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Yorodumi- PDB-8oip: 28S mammalian mitochondrial small ribosomal subunit with mtRF1 an... -
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-Basic information
Entry | Database: PDB / ID: 8oip | ||||||||||||
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Title | 28S mammalian mitochondrial small ribosomal subunit with mtRF1 and P-site tRNA | ||||||||||||
Components |
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Keywords | RIBOSOME / mammalian mitochondrial ribosome / release factor mtRF1 / non-canonical stop codon | ||||||||||||
Function / homology | Function and homology information Hormone ligand-binding receptors / gonadotropin-releasing hormone receptor binding / gonadotropin hormone-releasing hormone activity / G alpha (q) signalling events / Mitochondrial translation elongation / Mitochondrial translation termination / : / translation release factor activity / mitochondrial ribosome assembly / mitochondrial translational termination ...Hormone ligand-binding receptors / gonadotropin-releasing hormone receptor binding / gonadotropin hormone-releasing hormone activity / G alpha (q) signalling events / Mitochondrial translation elongation / Mitochondrial translation termination / : / translation release factor activity / mitochondrial ribosome assembly / mitochondrial translational termination / translation release factor activity, codon specific / Mitochondrial protein degradation / mitochondrial large ribosomal subunit / mitochondrial small ribosomal subunit / mitochondrial ribosome / mitochondrial translation / organelle membrane / positive regulation of proteolysis / ribosomal small subunit binding / regulation of translation / ribosomal small subunit assembly / small ribosomal subunit / small ribosomal subunit rRNA binding / nuclear membrane / cytosolic small ribosomal subunit / cytosolic large ribosomal subunit / rRNA binding / ribosome / structural constituent of ribosome / mitochondrial matrix / ribonucleoprotein complex / translation / mRNA binding / apoptotic process / synapse / nucleolus / mitochondrion / RNA binding / extracellular space / nucleoplasm Similarity search - Function | ||||||||||||
Biological species | Homo sapiens (human) Sus scrofa (pig) | ||||||||||||
Method | ELECTRON MICROSCOPY / single particle reconstruction / cryo EM / Resolution: 3.6 Å | ||||||||||||
Authors | Saurer, M. / Leibundgut, M. / Scaiola, A. / Schoenhut, T. / Ban, N. | ||||||||||||
Funding support | Switzerland, European Union, 3items
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Citation | Journal: Science / Year: 2023 Title: Molecular basis of translation termination at noncanonical stop codons in human mitochondria. Authors: Martin Saurer / Marc Leibundgut / Hima Priyanka Nadimpalli / Alain Scaiola / Tanja Schönhut / Richard G Lee / Stefan J Siira / Oliver Rackham / René Dreos / Tea Lenarčič / Eva Kummer / ...Authors: Martin Saurer / Marc Leibundgut / Hima Priyanka Nadimpalli / Alain Scaiola / Tanja Schönhut / Richard G Lee / Stefan J Siira / Oliver Rackham / René Dreos / Tea Lenarčič / Eva Kummer / David Gatfield / Aleksandra Filipovska / Nenad Ban / Abstract: The genetic code that specifies the identity of amino acids incorporated into proteins during protein synthesis is almost universally conserved. Mitochondrial genomes feature deviations from the ...The genetic code that specifies the identity of amino acids incorporated into proteins during protein synthesis is almost universally conserved. Mitochondrial genomes feature deviations from the standard genetic code, including the reassignment of two arginine codons to stop codons. The protein required for translation termination at these noncanonical stop codons to release the newly synthesized polypeptides is not currently known. In this study, we used gene editing and ribosomal profiling in combination with cryo-electron microscopy to establish that mitochondrial release factor 1 (mtRF1) detects noncanonical stop codons in human mitochondria by a previously unknown mechanism of codon recognition. We discovered that binding of mtRF1 to the decoding center of the ribosome stabilizes a highly unusual conformation in the messenger RNA in which the ribosomal RNA participates in specific recognition of the noncanonical stop codons. | ||||||||||||
