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Open data
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Basic information
Entry | Database: PDB / ID: 8fto | |||||||||
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Title | E. coli 70S ribosome with an improved MS2 tag inserted in H98 | |||||||||
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![]() | RIBOSOME / MS2-tag / H98 | |||||||||
Function / homology | ![]() negative regulation of cytoplasmic translational initiation / positive regulation of ribosome biogenesis / DnaA-L2 complex / negative regulation of translational initiation / negative regulation of DNA-templated DNA replication initiation / translational initiation / ribosome assembly / mRNA regulatory element binding translation repressor activity / assembly of large subunit precursor of preribosome / cytosolic ribosome assembly ...negative regulation of cytoplasmic translational initiation / positive regulation of ribosome biogenesis / DnaA-L2 complex / negative regulation of translational initiation / negative regulation of DNA-templated DNA replication initiation / translational initiation / ribosome assembly / mRNA regulatory element binding translation repressor activity / assembly of large subunit precursor of preribosome / cytosolic ribosome assembly / transcription antitermination / regulation of cell growth / DNA-templated transcription termination / maintenance of translational fidelity / ribosomal large subunit assembly / mRNA 5'-UTR binding / large ribosomal subunit / ribosome binding / 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 / tRNA binding / cytoplasmic translation / rRNA binding / negative regulation of translation / ribosome / structural constituent of ribosome / ribonucleoprotein complex / translation / response to antibiotic / mRNA binding / RNA binding / zinc ion binding / membrane / cytoplasm / cytosol Similarity search - Function | |||||||||
Biological species | ![]() ![]() | |||||||||
Method | ELECTRON MICROSCOPY / single particle reconstruction / cryo EM / Resolution: 1.85 Å | |||||||||
![]() | Nissley, A.J. / Cate, J.H.D. | |||||||||
Funding support | ![]()
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![]() | ![]() Title: Interactions between terminal ribosomal RNA helices stabilize the large ribosomal subunit. Authors: Amos J Nissley / Tammam S Kamal / Jamie H D Cate / ![]() Abstract: The ribosome is a large ribonucleoprotein assembly that uses diverse and complex molecular interactions to maintain proper folding. In vivo assembled ribosomes have been isolated using MS2 tags ...The ribosome is a large ribonucleoprotein assembly that uses diverse and complex molecular interactions to maintain proper folding. In vivo assembled ribosomes have been isolated using MS2 tags installed in either the 16S or 23S ribosomal RNAs (rRNAs), to enable studies of ribosome structure and function in vitro. RNA tags in the 50S subunit have commonly been inserted into an extended helix H98 in 23S rRNA, as this addition does not affect cellular growth or in vitro ribosome activity. Here, we find that 50S subunits with MS2 tags inserted in H98 are destabilized compared to wild-type (WT) 50S subunits. We identify the loss of RNA-RNA tertiary contacts that bridge helices H1, H94, and H98 as the cause of destabilization. Using cryogenic electron microscopy (cryo-EM), we show that this interaction is disrupted by the addition of the MS2 tag and can be restored through the insertion of a single adenosine in the extended H98 helix. This work establishes ways to improve MS2 tags in the 50S subunit that maintain ribosome stability and investigates a complex RNA tertiary structure that may be important for stability in various bacterial ribosomes. | |||||||||
History |
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Structure visualization
Structure viewer | Molecule: ![]() ![]() |
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Downloads & links
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Download
PDBx/mmCIF format | ![]() | 3.6 MB | Display | ![]() |
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PDB format | ![]() | Display | ![]() | |
PDBx/mmJSON format | ![]() | Tree view | ![]() | |
Others | ![]() |
-Validation report
Summary document | ![]() | 1.4 MB | Display | ![]() |
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Full document | ![]() | 1.4 MB | Display | |
Data in XML | ![]() | 191.5 KB | Display | |
Data in CIF | ![]() | 378.6 KB | Display | |
Arichive directory | ![]() ![]() | HTTPS FTP |
-Related structure data
Related structure data | ![]() 29449MC M: map data used to model this data C: citing same article ( |
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Similar structure data | Similarity search - Function & homology ![]() |
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Links
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Assembly
Deposited unit | ![]()
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1 |
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Components
+50S ribosomal protein ... , 28 types, 28 molecules 01234cdefghijklmnoprstuvwxyz
-RNA chain , 5 types, 5 molecules AXZab
#6: RNA chain | Mass: 499873.406 Da / Num. of mol.: 1 / Source method: isolated from a natural source / Source: (natural) ![]() ![]() |
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#27: RNA chain | Mass: 8707.231 Da / Num. of mol.: 1 / Source method: obtained synthetically / Source: (synth.) ![]() ![]() |
