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Yorodumi- PDB-6wnw: Active 70S ribosome without free 5S rRNA and bound with A- and P- tRNA -
+Open data
-Basic information
Entry | Database: PDB / ID: 6wnw | |||||||||||||||
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Title | Active 70S ribosome without free 5S rRNA and bound with A- and P- tRNA | |||||||||||||||
Components |
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Keywords | RIBOSOME / 23S-cp5S rRNA / engineering | |||||||||||||||
Function / homology | Function and homology information negative regulation of cytoplasmic translational initiation / stringent response / mRNA base-pairing translational repressor activity / ornithine decarboxylase inhibitor activity / misfolded RNA binding / transcription antitermination factor activity, RNA binding / Group I intron splicing / RNA folding / transcriptional attenuation / endoribonuclease inhibitor activity ...negative regulation of cytoplasmic translational initiation / stringent response / mRNA base-pairing translational repressor activity / ornithine decarboxylase inhibitor activity / misfolded RNA binding / transcription antitermination factor activity, 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 / translational termination / DnaA-L2 complex / negative regulation of translational initiation / four-way junction DNA binding / translation repressor activity / translational initiation / negative regulation of DNA-templated DNA replication initiation / regulation of mRNA stability / ribosome assembly / mRNA regulatory element binding translation repressor activity / response to reactive oxygen species / assembly of large subunit precursor of preribosome / transcription elongation factor complex / positive regulation of RNA splicing / cytosolic ribosome assembly / regulation of DNA-templated transcription elongation / transcription antitermination / DNA endonuclease activity / regulation of cell growth / DNA-templated transcription termination / maintenance of translational fidelity / : / response to radiation / mRNA 5'-UTR binding / ribosomal small subunit biogenesis / ribosomal small subunit assembly / small ribosomal subunit rRNA binding / cytosolic small ribosomal subunit / large ribosomal subunit rRNA binding / regulation of translation / ribosome biogenesis / ribosome binding / ribosomal large subunit assembly / large ribosomal subunit / 5S rRNA binding / cytosolic large ribosomal subunit / small ribosomal subunit / cytoplasmic translation / transferase activity / tRNA binding / negative regulation of translation / molecular adaptor activity / ribosome / rRNA binding / structural constituent of ribosome / translation / response to antibiotic / mRNA binding / negative regulation of DNA-templated transcription / DNA binding / RNA binding / zinc ion binding / membrane / cytosol / cytoplasm Similarity search - Function | |||||||||||||||
Biological species | Escherichia coli (E. coli) | |||||||||||||||
Method | ELECTRON MICROSCOPY / single particle reconstruction / cryo EM / Resolution: 3.2 Å | |||||||||||||||
Authors | Loveland, A.B. / Korostelev, A.A. / Mankin, A.S. / Huang, S. / Aleksashin, N.A. / Klepacki, D. / Reier, K. / Kefi, A. / Szal, A. / Remme, J. ...Loveland, A.B. / Korostelev, A.A. / Mankin, A.S. / Huang, S. / Aleksashin, N.A. / Klepacki, D. / Reier, K. / Kefi, A. / Szal, A. / Remme, J. / Jaeger, L. / Vazquez-Laslop, N. | |||||||||||||||
Funding support | United States, Estonia, 4items
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Citation | Journal: Nat Commun / Year: 2020 Title: Ribosome engineering reveals the importance of 5S rRNA autonomy for ribosome assembly. Authors: Shijie Huang / Nikolay A Aleksashin / Anna B Loveland / Dorota Klepacki / Kaspar Reier / Amira Kefi / Teresa Szal / Jaanus Remme / Luc Jaeger / Nora Vázquez-Laslop / Andrei A Korostelev / Alexander S Mankin / Abstract: 5S rRNA is an indispensable component of cytoplasmic ribosomes in all species. The functions of 5S rRNA and the reasons for its evolutionary preservation as an independent molecule remain unclear. ...5S rRNA is an indispensable component of cytoplasmic ribosomes in all species. The functions of 5S rRNA and the reasons for its evolutionary preservation as an independent molecule remain unclear. Here we used ribosome engineering to investigate whether 5S rRNA autonomy is critical for