled ribosomes to the normal elongation cycle and translation termination. Two additional rescue systems, ArfA-RF2 (refs 13, 14, 15, 16) and ArfB (formerly known as YaeJ), are also present in many bacterial species, but their mechanisms are not fully understood. Here, using cryo-electron microscopy, we characterize the structure of the Escherichia coli 70S ribosome bound with ArfA, the release factor RF2, a short non-stop mRNA and a cognate P-site tRNA. The C-terminal loop of ArfA occupies the mRNA entry channel on the 30S subunit, whereas its N terminus is sandwiched between the decoding centre and the switch loop of RF2, leading to marked conformational changes in both the decoding centre and RF2. Despite the distinct conformation of RF2, its conserved catalytic GGQ motif is precisely positioned next to the CCA-end of the P-site tRNA. These data illustrate a stop-codon surrogate mechanism for ArfA in facilitating the termination of non-stop ribosomal complexes by RF2. | |||||||||
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_6667.map.gz | 16.4 MB | EMDB map data format | |
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Header (meta data) | emd-6667-v30.xml emd-6667.xml | 77.6 KB 77.6 KB | Display Display | EMDB header |
Images | emd_6667.png | 72.4 KB | ||
Filedesc metadata | emd-6667.cif.gz | 13.3 KB | ||
Archive directory | http://ftp.pdbj.org/pub/emdb/structures/EMD-6667 ftp://ftp.pdbj.org/pub/emdb/structures/EMD-6667 | HTTPS FTP |
-Validation report
Summary document | emd_6667_validation.pdf.gz | 478.6 KB | Display | EMDB validaton report |
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Full document | emd_6667_full_validation.pdf.gz | 478.2 KB | Display | |
Data in XML | emd_6667_validation.xml.gz | 7 KB | Display | |
Data in CIF | emd_6667_validation.cif.gz | 8 KB | Display | |
Arichive directory | https://ftp.pdbj.org/pub/emdb/validation_reports/EMD-6667 ftp://ftp.pdbj.org/pub/emdb/validation_reports/EMD-6667 | HTTPS FTP |
-Related structure data
Related structure data | 5h5uMC M: atomic model generated by this map C: citing same article (ref.) |
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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_6667.map.gz / Format: CCP4 / Size: 178 MB / Type: IMAGE STORED AS FLOATING POINT NUMBER (4 BYTES) | ||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||
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Projections & slices | Image control
Images are generated by Spider. | ||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||
Voxel size | X=Y=Z: 1.08 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 : 70S-ArfA-RF2 complex
+Supramolecule #1: 70S-ArfA-RF2 complex
+Supramolecule #2: Rf2-ArfA
+Supramolecule #3: rRNA
+Macromolecule #1: 30S ribosomal protein S20
+Macromolecule #2: 30S ribosomal protein S21
+Macromolecule #3: Alternative ribosome-rescue factor A
+Macromolecule #4: Peptide chain release factor 2
+Macromolecule #9: 50S ribosomal protein L2
+Macromolecule #10: 50S ribosomal protein L3
+Macromolecule #11: 50S ribosomal protein L4
+Macromolecule #12: 50S ribosomal protein L5
+Macromolecule #13: 50S ribosomal protein L6
+Macromolecule #14: 50S ribosomal protein L9
+Macromolecule #15: 50S ribosomal protein L10
+Macromolecule #16: 50S ribosomal protein L11
+Macromolecule #17: 50S ribosomal protein L13
+Macromolecule #18: 50S ribosomal protein L14
+Macromolecule #19: 50S ribosomal protein L15
+Macromolecule #20: 50S ribosomal protein L16
+Macromolecule #21: 50S ribosomal protein L17
+Macromolecule #22: 50S ribosomal protein L18
+Macromolecule #23: 50S ribosomal protein L19
+Macromolecule #24: 50S ribosomal protein L20
+Macromolecule #25: 50S ribosomal protein L21
+Macromolecule #26: 50S ribosomal protein L22
+Macromolecule #27: 50S ribosomal protein L23
+Macromolecule #28: 50S ribosomal protein L24
+Macromolecule #29: 50S ribosomal protein L25
+Macromolecule #30: 50S ribosomal protein L27
+Macromolecule #31: 50S ribosomal protein L28
+Macromolecule #32: 50S ribosomal protein L29
+Macromolecule #33: 50S ribosomal protein L30
+Macromolecule #34: 50S ribosomal protein L32
+Macromolecule #35: 50S ribosomal protein L33
+Macromolecule #36: 50S ribosomal protein L34
+Macromolecule #37: 50S ribosomal protein L35
+Macromolecule #38: 50S ribosomal protein L36
+Macromolecule #40: 30S ribosomal protein S2
+Macromolecule #41: 30S ribosomal protein S3
+Macromolecule #42: 30S ribosomal protein S4
+Macromolecule #43: 30S ribosomal protein S5
+Macromolecule #44: 30S ribosomal protein S6
+Macromolecule #45: 30S ribosomal protein S7
+Macromolecule #46: 30S ribosomal protein S8
+Macromolecule #47: 30S ribosomal protein S9
+Macromolecule #48: 30S ribosomal protein S10
+Macromolecule #49: 30S ribosomal protein S11
+Macromolecule #50: 30S ribosomal protein S12
+Macromolecule #51: 30S ribosomal protein S13
+Macromolecule #52: 30S ribosomal protein S14
+Macromolecule #53: 30S ribosomal protein S15
+Macromolecule #54: 30S ribosomal protein S16
+Macromolecule #55: 30S ribosomal protein S17
+Macromolecule #56: 30S ribosomal protein S18
+Macromolecule #57: 30S ribosomal protein S19
+Macromolecule #5: P-site tRNA
+Macromolecule #6: mRNA
+Macromolecule #7: 23S rRNA
+Macromolecule #8: 5S Ribosomal RNA
+Macromolecule #39: 16S rRNA
-Experimental details
-Structure determination
Method | cryo EM |
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Processing | single particle reconstruction |
Aggregation state | particle |
-Sample preparation
Buffer | pH: 7.6 |
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Grid | Material: COPPER / Support film - Material: CARBON / Support film - topology: HOLEY / Pretreatment - Type: GLOW DISCHARGE / Pretreatment - Time: 30 sec. |
Vitrification | Cryogen name: ETHANE / Chamber humidity: 100 % / Chamber temperature: 277.15 K |
-Electron microscopy
Microscope | FEI TITAN KRIOS |
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Image recording | Film or detector model: FEI FALCON II (4k x 4k) / Detector mode: INTEGRATING / Average electron dose: 2.0 e/Å2 |
Electron beam | Acceleration voltage: 300 kV / Electron source: FIELD EMISSION GUN |
Electron optics | Illumination mode: FLOOD BEAM / Imaging mode: BRIGHT FIELD |
Sample stage | Specimen holder model: FEI TITAN KRIOS AUTOGRID HOLDER / Cooling holder cryogen: NITROGEN |
Experimental equipment | Model: Titan Krios / Image courtesy: FEI Company |
-Image processing
Startup model | Type of model: OTHER |
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Final reconstruction | Number classes used: 8 / Resolution.type: BY AUTHOR / Resolution: 3.01 Å / Resolution method: FSC 0.143 CUT-OFF / Software - Name: RELION (ver. 1.3) / Number images used: 13693 |
Initial angle assignment | Type: PROJECTION MATCHING |
Final angle assignment | Type: PROJECTION MATCHING |