#28: RNA chain | Mass: 24496.617 Da / Num. of mol.: 1 / Source method: obtained synthetically / Source: (synth.) ![]() ![]() |
#29: RNA chain | Mass: 952350.188 Da / Num. of mol.: 1 / Source method: isolated from a natural source / Source: (natural) ![]() ![]() |
#30: RNA chain | Mass: 38790.090 Da / Num. of mol.: 1 / Source method: isolated from a natural source / Source: (natural) ![]() ![]() |
-30S ribosomal protein ... , 20 types, 20 molecules BCDEFGHIJKLMNOPQRSTU
#7: Protein | Mass: 26781.670 Da / Num. of mol.: 1 / Source method: isolated from a natural source / Source: (natural) ![]() ![]() |
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#8: Protein | Mass: 26031.316 Da / Num. of mol.: 1 / Source method: isolated from a natural source / Source: (natural) ![]() ![]() |
#9: Protein | Mass: 23514.199 Da / Num. of mol.: 1 / Source method: isolated from a natural source / Source: (natural) ![]() ![]() |
#10: Protein | Mass: 17629.398 Da / Num. of mol.: 1 / Source method: isolated from a natural source / Source: (natural) ![]() ![]() |
#11: Protein | Mass: 15727.512 Da / Num. of mol.: 1 / Source method: isolated from a natural source / Source: (natural) ![]() ![]() |
#12: Protein | Mass: 20055.156 Da / Num. of mol.: 1 / Source method: isolated from a natural source / Source: (natural) ![]() ![]() |
#13: Protein | Mass: 14146.557 Da / Num. of mol.: 1 / Source method: isolated from a natural source / Source: (natural) ![]() ![]() |
#14: Protein | Mass: 14886.270 Da / Num. of mol.: 1 / Source method: isolated from a natural source / Source: (natural) ![]() ![]() |
#15: Protein | Mass: 11755.597 Da / Num. of mol.: 1 / Source method: isolated from a natural source / Source: (natural) ![]() ![]() |
#16: Protein | Mass: 13871.959 Da / Num. of mol.: 1 / Source method: isolated from a natural source / Source: (natural) ![]() ![]() |
#17: Protein | Mass: 13814.249 Da / Num. of mol.: 1 / Source method: isolated from a natural source / Source: (natural) ![]() ![]() |
#18: Protein | Mass: 13128.467 Da / Num. of mol.: 1 / Source method: isolated from a natural source / Source: (natural) ![]() ![]() |
#19: Protein | Mass: 11606.560 Da / Num. of mol.: 1 / Source method: isolated from a natural source / Source: (natural) ![]() ![]() |
#20: Protein | Mass: 10290.816 Da / Num. of mol.: 1 / Source method: isolated from a natural source / Source: (natural) ![]() ![]() |
#21: Protein | Mass: 9207.572 Da / Num. of mol.: 1 / Source method: isolated from a natural source / Source: (natural) ![]() ![]() |
#22: Protein | Mass: 9724.491 Da / Num. of mol.: 1 / Source method: isolated from a natural source / Source: (natural) ![]() ![]() |
#23: Protein | Mass: 9005.472 Da / Num. of mol.: 1 / Source method: isolated from a natural source / Source: (natural) ![]() ![]() |
#24: Protein | Mass: 10455.355 Da / Num. of mol.: 1 / Source method: isolated from a natural source / Source: (natural) ![]() ![]() |
#25: Protein | Mass: 9708.464 Da / Num. of mol.: 1 / Source method: isolated from a natural source / Source: (natural) ![]() ![]() |
#26: Protein | Mass: 8524.039 Da / Num. of mol.: 1 / Source method: isolated from a natural source / Source: (natural) ![]() ![]() |
-Protein , 1 types, 1 molecules q
#45: Protein | Mass: 11586.374 Da / Num. of mol.: 1 / Source method: isolated from a natural source / Source: (natural) ![]() ![]() |
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-Non-polymers , 3 types, 6399 molecules ![](data/chem/img/MG.gif)
![](data/chem/img/FME.gif)
![](data/chem/img/HOH.gif)
![](data/chem/img/FME.gif)
![](data/chem/img/HOH.gif)
#55: Chemical | ChemComp-MG / #56: Chemical | ChemComp-FME / | #57: Water | ChemComp-HOH / | |
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-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 |
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Sample preparation
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Source (natural) |
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Buffer solution | pH: 7.5 | ||||||||||||||||||||||||||||||||||||
Buffer component |
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Specimen | Conc.: 0.27 mg/ml / Embedding applied: NO / Shadowing applied: NO / Staining applied: NO / Vitrification applied: YES | ||||||||||||||||||||||||||||||||||||
Specimen support | Grid material: GOLD / Grid mesh size: 300 divisions/in. / Grid type: UltrAuFoil R1.2/1.3 | ||||||||||||||||||||||||||||||||||||
Vitrification | Instrument: FEI VITROBOT MARK III / Cryogen name: ETHANE / Humidity: 100 % / Chamber temperature: 277 K |
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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: ![]() |
Electron lens | Mode: BRIGHT FIELD / Nominal defocus max: 1500 nm / Nominal defocus min: 500 nm |
Image recording | Electron dose: 40 e/Å2 / Film or detector model: GATAN K3 (6k x 4k) / Num. of grids imaged: 1 / Num. of real images: 11736 |
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Processing
EM software |
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CTF correction | Type: PHASE FLIPPING AND AMPLITUDE CORRECTION | ||||||||||||||||||||||||||||||||||||||||||||
Particle selection | Num. of particles selected: 2475999 | ||||||||||||||||||||||||||||||||||||||||||||
3D reconstruction | Resolution: 1.85 Å / Resolution method: FSC 0.143 CUT-OFF / Num. of particles: 601617 / Symmetry type: POINT | ||||||||||||||||||||||||||||||||||||||||||||
Atomic model building | Protocol: RIGID BODY FIT |