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Yorodumi- PDB-5h5u: Mechanistic insights into the alternative translation termination... -
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-Basic information
Entry | Database: PDB / ID: 5h5u | ||||||
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Title | Mechanistic insights into the alternative translation termination by ArfA and RF2 | ||||||
Components |
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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 translational initiation / negative regulation of DNA-templated DNA replication initiation / regulation of mRNA stability / rescue of stalled ribosome / mRNA regulatory element binding translation repressor activity / ribosome assembly / positive regulation of RNA splicing / assembly of large subunit precursor of preribosome / transcription elongation factor complex / cytosolic ribosome assembly / regulation of DNA-templated transcription elongation / DNA endonuclease activity / ribosomal large subunit assembly / transcription antitermination / response to reactive oxygen species / regulation of cell growth / DNA-templated transcription termination / maintenance of translational fidelity / response to radiation / mRNA 5'-UTR binding / large ribosomal subunit / ribosome biogenesis / ribosome binding / regulation of translation / ribosomal small subunit biogenesis / ribosomal small subunit assembly / small ribosomal subunit / small ribosomal subunit rRNA binding / transferase activity / 5S rRNA binding / large ribosomal subunit rRNA binding / cytosolic small ribosomal subunit / cytosolic large ribosomal subunit / cytoplasmic translation / tRNA binding / molecular adaptor activity / rRNA binding / negative regulation of translation / ribosome / structural constituent of ribosome / translation / response to antibiotic / negative regulation of DNA-templated transcription / mRNA binding / DNA binding / RNA binding / zinc ion binding / membrane / cytoplasm / cytosol Similarity search - Function | ||||||
Biological species | Escherichia coli K-12 (bacteria) Escherichia coli (E. coli) | ||||||
Method | ELECTRON MICROSCOPY / single particle reconstruction / cryo EM / Resolution: 3.01 Å | ||||||
Authors | Ma, C. / Kurita, D. / Li, N. / Chen, Y. / Himeno, H. / Gao, N. | ||||||
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 | Molecule: MolmilJmol/JSmol |
-Downloads & links
-Download
PDBx/mmCIF format | 5h5u.cif.gz | 3.7 MB | Display | PDBx/mmCIF format |
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PDB format | pdb5h5u.ent.gz | Display | PDB format | |
PDBx/mmJSON format | 5h5u.json.gz | Tree view | PDBx/mmJSON format | |
Others | Other downloads |
-Validation report
Summary document | 5h5u_validation.pdf.gz | 1.2 MB | Display | wwPDB validaton report |
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Full document | 5h5u_full_validation.pdf.gz | 1.3 MB | Display | |
Data in XML | 5h5u_validation.xml.gz | 200 KB | Display | |
Data in CIF | 5h5u_validation.cif.gz | 364 KB | Display | |
Arichive directory | https://data.pdbj.org/pub/pdb/validation_reports/h5/5h5u ftp://data.pdbj.org/pub/pdb/validation_reports/h5/5h5u | HTTPS FTP |
-Related structure data
Related structure data | 6667MC M: map data used to model this data C: citing same article (ref.) |
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Similar structure data |
-Links
-Assembly
Deposited unit |
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-Components
-30S ribosomal protein ... , 20 types, 20 molecules 12ijklmnopqrstuvwxyz
#1: Protein | Mass: 9577.268 Da / Num. of mol.: 1 / Source method: isolated from a natural source / Source: (natural) Escherichia coli K-12 (bacteria) / Strain: K-12 / References: UniProt: P0A7U7 |
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#2: Protein | Mass: 8392.844 Da / Num. of mol.: 1 / Source method: isolated from a natural source / Source: (natural) Escherichia coli K-12 (bacteria) / Strain: K-12 / References: UniProt: P68679 |