ribosome function and cell survival. By linking circularly permutated 5S rRNA with 23S rRNA we generated a bacterial strain devoid of free 5S rRNA. Viability of the engineered cells demonstrates that autonomous 5S rRNA is dispensable for cell growth under standard conditions and is unlikely to have essential functions outside the ribosome. The fully assembled ribosomes carrying 23S-5S rRNA are highly active in translation. However, the engineered cells accumulate aberrant 50S subunits unable to form stable 70S ribosomes. Cryo-EM analysis revealed a malformed peptidyl transferase center in the misassembled 50S subunits. Our results argue that the autonomy of 5S rRNA is preserved due to its role in ribosome biogenesis. | |||||||||||||||
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 | 6wnw.cif.gz | 3.2 MB | Display | PDBx/mmCIF format |
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PDB format | pdb6wnw.ent.gz | Display | PDB format | |
PDBx/mmJSON format | 6wnw.json.gz | Tree view | PDBx/mmJSON format | |
Others | Other downloads |
-Validation report
Arichive directory | https://data.pdbj.org/pub/pdb/validation_reports/wn/6wnw ftp://data.pdbj.org/pub/pdb/validation_reports/wn/6wnw | HTTPS FTP |
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-Related structure data
Related structure data | 21858MC 6wntC 6wnvC 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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1 |
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NMR ensembles |
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-Components
+50S ribosomal protein ... , 32 types, 32 molecules bcdefghijklmnopqrstuvwxyzABCDEFa
-30S ribosomal protein ... , 20 types, 20 molecules GHIJKLMNOPQRSTUVWXYZ
#32: Protein | Mass: 25015.816 Da / Num. of mol.: 1 / Source method: isolated from a natural source / Details: cell / Source: (natural) Escherichia coli (E. coli) / Strain: MRE600 / References: UniProt: P0A7V0 |
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#33: Protein | Mass: 23078.785 Da / Num. of mol.: 1 / Source method: isolated from a natural source / Details: cell / Source: (natural) Escherichia coli (E. coli) / Strain: MRE600 / References: UniProt: P0A7V3 |
#34: Protein | Mass: 23383.002 Da / Num. of mol.: 1 / Source method: isolated from a natural source / Details: cell / Source: (natural) Escherichia coli (E. coli) / Strain: MRE600 / References: UniProt: P0A7V8 |
#35: Protein | Mass: 16532.088 Da / Num. of mol.: 1 / Source method: isolated from a natural source / Details: cell / Source: (natural) Escherichia coli (E. coli) / Strain: MRE600 / References: UniProt: P0A7W1 |
#36: Protein | Mass: 11669.371 Da / Num. of mol.: 1 / Source method: isolated from a natural source / Details: cell / Source: (natural) Escherichia coli (E. coli) / Strain: MRE600 / References: UniProt: P02358 |
#37: Protein | Mass: 16861.523 Da / Num. of mol.: 1 / Source method: isolated from a natural source / Details: cell / Source: (natural) Escherichia coli (E. coli) / Strain: MRE600 / References: UniProt: P02359 |
#38: Protein | Mass: 14015.361 Da / Num. of mol.: 1 / Source method: isolated from a natural source / Details: cell / Source: (natural) Escherichia coli (E. coli) / Strain: MRE600 / References: UniProt: P0A7W7 |
#39: Protein | Mass: 14554.882 Da / Num. of mol.: 1 / Source method: isolated from a natural source / Details: cell / Source: (natural) Escherichia coli (E. coli) / Strain: MRE600 / References: UniProt: P0A7X3 |
#40: Protein | Mass: 11196.988 Da / Num. of mol.: 1 / Source method: isolated from a natural source / Details: cell / Source: (natural) Escherichia coli (E. coli) / Strain: MRE600 / References: UniProt: P0A7R5 |
#41: Protein | Mass: 12388.068 Da / Num. of mol.: 1 / Source method: isolated from a natural source / Details: cell / Source: (natural) Escherichia coli (E. coli) / Strain: MRE600 / References: UniProt: P0A7R9 |
#42: Protein | Mass: 13636.961 Da / Num. of mol.: 1 / Source method: isolated from a natural source / Details: cell / Source: (natural) Escherichia coli (E. coli) / Strain: MRE600 / References: UniProt: P0A7S3 |
#43: Protein | Mass: 12625.753 Da / Num. of mol.: 1 / Source method: isolated from a natural source / Details: cell / Source: (natural) Escherichia coli (E. coli) / Strain: MRE600 / References: UniProt: P0A7S9 |