History |
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-Structure visualization
Structure viewer | Molecule: MolmilJmol/JSmol |
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-Downloads & links
-Download
PDBx/mmCIF format | 8oip.cif.gz | 1.6 MB | Display | PDBx/mmCIF format |
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PDB format | pdb8oip.ent.gz | Display | PDB format | |
PDBx/mmJSON format | 8oip.json.gz | Tree view | PDBx/mmJSON format | |
Others | Other downloads |
-Validation report
Summary document | 8oip_validation.pdf.gz | 2.1 MB | Display | wwPDB validaton report |
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Full document | 8oip_full_validation.pdf.gz | 2.1 MB | Display | |
Data in XML | 8oip_validation.xml.gz | 174.5 KB | Display | |
Data in CIF | 8oip_validation.cif.gz | 280.5 KB | Display | |
Arichive directory | https://data.pdbj.org/pub/pdb/validation_reports/oi/8oip ftp://data.pdbj.org/pub/pdb/validation_reports/oi/8oip | HTTPS FTP |
-Related structure data
Related structure data | 16895MC 8oinC 8oiqC 8oirC 8oisC 8oitC C: citing same article (ref.) M: map data used to model this data |
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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
-Protein , 12 types, 12 molecules BXBdADAEANASAWAXAaAcAeAg
#1: Protein | Mass: 34528.809 Da / Num. of mol.: 1 / Source method: isolated from a natural source / Source: (natural) Sus scrofa (pig) / Organ: Liver / References: UniProt: I3LNJ0 |
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#2: Protein | Mass: 15156.537 Da / Num. of mol.: 1 / Source method: isolated from a natural source / Source: (natural) Sus scrofa (pig) / Organ: Liver / References: UniProt: I3L9J0 |
#6: Protein | Mass: 22843.941 Da / Num. of mol.: 1 / Source method: isolated from a natural source / Source: (natural) Sus scrofa (pig) / Organ: Liver / References: UniProt: A0A287B3R8 |
#7: Protein | Mass: 41085.398 Da / Num. of mol.: 1 / Source method: isolated from a natural source Details: NCBI Reference Sequence: XP_020925505.1,NCBI Reference Sequence: XP_020925505.1 Source: (natural) Sus scrofa (pig) / Organ: Liver / References: UniProt: A0A4X1TSM9 |
#16: Protein | Mass: 14452.014 Da / Num. of mol.: 1 / Source method: isolated from a natural source / Source: (natural) Sus scrofa (pig) / Organ: Liver / References: UniProt: F1RIU0 |
#21: Protein | Mass: 21666.674 Da / Num. of mol.: 1 / Source method: isolated from a natural source / Details: NCBI Reference Sequence: XP_005669023.1 / Source: (natural) Sus scrofa (pig) / Organ: Liver / References: UniProt: A0A287AEW3 |
#25: Protein | Mass: 21014.912 Da / Num. of mol.: 1 / Source method: isolated from a natural source / Source: (natural) Sus scrofa (pig) / Organ: Liver / References: UniProt: A0A0M3KL56 |
#26: Protein | Mass: 46966.062 Da / Num. of mol.: 1 / Source method: isolated from a natural source / Source: (natural) Sus scrofa (pig) / Organ: Liver / References: UniProt: I3LS10 |
#29: Protein | Mass: 55569.742 Da / Num. of mol.: 1 Source method: isolated from a genetically manipulated source Details: Human mitochondrial release factor 1 (mtRF1), with a C-terminal linker (GGSGGSG) and a 3xFLAG-tag (DYKDHDGDYKDHDIDYKDDDDK), expressed from a pcDNA3.1(+) plasmid in HEK293 EBNA cells. Source: (gene. exp.) Homo sapiens (human) / Gene: MTRF1 / Plasmid: pcDNA3.1(+) / Cell line (production host): HEK293 EBNA cells / Production host: Homo sapiens (human) / References: UniProt: O75570 |
#31: Protein | Mass: 13603.939 Da / Num. of mol.: 1 / Source method: isolated from a natural source / Source: (natural) Sus scrofa (pig) / Organ: Liver / References: UniProt: F1SU49 |
#33: Protein | Mass: 78518.156 Da / Num. of mol.: 1 / Source method: isolated from a natural source / Details: NCBI Reference Sequence: XP_005662461.1 / Source: (natural) Sus scrofa (pig) / Organ: Liver / References: UniProt: K7GKS8 |
#34: Protein | Mass: 45604.570 Da / Num. of mol.: 1 / Source method: isolated from a natural source / Source: (natural) Sus scrofa (pig) / Organ: Liver / References: UniProt: I3LU08 |