#40: Protein | Mass: 26650.475 Da / Num. of mol.: 1 / Source method: isolated from a natural source / Source: (natural) Escherichia coli K-12 (bacteria) / Strain: K-12 / References: UniProt: P0A7V0 |
#41: Protein | Mass: 25900.117 Da / Num. of mol.: 1 / Source method: isolated from a natural source / Source: (natural) Escherichia coli K-12 (bacteria) / Strain: K-12 / References: UniProt: P0A7V3 |
#42: Protein | Mass: 23383.002 Da / Num. of mol.: 1 / Source method: isolated from a natural source / Source: (natural) Escherichia coli K-12 (bacteria) / Strain: K-12 / References: UniProt: P0A7V8 |
#43: Protein | Mass: 17498.203 Da / Num. of mol.: 1 / Source method: isolated from a natural source / Source: (natural) Escherichia coli K-12 (bacteria) / Strain: K-12 / References: UniProt: P0A7W1 |
#44: Protein | Mass: 15727.512 Da / Num. of mol.: 1 / Source method: isolated from a natural source / Source: (natural) Escherichia coli K-12 (bacteria) / Strain: K-12 / References: UniProt: P02358 |
#45: Protein | Mass: 19923.959 Da / Num. of mol.: 1 / Source method: isolated from a natural source / Source: (natural) Escherichia coli K-12 (bacteria) / Strain: K-12 / References: UniProt: P02359 |
#46: Protein | Mass: 14015.361 Da / Num. of mol.: 1 / Source method: isolated from a natural source / Source: (natural) Escherichia coli K-12 (bacteria) / Strain: K-12 / References: UniProt: P0A7W7 |
#47: Protein | Mass: 14755.074 Da / Num. of mol.: 1 / Source method: isolated from a natural source / Source: (natural) Escherichia coli K-12 (bacteria) / Strain: K-12 / References: UniProt: P0A7X3 |
#48: Protein | Mass: 11755.597 Da / Num. of mol.: 1 / Source method: isolated from a natural source / Source: (natural) Escherichia coli K-12 (bacteria) / Strain: K-12 / References: UniProt: P0A7R5 |
#49: Protein | Mass: 13739.778 Da / Num. of mol.: 1 / Source method: isolated from a natural source / Source: (natural) Escherichia coli K-12 (bacteria) / Strain: K-12 / References: UniProt: P0A7R9 |
#50: Protein | Mass: 13636.961 Da / Num. of mol.: 1 / Source method: isolated from a natural source / Source: (natural) Escherichia coli K-12 (bacteria) / Strain: K-12 / References: UniProt: P0A7S3 |
#51: Protein | Mass: 12997.271 Da / Num. of mol.: 1 / Source method: isolated from a natural source / Source: (natural) Escherichia coli K-12 (bacteria) / Strain: K-12 / References: UniProt: P0A7S9 |
#52: Protein | Mass: 11475.364 Da / Num. of mol.: 1 / Source method: isolated from a natural source / Source: (natural) Escherichia coli K-12 (bacteria) / Strain: K-12 / References: UniProt: P0AG59 |
#53: Protein | Mass: 10159.621 Da / Num. of mol.: 1 / Source method: isolated from a natural source / Source: (natural) Escherichia coli K-12 (bacteria) / Strain: K-12 / References: UniProt: P0ADZ4 |
#54: Protein | Mass: 9207.572 Da / Num. of mol.: 1 / Source method: isolated from a natural source / Source: (natural) Escherichia coli K-12 (bacteria) / Strain: K-12 / References: UniProt: P0A7T3 |
#55: Protein | Mass: 9593.296 Da / Num. of mol.: 1 / Source method: isolated from a natural source / Source: (natural) Escherichia coli K-12 (bacteria) / Strain: K-12 / References: UniProt: P0AG63 |
#56: Protein | Mass: 8874.276 Da / Num. of mol.: 1 / Source method: isolated from a natural source / Source: (natural) Escherichia coli K-12 (bacteria) / Strain: K-12 / References: UniProt: P0A7T7 |
#57: Protein | Mass: 10324.160 Da / Num. of mol.: 1 / Source method: isolated from a natural source / Source: (natural) Escherichia coli K-12 (bacteria) / Strain: K-12 / References: UniProt: P0A7U3 |
-Protein , 2 types, 2 molecules 34
#3: Protein | Mass: 8190.438 Da / Num. of mol.: 1 / Source method: isolated from a natural source / Source: (natural) Escherichia coli K-12 (bacteria) / Strain: K-12 / References: UniProt: P36675 |