#44: Protein | Mass: 11475.364 Da / Num. of mol.: 1 / Source method: isolated from a natural source / Details: cell / Source: (natural) Escherichia coli (E. coli) / Strain: MRE600 / References: UniProt: P0AG59 |
#45: Protein | Mass: 10159.621 Da / Num. of mol.: 1 / Source method: isolated from a natural source / Details: cell / Source: (natural) Escherichia coli (E. coli) / Strain: MRE600 / References: UniProt: P0ADZ4 |
#46: Protein | Mass: 9207.572 Da / Num. of mol.: 1 / Source method: isolated from a natural source / Details: cell / Source: (natural) Escherichia coli (E. coli) / Strain: MRE600 / References: UniProt: P0A7T3 |
#47: Protein | Mass: 9263.946 Da / Num. of mol.: 1 / Source method: isolated from a natural source / Details: cell / Source: (natural) Escherichia coli (E. coli) / Strain: MRE600 / References: UniProt: P0AG63 |
#48: Protein | Mass: 7606.768 Da / Num. of mol.: 1 / Source method: isolated from a natural source / Details: cell / Source: (natural) Escherichia coli (E. coli) / Strain: MRE600 / References: UniProt: P0A7T7 |
#49: Protein | Mass: 9057.626 Da / Num. of mol.: 1 / Source method: isolated from a natural source / Details: cell / Source: (natural) Escherichia coli (E. coli) / Strain: MRE600 / References: UniProt: P0A7U3 |
#50: Protein | Mass: 9506.190 Da / Num. of mol.: 1 / Source method: isolated from a natural source / Details: cell / Source: (natural) Escherichia coli (E. coli) / Strain: MRE600 / References: UniProt: P0A7U7 |
#51: Protein | Mass: 7763.073 Da / Num. of mol.: 1 / Source method: isolated from a natural source / Details: cell / Source: (natural) Escherichia coli (E. coli) / Strain: MRE600 / References: UniProt: P68679 |
-RNA chain , 5 types, 5 molecules 34517
#53: RNA chain | Mass: 498725.406 Da / Num. of mol.: 1 / Source method: isolated from a natural source / Details: cell / Source: (natural) Escherichia coli (E. coli) / Strain: MRE600 / References: GenBank: 1126835768 |
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#54: RNA chain | Mass: 982729.812 Da / Num. of mol.: 1 Source method: isolated from a genetically manipulated source Details: Engineered 23S rRNA with inserted circularly permuted 5S rRNA Source: (gene. exp.) Escherichia coli (E. coli) / Strain: MRE600 / Production host: Escherichia coli (E. coli) |
#55: RNA chain | Mass: 24802.785 Da / Num. of mol.: 1 / Source method: isolated from a natural source / Details: cell / Source: (natural) Escherichia coli (E. coli) / Strain: MRE600 / References: GenBank: 1275521620 |
#56: RNA chain | Mass: 3169.956 Da / Num. of mol.: 1 / Source method: isolated from a natural source / Details: cell / Source: (natural) Escherichia coli (E. coli) / Strain: MRE600 |
#57: RNA chain | Mass: 24485.539 Da / Num. of mol.: 1 / Source method: isolated from a natural source / Details: cell / Source: (natural) Escherichia coli (E. coli) / Strain: MRE600 / References: GenBank: 1275521620 |
-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: Active 70S ribosome without free 5S rRNA and bound with A- and P- tRNA Type: RIBOSOME / Entity ID: all / Source: RECOMBINANT |
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Molecular weight | Value: 2.7 MDa / Experimental value: NO |
Source (natural) | Organism: Escherichia coli (E. coli) / Strain: MRE600 |
Source (recombinant) | Organism: Escherichia coli (E. coli) |
Buffer solution | pH: 7 |
Specimen | Conc.: 0.1 mg/ml / Embedding applied: NO / Shadowing applied: NO / Staining applied: NO / Vitrification applied: YES |
Vitrification | Cryogen name: ETHANE |
-Electron microscopy imaging
Microscopy | Model: TFS TALOS |
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Electron gun | Electron source: FIELD EMISSION GUN / Accelerating voltage: 200 kV / Illumination mode: FLOOD BEAM |
Electron lens | Mode: BRIGHT FIELDBright-field microscopy |
Image recording | Electron dose: 30 e/Å2 / Film or detector model: GATAN K3 (6k x 4k) |
-Processing
EM software |
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CTF correction | Type: PHASE FLIPPING ONLY | ||||||||||||||||||||||||||||
Particle selection | Num. of particles selected: 489732 | ||||||||||||||||||||||||||||
Symmetry | Point symmetry: C1 (asymmetric) | ||||||||||||||||||||||||||||
3D reconstruction | Resolution: 3.2 Å / Resolution method: FSC 0.143 CUT-OFF / Num. of particles: 26717 / Symmetry type: POINT | ||||||||||||||||||||||||||||
NMR ensemble | Conformers submitted total number: 1 |