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Yorodumi- PDB-8oit: 39S human mitochondrial large ribosomal subunit with mtRF1 and P-... -
+Open data
-Basic information
Entry | Database: PDB / ID: 8oit | ||||||||||||
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Title | 39S human mitochondrial large ribosomal subunit with mtRF1 and P-site tRNA | ||||||||||||
Components |
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Keywords | RIBOSOME / human mitochondrial ribosome / release factor mtRF1 / non-canonical stop codon | ||||||||||||
Function / homology | Function and homology information translation release factor activity / rRNA import into mitochondrion / mitochondrial translational termination / mitochondrial transcription / mitochondrial translational elongation / translation release factor activity, codon nonspecific / translation release factor activity, codon specific / microprocessor complex / Mitochondrial translation elongation / Mitochondrial translation termination ...translation release factor activity / rRNA import into mitochondrion / mitochondrial translational termination / mitochondrial transcription / mitochondrial translational elongation / translation release factor activity, codon nonspecific / translation release factor activity, codon specific / microprocessor complex / Mitochondrial translation elongation / Mitochondrial translation termination / Mitochondrial translation initiation / mitochondrial large ribosomal subunit / Hydrolases; Acting on ester bonds; Endoribonucleases producing 5'-phosphomonoesters / peptidyl-tRNA hydrolase / mitochondrial small ribosomal subunit / aminoacyl-tRNA hydrolase activity / mitochondrial ribosome / mitochondrial translation / anatomical structure morphogenesis / RNA processing / rescue of stalled ribosome / cellular response to leukemia inhibitory factor / small ribosomal subunit rRNA binding / fibrillar center / large ribosomal subunit rRNA binding / double-stranded RNA binding / cell junction / 5S rRNA binding / endonuclease activity / mitochondrial inner membrane / negative regulation of translation / rRNA binding / nuclear body / ribosome / mitochondrial matrix / structural constituent of ribosome / cell cycle / translation / ribonucleoprotein complex / protein domain specific binding / nucleotide binding / mRNA binding / synapse / apoptotic process / nucleolus / positive regulation of DNA-templated transcription / mitochondrion / extracellular space / RNA binding / nucleoplasm / nucleus / plasma membrane / cytosol Similarity search - Function | ||||||||||||
Biological species | Homo sapiens (human) | ||||||||||||
Method | ELECTRON MICROSCOPY / single particle reconstruction / cryo EM / Resolution: 2.9 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 | 8oit.cif.gz | 2.8 MB | Display | PDBx/mmCIF format |
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PDB format | pdb8oit.ent.gz | Display | PDB format | |
PDBx/mmJSON format | 8oit.json.gz | Tree view | PDBx/mmJSON format | |
Others | Other downloads |
-Validation report
Arichive directory | https://data.pdbj.org/pub/pdb/validation_reports/oi/8oit ftp://data.pdbj.org/pub/pdb/validation_reports/oi/8oit | HTTPS FTP |
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-Related structure data
Related structure data | 16899MC 8oinC 8oipC 8oiqC 8oirC 8oisC 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
+39S ribosomal protein ... , 49 types, 54 molecules B1B2B3B4B5B6BABBBCBDBEBFBGBHBIBJBKBLBMBNBOBPBQBRBSBTBUBVBWBX...
