+Open data
-Basic information
Entry | Database: PDB / ID: 8b0x | |||||||||||||||
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Title | Translating 70S ribosome in the unrotated state (P and E, tRNAs) | |||||||||||||||
Components |
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Keywords | RIBOSOME / 70S / bacterial / translation / high-resolution | |||||||||||||||
Function / homology | Function and homology information 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 ...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 / DnaA-L2 complex / four-way junction DNA binding / translation repressor activity / negative regulation of translational initiation / negative regulation of DNA-templated DNA replication initiation / regulation of mRNA stability / 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 / transcription antitermination / response to reactive oxygen species / regulation of cell growth / ribosomal large subunit assembly / 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 / cytosol / cytoplasm Similarity search - Function | |||||||||||||||
Biological species | Escherichia coli B (bacteria) | |||||||||||||||
Method | ELECTRON MICROSCOPY / single particle reconstruction / cryo EM / Resolution: 1.55 Å | |||||||||||||||
Authors | Fromm, S.A. / O'Connor, K.M. / Purdy, M. / Bhatt, P.R. / Loughran, G. / Atkins, J.F. / Jomaa, A. / Mattei, S. | |||||||||||||||
Funding support | Ireland, European Union, 4items
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Citation | Journal: Nat Commun / Year: 2023 Title: The translating bacterial ribosome at 1.55 Å resolution generated by cryo-EM imaging services. Authors: Simon A Fromm / Kate M O'Connor / Michael Purdy / Pramod R Bhatt / Gary Loughran / John F Atkins / Ahmad Jomaa / Simone Mattei / Abstract: Our understanding of protein synthesis has been conceptualised around the structure and function of the bacterial ribosome. This complex macromolecular machine is the target of important ...Our understanding of protein synthesis has been conceptualised around the structure and function of the bacterial ribosome. This complex macromolecular machine is the target of important antimicrobial drugs, an integral line of defence against infectious diseases. Here, we describe how open access to cryo-electron microscopy facilities combined with bespoke user support enabled structural determination of the translating ribosome from Escherichia coli at 1.55 Å resolution. The obtained structures allow for direct determination of the rRNA sequence to identify ribosome polymorphism sites in the E. coli strain used in this study and enable interpretation of the ribosomal active and peripheral sites at unprecedented resolution. This includes scarcely populated chimeric hybrid states of the ribosome engaged in several tRNA translocation steps resolved at ~2 Å resolution. The current map not only improves our understanding of protein synthesis but also allows for more precise structure-based drug design of antibiotics to tackle rising bacterial resistance. #1: Journal: Biorxiv / Year: 2022 Title: The translating bacterial ribosome at 1.55 angstrom resolution by open access cryo-EM Authors: Fromm, S.A. / O'Connor, K.M. / Purdy, M. / Bhatt, P.R. / Loughran, G. / Atkins, J.F. / Jomaa, A. / Mattei, S. | |||||||||||||||
History |
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-Structure visualization
Structure viewer | Molecule: MolmilJmol/JSmol |
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-Downloads & links
-Download
PDBx/mmCIF format | 8b0x.cif.gz | 3.8 MB | Display | PDBx/mmCIF format |
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PDB format | pdb8b0x.ent.gz | Display | PDB format | |
PDBx/mmJSON format | 8b0x.json.gz | Tree view | PDBx/mmJSON format | |
Others | Other downloads |
-Validation report
Arichive directory | https://data.pdbj.org/pub/pdb/validation_reports/b0/8b0x ftp://data.pdbj.org/pub/pdb/validation_reports/b0/8b0x | HTTPS FTP |
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-Related structure data