-RNA chain , 3 types, 3 molecules AAAGAI
#3: RNA chain | Mass: 309087.250 Da / Num. of mol.: 1 / Source method: isolated from a natural source Details: Corresponds to NCBI reference sequence NC_012095.1 (positions 70-1031) Source: (natural) Sus scrofa (pig) / Organ: Liver / References: GenBank: NC_012095.1 |
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#9: RNA chain | Mass: 22823.703 Da / Num. of mol.: 1 / Source method: obtained synthetically Details: Corresponds to GenBank sequence ID LC530718.1 (positions 1-71). In vitro transcribed human tRNA with formyl-methionine attached to the CCA-end of the tRNA. Source: (synth.) Homo sapiens (human) |
#11: RNA chain | Mass: 2902.831 Da / Num. of mol.: 1 / Source method: obtained synthetically / Details: Synthetic mRNA / Source: (synth.) Homo sapiens (human) |
-28S ribosomal protein ... , 7 types, 7 molecules ABALAMAOAQAYAd
#4: Protein | Mass: 41357.789 Da / Num. of mol.: 1 / Source method: isolated from a natural source / Source: (natural) Sus scrofa (pig) / Organ: Liver / References: UniProt: F1SG95 |
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#14: Protein | Mass: 29596.867 Da / Num. of mol.: 1 / Source method: isolated from a natural source / Source: (natural) Sus scrofa (pig) / Organ: Liver / References: UniProt: A0A287AKS1 |
#15: Protein | Mass: 15182.668 Da / Num. of mol.: 1 / Source method: isolated from a natural source / Source: (natural) Sus scrofa (pig) / Organ: Liver / References: UniProt: A0A480JTD7 |
#17: Protein | Mass: 29220.465 Da / Num. of mol.: 1 / Source method: isolated from a natural source / Source: (natural) Sus scrofa (pig) / Organ: Liver / References: UniProt: Q767K8 |
#19: Protein | Mass: 10724.543 Da / Num. of mol.: 1 / Source method: isolated from a natural source / Source: (natural) Sus scrofa (pig) / Organ: Liver / References: UniProt: A0A286ZNB3 |
#27: Protein | Mass: 43516.496 Da / Num. of mol.: 1 / Source method: isolated from a natural source / Source: (natural) Sus scrofa (pig) / Organ: Liver / References: UniProt: F1RME2 |
#32: Protein | Mass: 48213.074 Da / Num. of mol.: 1 / Source method: isolated from a natural source / Source: (natural) Sus scrofa (pig) / Organ: Liver / References: UniProt: F1SU66 |
-Mitochondrial ribosomal protein ... , 14 types, 14 molecules ACAFAHAJAKAPARATAUAVAZAbAiAj
#5: Protein | Mass: 18935.172 Da / Num. of mol.: 1 / Source method: isolated from a natural source / Source: (natural) Sus scrofa (pig) / Organ: Liver / References: UniProt: A0A287ANU4 |
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#8: Protein | Mass: 28150.918 Da / Num. of mol.: 1 / Source method: isolated from a natural source / Source: (natural) Sus scrofa (pig) / Organ: Liver / References: UniProt: A0A286ZJ25 |
#10: Protein | Mass: 23105.678 Da / Num. of mol.: 1 / Source method: isolated from a natural source / Source: (natural) Sus scrofa (pig) / Organ: Liver / References: UniProt: F1RUU2 |
#12: Protein | Mass: 15257.107 Da / Num. of mol.: 1 / Source method: isolated from a natural source / Source: (natural) Sus scrofa (pig) / Organ: Liver / References: UniProt: A0A0M3KL52 |
#13: Protein | Mass: 15120.739 Da / Num. of mol.: 1 / Source method: isolated from a natural source / Source: (natural) Sus scrofa (pig) / Organ: Liver / References: UniProt: A0A0M3KL53 |
#18: Protein | Mass: 16211.086 Da / Num. of mol.: 1 / Source method: isolated from a natural source / Source: (natural) Sus scrofa (pig) / Organ: Liver / References: UniProt: A0A0M3KL54 |
#20: Protein | Mass: 43234.645 Da / Num. of mol.: 1 / Source method: isolated from a natural source / Source: (natural) Sus scrofa (pig) / Organ: Liver / References: UniProt: A0A287AQT5 |
#22: Protein | Mass: 19954.186 Da / Num. of mol.: 1 / Source method: isolated from a natural source / Source: (natural) Sus scrofa (pig) / Organ: Liver / References: UniProt: A0A0M3KL55 |
#23: Protein | Mass: 23637.900 Da / Num. of mol.: 1 / Source method: isolated from a natural source / Details: NCBI Reference Sequence: XP_001924843.2 / Source: (natural) Sus scrofa (pig) / Organ: Liver / References: UniProt: A0A287AY01 |