-RNA chain , 4 types, 4 molecules B7B8B9AG
#2: RNA chain | Mass: 894.612 Da / Num. of mol.: 1 / Source method: isolated from a natural source / Source: (natural) Homo sapiens (human) / Cell line: HEK293 EBNA |
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#3: RNA chain | Mass: 500727.125 Da / Num. of mol.: 1 / Source method: isolated from a natural source Details: NCBI Reference Sequence: NC_012920.1 added 3' GU (encoded on genome but not annotated as part of the 16S rRNA), N1437>U as in MT079070.1 Source: (natural) Homo sapiens (human) / Cell line: HEK293 EBNA / References: GenBank: 1563835895 |
#4: RNA chain | Mass: 22989.752 Da / Num. of mol.: 1 / Source method: isolated from a natural source / Details: NCBI Reference Sequence: NC_012920.1 / Source: (natural) Homo sapiens (human) / Cell line: HEK293 EBNA / References: GenBank: NC_012920.1 |
#56: RNA chain | Mass: 22692.508 Da / Num. of mol.: 1 / Source method: isolated from a natural source Details: NCBI Reference Sequence: NC_012920.1 + aminoacylated CCA-end Source: (natural) Homo sapiens (human) / Cell line: HEK293 EBNA / References: GenBank: NC_012920.1 |
-Protein , 4 types, 4 molecules BeBfBgAa
#35: Protein | Mass: 12292.333 Da / Num. of mol.: 1 / Source method: isolated from a natural source / Source: (natural) Homo sapiens (human) / Cell line: HEK293 EBNA / References: UniProt: Q9BQC6 |
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#36: Protein | Mass: 23674.203 Da / Num. of mol.: 1 / Source method: isolated from a natural source / Source: (natural) Homo sapiens (human) / Cell line: HEK293 EBNA / References: UniProt: Q14197, peptidyl-tRNA hydrolase |
#37: Protein | Mass: 25426.895 Da / Num. of mol.: 1 / Source method: isolated from a natural source / Source: (natural) Homo sapiens (human) / Cell line: HEK293 EBNA / References: UniProt: Q8TAE8 |
#57: Protein | Mass: 56258.391 Da / Num. of mol.: 1 / Mutation: 311GGQ313 > AAQ Source method: isolated from a genetically manipulated source Details: Human mitochondrial release factor 1 (mtRF1), containing a 311GGQ>AAQ mutation, a C-terminal linker (GGSGGSGGSGGSGGSGG) and a 3xFLAG-tag (DYKDHDGDYKDHDIDYKDDDDK), expressed from a pcDNA3.1(+) ...Details: Human mitochondrial release factor 1 (mtRF1), containing a 311GGQ>AAQ mutation, a C-terminal linker (GGSGGSGGSGGSGGSGG) 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 / Production host: Homo sapiens (human) / References: UniProt: O75570 |
-Non-polymers , 6 types, 267 molecules
#58: Chemical | ChemComp-K / #59: Chemical | ChemComp-MG / #60: Chemical | ChemComp-VAL / | #61: Chemical | #62: Chemical | ChemComp-FS2 / | #63: Chemical | ChemComp-MET / | |
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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: 39S human mitochondrial ribosome with mtRF1 and P-site tRNA Type: RIBOSOME Details: 39S human mitochondrial small ribosomal subunit in complex with mtRF1(AAQ)-3xFLAG, human methionine-tRNA(Met), and an mRNA containing a non-canonical stop codon, purified from HEK293 EBNA cells. Entity ID: #1-#57 / Source: NATURAL | |||||||||||||||||||||||||
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Molecular weight |
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Source (natural) |
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Source (recombinant) |
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Buffer solution | pH: 7.5 | |||||||||||||||||||||||||
Buffer component |
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Specimen | Embedding applied: NO / Shadowing applied: NO / Staining applied: NO / Vitrification applied: YES Details: Affinity-purified 55S mitoribosome. Refinement focused on 39S large 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 FIELDBright-field microscopy / Nominal magnification: 81000 X / Nominal defocus max: 3000 nm / Nominal defocus min: 600 nm / Cs: 2.7 mm / C2 aperture diameter: 100 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 (6k x 4k) / Num. of grids imaged: 2 / Num. of real images: 43882 |
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. | ||||||||||||||||||||||||||||||||||||
CTF correction | Type: PHASE FLIPPING AND AMPLITUDE CORRECTION | ||||||||||||||||||||||||||||||||||||
Particle selection | Num. of particles selected: 1471587 Details: Particles were picked with blob picker in Cryosparc. | ||||||||||||||||||||||||||||||||||||
3D reconstruction | Resolution: 2.9 Å / Resolution method: FSC 0.143 CUT-OFF / Num. of particles: 41288 / Symmetry type: POINT | ||||||||||||||||||||||||||||||||||||
Atomic model building | Protocol: FLEXIBLE FIT / Space: REAL | ||||||||||||||||||||||||||||||||||||
Atomic model building | 3D fitting-ID: 1
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