Related structure data | 15793MC 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
+50S ribosomal protein ... , 28 types, 28 molecules 0123cdefghijklmnopqrstuvwxyz
-RNA chain , 5 types, 5 molecules AXZab
#5: RNA chain | Mass: 503775.938 Da / Num. of mol.: 1 / Source method: isolated from a natural source / Source: (natural) Escherichia coli B (bacteria) / References: GenBank: 991965528 |
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#26: RNA chain | Mass: 1923.237 Da / Num. of mol.: 1 / Source method: obtained synthetically / Source: (synth.) Escherichia coli B (bacteria) |
#27: RNA chain | Mass: 24842.811 Da / Num. of mol.: 1 / Source method: isolated from a natural source / Source: (natural) Escherichia coli B (bacteria) / References: GenBank: 1804121330 |
#28: RNA chain | Mass: 948625.938 Da / Num. of mol.: 1 / Source method: isolated from a natural source / Source: (natural) Escherichia coli B (bacteria) |
#29: RNA chain | Mass: 38483.926 Da / Num. of mol.: 1 / Source method: isolated from a natural source / Source: (natural) Escherichia coli B (bacteria) / References: GenBank: 1845258627 |
-30S ribosomal protein ... , 20 types, 20 molecules BCDEFGHIJKLMNOPQRSTU
#6: Protein | Mass: 26781.670 Da / Num. of mol.: 1 / Source method: isolated from a natural source / Source: (natural) Escherichia coli B (bacteria) / References: UniProt: P0A7V0 |
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#7: Protein | Mass: 26031.316 Da / Num. of mol.: 1 / Source method: isolated from a natural source / Source: (natural) Escherichia coli B (bacteria) / Strain: K12 / References: UniProt: P0A7V3 |
#8: Protein | Mass: 23514.199 Da / Num. of mol.: 1 / Source method: isolated from a natural source / Source: (natural) Escherichia coli B (bacteria) / Strain: K12 / References: UniProt: P0A7V8 |
#9: Protein | Mass: 17629.398 Da / Num. of mol.: 1 / Source method: isolated from a natural source / Source: (natural) Escherichia coli B (bacteria) / Strain: K12 / References: UniProt: P0A7W1 |
#10: Protein | Mass: 15727.512 Da / Num. of mol.: 1 / Source method: isolated from a natural source / Source: (natural) Escherichia coli B (bacteria) / References: UniProt: P02358 |
#11: Protein | Mass: 20055.156 Da / Num. of mol.: 1 / Source method: isolated from a natural source / Source: (natural) Escherichia coli B (bacteria) / References: UniProt: P02359 |
#12: Protein | Mass: 14146.557 Da / Num. of mol.: 1 / Source method: isolated from a natural source / Source: (natural) Escherichia coli B (bacteria) / Strain: K12 / References: UniProt: P0A7W7 |
#13: Protein | Mass: 14886.270 Da / Num. of mol.: 1 / Source method: isolated from a natural source / Source: (natural) Escherichia coli B (bacteria) / Strain: K12 / References: UniProt: P0A7X3 |
#14: Protein | Mass: 11755.597 Da / Num. of mol.: 1 / Source method: isolated from a natural source / Source: (natural) Escherichia coli B (bacteria) / Strain: K12 / References: UniProt: P0A7R5 |
#15: Protein | Mass: 13871.959 Da / Num. of mol.: 1 / Source method: isolated from a natural source / Source: (natural) Escherichia coli B (bacteria) / Strain: K12 / References: UniProt: P0A7R9 |
#16: Protein | Mass: 13814.249 Da / Num. of mol.: 1 / Source method: isolated from a natural source / Source: (natural) Escherichia coli B (bacteria) / Strain: K12 / References: UniProt: P0A7S3 |
#17: Protein | Mass: 13128.467 Da / Num. of mol.: 1 / Source method: isolated from a natural source / Source: (natural) Escherichia coli B (bacteria) / Strain: K12 / References: UniProt: P0A7S9 |
#18: Protein | Mass: 11606.560 Da / Num. of mol.: 1 / Source method: isolated from a natural source / Source: (natural) Escherichia coli B (bacteria) / Strain: K12 / References: UniProt: P0AG59 |
#19: Protein | Mass: 10290.816 Da / Num. of mol.: 1 / Source method: isolated from a natural source / Source: (natural) Escherichia coli B (bacteria) / Strain: K12 / References: UniProt: P0ADZ4 |
#20: Protein | Mass: 9207.572 Da / Num. of mol.: 1 / Source method: isolated from a natural source / Source: (natural) Escherichia coli B (bacteria) / Strain: K12 / References: UniProt: P0A7T3 |