#24: Protein | Mass: 45761.789 Da / Num. of mol.: 1 / Source method: isolated from a natural source / Source: (natural) Sus scrofa (pig) / Organ: Liver / References: UniProt: A0A5G2R1I0 |
#28: Protein | Mass: 16971.662 Da / Num. of mol.: 1 / Source method: isolated from a natural source / Source: (natural) Sus scrofa (pig) / Organ: Liver / References: UniProt: F1S8G3 |
#30: Protein | Mass: 31731.604 Da / Num. of mol.: 1 / Source method: isolated from a natural source / Source: (natural) Sus scrofa (pig) / Organ: Liver / References: UniProt: F1S001 |
#35: Protein | Mass: 20856.934 Da / Num. of mol.: 1 / Source method: isolated from a natural source / Source: (natural) Sus scrofa (pig) / Organ: Liver / References: UniProt: A0A286ZJJ6 |
#36: Protein | Mass: 55714.398 Da / Num. of mol.: 1 / Source method: isolated from a natural source / Source: (natural) Sus scrofa (pig) / Organ: Liver / References: UniProt: A0A287AVS9 |
-Non-polymers , 9 types, 148 molecules
#37: Chemical | ChemComp-MG / #38: Chemical | ChemComp-SPM / | #39: Chemical | ChemComp-SPD / | #40: Chemical | ChemComp-K / #41: Chemical | ChemComp-ZN / | #42: Chemical | #43: Chemical | ChemComp-ATP / | #44: Chemical | ChemComp-GDP / | #45: 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 |
-Sample preparation
Component | Name: 28S mammalian mitochondrial small ribosomal subunit with mtRF1, mRNA, and P-site tRNA Type: RIBOSOME Details: Mammalian mitochondrial translation termination complex, in vitro reconstituted from porcine mitoribosomal subunits, human mtRF1-3xFLAG, human methionine-tRNA(formyl-methionine), and a ...Details: Mammalian mitochondrial translation termination complex, in vitro reconstituted from porcine mitoribosomal subunits, human mtRF1-3xFLAG, human methionine-tRNA(formyl-methionine), and a synthetic mRNA (5'-A AUG AGA CAA-3') containing the non-canonical stop codon AGA. Entity ID: #1-#36 / Source: NATURAL | ||||||||||||||||||||
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Molecular weight | Value: 1.1 MDa / Experimental value: NO | ||||||||||||||||||||
Source (natural) | Organism: Sus scrofa (pig) / Cellular location: Mitochondrial matrix / Organelle: Mitochondria / Tissue: Liver | ||||||||||||||||||||
Buffer solution | pH: 7.5 Details: 20mM HEPES-KOH pH=7.5, 100mM KCl, 20mM MgCl2, ~1% sucrose (from mtRF1 storage buffer) | ||||||||||||||||||||
Buffer component |
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Specimen | Embedding applied: NO / Shadowing applied: NO / Staining applied: NO / Vitrification applied: YES Details: In vitro reconstituted complex (55S). Focused refinement on small subunit. | ||||||||||||||||||||
Specimen support | Grid material: COPPER / Grid type: Quantifoil R2/2 | ||||||||||||||||||||
Vitrification | Instrument: FEI VITROBOT MARK IV / Cryogen name: ETHANE-PROPANE / 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 / Nominal magnification: 81000 X / Nominal defocus max: 3000 nm / Nominal defocus min: 600 nm / Cs: 2.7 mm / C2 aperture diameter: 70 µm / Alignment procedure: COMA FREE |
Specimen holder | Cryogen: NITROGEN / Specimen holder model: FEI TITAN KRIOS AUTOGRID HOLDER |
Image recording | Electron dose: 60 e/Å2 / Film or detector model: GATAN K3 BIOQUANTUM (6k x 4k) / Num. of grids imaged: 1 / Num. of real images: 6973 |
EM imaging optics | Energyfilter slit width: 20 eV |
-Processing
EM software |
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Image processing | Details: Micrographs were motion corrected and CTF parameters were estimated using Cryosparc live v4. | ||||||||||||||||||||||||||||||||||||
CTF correction | Type: PHASE FLIPPING AND AMPLITUDE CORRECTION | ||||||||||||||||||||||||||||||||||||
Particle selection | Num. of particles selected: 1471587 Details: Particles were picked with blob picker in Cryosparc live v4. | ||||||||||||||||||||||||||||||||||||
3D reconstruction | Resolution: 3.6 Å / Resolution method: FSC 0.143 CUT-OFF / Num. of particles: 50622 / Symmetry type: POINT | ||||||||||||||||||||||||||||||||||||
Atomic model building | Protocol: FLEXIBLE FIT / Space: REAL | ||||||||||||||||||||||||||||||||||||
Atomic model building | 3D fitting-ID: 1
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