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#4: Protein | Mass: 41300.660 Da / Num. of mol.: 1 / Source method: isolated from a natural source / Source: (natural) Escherichia coli K-12 (bacteria) / Strain: K-12 / References: UniProt: P07012 |
-RNA chain , 5 types, 5 molecules 57ABh
#5: RNA chain | Mass: 24497.602 Da / Num. of mol.: 1 / Source method: obtained synthetically / Source: (synth.) Escherichia coli (E. coli) |
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#6: RNA chain | Mass: 1876.173 Da / Num. of mol.: 1 / Source method: obtained synthetically / Source: (synth.) Escherichia coli (E. coli) |
#7: RNA chain | Mass: 941305.188 Da / Num. of mol.: 1 / Source method: isolated from a natural source / Source: (natural) Escherichia coli K-12 (bacteria) / Strain: K-12 / References: GenBank: 802133627 |
#8: RNA chain | Mass: 38790.090 Da / Num. of mol.: 1 / Source method: isolated from a natural source / Source: (natural) Escherichia coli K-12 (bacteria) / Strain: K-12 / References: GenBank: 1063812051 |
#39: RNA chain | Mass: 497221.594 Da / Num. of mol.: 1 / Source method: isolated from a natural source / Source: (natural) Escherichia coli K-12 (bacteria) / Strain: K-12 / References: GenBank: 802133627 |
+50S ribosomal protein ... , 30 types, 30 molecules CDEFGHIJKLMNOPQRSTUVWXYZabcdef
-Experimental details
-Experiment
Experiment | Method: ELECTRON MICROSCOPY |
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EM experiment | Aggregation state: PARTICLE / 3D reconstruction method: single particle reconstruction |
-Sample preparation
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Source (natural) |
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Source (recombinant) |
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Buffer solution | pH: 7.6 | ||||||||||||||||||||||||
Specimen | Embedding applied: NO / Shadowing applied: NO / Staining applied: NO / Vitrification applied: YES | ||||||||||||||||||||||||
Specimen support | Grid material: COPPER | ||||||||||||||||||||||||
Vitrification | Cryogen name: ETHANE / Humidity: 100 % / Chamber temperature: 277.15 K |
-Electron microscopy imaging
Experimental equipment | Model: Titan Krios / Image courtesy: FEI Company |
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Microscopy | Model: FEI TITAN KRIOS |
Electron gun | Electron source: FIELD EMISSION GUN / Accelerating voltage: 300 kV / Illumination mode: FLOOD BEAM |
Electron lens | Mode: BRIGHT FIELD |
Specimen holder | Cryogen: NITROGEN / Specimen holder model: FEI TITAN KRIOS AUTOGRID HOLDER |
Image recording | Electron dose: 2 e/Å2 / Detector mode: INTEGRATING / Film or detector model: FEI FALCON II (4k x 4k) |
-Processing
Software | Name: REFMAC / Version: 5.8.0088 / Classification: refinement | |||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||
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EM software |
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CTF correction | Type: NONE | |||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||
3D reconstruction | Resolution: 3.01 Å / Resolution method: FSC 0.143 CUT-OFF / Num. of particles: 13693 / Num. of class averages: 8 / Symmetry type: POINT | |||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||
Refinement | Resolution: 3→270 Å / Cor.coef. Fo:Fc: 0.971 / SU B: 9.616 / SU ML: 0.159 / ESU R: 0.249 Stereochemistry target values: MAXIMUM LIKELIHOOD WITH PHASES Details: HYDROGENS HAVE BEEN ADDED IN THE RIDING POSITIONS
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Solvent computation | Ion probe radii: 0.8 Å / Shrinkage radii: 0.8 Å / VDW probe radii: 1.2 Å / Solvent model: MASK | |||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||
Displacement parameters | Biso mean: 144.877 Å2
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Refinement step | Cycle: 1 / Total: 149802 | |||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||
Refine LS restraints |
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