#21: Protein | Mass: 9724.491 Da / Num. of mol.: 1 / Source method: isolated from a natural source / Source: (natural) Escherichia coli B (bacteria) / Strain: K12 / References: UniProt: P0AG63 |
#22: Protein | Mass: 9005.472 Da / Num. of mol.: 1 / Source method: isolated from a natural source / Source: (natural) Escherichia coli B (bacteria) / Strain: K12 / References: UniProt: P0A7T7 |
#23: Protein | Mass: 10455.355 Da / Num. of mol.: 1 / Source method: isolated from a natural source / Source: (natural) Escherichia coli B (bacteria) / Strain: K12 / References: UniProt: P0A7U3 |
#24: Protein | Mass: 9708.464 Da / Num. of mol.: 1 / Source method: isolated from a natural source / Source: (natural) Escherichia coli B (bacteria) / References: UniProt: P0A7U7 |
#25: Protein | Mass: 8524.039 Da / Num. of mol.: 1 / Source method: isolated from a natural source / Source: (natural) Escherichia coli B (bacteria) / Strain: K12 / References: UniProt: P68679 |
-Non-polymers , 4 types, 11991 molecules
#54: Chemical | ChemComp-ZN / | ||||
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#55: Chemical | ChemComp-K / #56: Chemical | ChemComp-MG / #57: Water | ChemComp-HOH / | |
-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: 70S ribosome / Type: RIBOSOME / Details: translating with P-site tRNA and mRNA / Entity ID: #1-#14, #16-#38, #40-#53 / Source: NATURAL | ||||||||||||||||||||
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Molecular weight | Value: 2.5 MDa / Experimental value: NO | ||||||||||||||||||||
Source (natural) | Organism: Escherichia coli B (bacteria) | ||||||||||||||||||||
Buffer solution | pH: 7.4 | ||||||||||||||||||||
Buffer component |
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Specimen | Conc.: 0.5 mg/ml / Embedding applied: NO / Shadowing applied: NO / Staining applied: NO / Vitrification applied: YES | ||||||||||||||||||||
Specimen support | Details: Fischione 1070 / Grid material: COPPER / Grid mesh size: 300 divisions/in. / Grid type: Quantifoil R2/1 | ||||||||||||||||||||
Vitrification | Instrument: FEI VITROBOT MARK IV / Cryogen name: ETHANE / Humidity: 100 % / Chamber temperature: 279 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 / Nominal magnification: 165000 X / Nominal defocus max: 1500 nm / Nominal defocus min: 500 nm / Cs: 2.7 mm / C2 aperture diameter: 50 µm / Alignment procedure: COMA FREE |
Specimen holder | Cryogen: NITROGEN / Specimen holder model: FEI TITAN KRIOS AUTOGRID HOLDER |
Image recording | Average exposure time: 5.2 sec. / Electron dose: 40 e/Å2 / Film or detector model: FEI FALCON IV (4k x 4k) / Num. of grids imaged: 1 / Num. of real images: 19449 Details: saved in EER format; 16 exposures per hole/stage movement. 11 in outer ring around the hole edge, 5 in inner ring around the hole center. |
EM imaging optics | Energyfilter name: TFS Selectris X / Energyfilter slit width: 10 eV |
Image scans | Sampling size: 14 µm / Width: 4096 / Height: 4096 |
-Processing
Software | Name: UCSF ChimeraX / Version: 1.4/v9 / Classification: model building / URL: https://www.rbvi.ucsf.edu/chimerax/ / Os: macOS / Type: package | ||||||||||||||||||||||||||||||
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EM software |
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CTF correction | Details: per-particle CTF correction / Type: PHASE FLIPPING AND AMPLITUDE CORRECTION | ||||||||||||||||||||||||||||||
Particle selection | Num. of particles selected: 1579392 | ||||||||||||||||||||||||||||||
Symmetry | Point symmetry: C1 (asymmetric) | ||||||||||||||||||||||||||||||
3D reconstruction | Resolution: 1.55 Å / Resolution method: FSC 0.143 CUT-OFF / Num. of particles: 506020 / Num. of class averages: 2 / Symmetry type: POINT | ||||||||||||||||||||||||||||||
Atomic model building | Protocol: FLEXIBLE FIT / Space: REAL | ||||||||||||||||||||||||||||||
Atomic model building | PDB-ID: 7K00 Accession code: 7K00 / Source name: PDB / Type: experimental model |