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- PDB-8oip: 28S mammalian mitochondrial small ribosomal subunit with mtRF1 an... -

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Basic information

Entry
Database: PDB / ID: 8oip
Title28S mammalian mitochondrial small ribosomal subunit with mtRF1 and P-site tRNA
Components
  • (28S ribosomal protein ...) x 7
  • (Mitochondrial ribosomal protein ...) x 14
  • 12S rRNA
  • 39S ribosomal protein L19, mitochondrial
  • Aurora kinase A interacting protein 1
  • Coiled-coil-helix-coiled-coil-helix domain containing 1
  • Death associated protein 3
  • Mitoribosomal protein ms28, mrps28
  • P-site Met-tRNA(fMet)
  • Peptide chain release factor 1, mitochondrial
  • bL31m
  • bS6m,Mitochondrial ribosomal protein S6
  • mRNA
  • mS23
  • mS39
  • uS17m
  • uS9m
KeywordsRIBOSOME / 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 / ribosomal small subunit assembly / small ribosomal subunit rRNA binding / regulation of translation / small ribosomal subunit / cytosolic small ribosomal subunit / nuclear membrane / cytosolic large ribosomal subunit / rRNA binding / ribosome / structural constituent of ribosome / mitochondrial matrix / ribonucleoprotein complex / translation / mRNA binding / apoptotic process / synapse / nucleolus / mitochondrion / extracellular space / RNA binding / nucleoplasm
Similarity search - Function
Gonadotropin-releasing hormone / Gonadoliberin / Gonadotropin-releasing hormone / Gonadotropin-releasing hormones signature. / Coiled-coil-helix-coiled-coil-helix domain-containing protein 1 / Cysteine alpha-hairpin motif superfamily / Peptide chain release factor / PCRF domain / PCRF / 28S ribosomal protein S27, mitochondrial ...Gonadotropin-releasing hormone / Gonadoliberin / Gonadotropin-releasing hormone / Gonadotropin-releasing hormones signature. / Coiled-coil-helix-coiled-coil-helix domain-containing protein 1 / Cysteine alpha-hairpin motif superfamily / Peptide chain release factor / PCRF domain / PCRF / 28S ribosomal protein S27, mitochondrial / MRPS27/PTCD2 / Mitochondrial 28S ribosomal protein S27 / Ribosomal protein S22, mitochondrial / Ribosomal protein S23, mitochondrial / Ribosomal protein S23/S25, mitochondrial / Mitochondrial 28S ribosomal protein S31 / Mitochondrial 28S ribosomal protein S22 / Mitochondrial ribosomal protein S23 / Mitochondrial 28S ribosomal protein S31 / Ribosomal protein S29, mitochondrial / Ribosomal protein S28, mitochondrial / 28S ribosomal protein S10, mitochondrial / Mitochondrial ribosomal protein MRP-S35 / Peptide chain release factor class I superfamily / 28S ribosomal protein S24, mitochondrial / Mitochondrial 28S ribosomal protein S34 / Pentatricopeptide repeat domain-containing protein 3 / 28S ribosomal protein S17, mitochondrial / 28S ribosomal protein S18b, mitochondrial / : / Mitochondrial ribosome subunit S24 / Mitochondrial 28S ribosomal protein S34 / Small ribosomal subunit protein uS5m, N-terminal / 28S ribosomal protein S25, mitochondrial / Prokaryotic-type class I peptide chain release factors signature. / Pentatricopeptide repeat domain / Ribosomal protein L55, mitochondrial / Mitochondrial ribosomal protein L55 superfamily / Mitochondrial ribosomal protein L55 / Ribosomal protein S27/S33, mitochondrial / Ribosomal protein S24/S35, mitochondrial / Mitochondrial ribosomal subunit S27 / Ribosomal protein S24/S35, mitochondrial, conserved domain / Mitochondrial ribosomal subunit protein / Pentatricopeptide (PPR) repeat profile. / Ribosomal protein S23/S29, mitochondrial / Mitochondrial ribosomal death-associated protein 3 / Pentatricopeptide repeat / Mitochondrial mRNA-processing protein COX24, C-terminal / Mitochondrial mRNA-processing protein COX24, C-terminal / Mitochondrial domain of unknown function (DUF1713) / Peptide chain release factor class I / RF-1 domain / CHCH / CHCH domain / Ribosomal protein/NADH dehydrogenase domain / Mitochondrial ribosomal protein L51 / S25 / CI-B8 domain / Mitochondrial ribosomal protein L51 / S25 / CI-B8 domain / Ribosomal protein S21 superfamily / Ribosomal protein S21 / Ribosomal protein S21 / Ribosomal protein S2, bacteria/mitochondria/plastid / Ribosomal protein S18, conserved site / Ribosomal protein S18 signature. / Ribosomal protein S16 / Ribosomal protein S16 / Ribosomal protein S16 domain superfamily / Ribosomal protein S15, bacterial-type / Ribosomal protein L19 / Ribosomal protein L19 superfamily / Ribosomal protein L19 / Ribosomal protein S6 / Ribosomal protein S6 / Ribosomal protein S6 superfamily / Ribosomal protein S12, bacterial-type / Translation elongation factor EF1B/ribosomal protein S6 / Ribosomal protein S18 / Ribosomal protein S18 / Ribosomal protein S18 superfamily / Ribosomal protein S2 signature 1. / Ribosomal protein S2, conserved site / Ribosomal protein S5, N-terminal, conserved site / Ribosomal protein S5 signature. / Ribosomal protein S2 / Ribosomal protein S2, flavodoxin-like domain superfamily / Ribosomal protein S2 / S5 double stranded RNA-binding domain profile. / Ribosomal protein S5 / Ribosomal protein S5, N-terminal / Ribosomal protein S5, N-terminal domain / Ribosomal protein S5, C-terminal / Ribosomal protein S5, C-terminal domain / Ribosomal protein S14 / Ribosomal protein S14p/S29e / Ribosomal S11, conserved site / Ribosomal protein S11 / Ribosomal protein S11 / Ribosomal protein S11 signature. / Ribosomal protein S10p/S20e / Ribosomal protein S10 domain
Similarity search - Domain/homology
ADENOSINE-5'-TRIPHOSPHATE / FE-S-O HYBRID CLUSTER / GUANOSINE-5'-DIPHOSPHATE / : / SPERMIDINE / SPERMINE / : / RNA / RNA (> 10) / RNA (> 100) ...ADENOSINE-5'-TRIPHOSPHATE / FE-S-O HYBRID CLUSTER / GUANOSINE-5'-DIPHOSPHATE / : / SPERMIDINE / SPERMINE / : / RNA / RNA (> 10) / RNA (> 100) / Small ribosomal subunit protein uS12m / 28S ribosomal protein S14, mitochondrial / 28S ribosomal protein S18c, mitochondrial isoform 1 / Small ribosomal subunit protein mS25 / 28S ribosomal protein S28, mitochondrial / Small ribosomal subunit protein uS7m / Mitochondrial ribosomal protein S11 / 28S ribosomal protein S21, mitochondrial / Mitochondrial ribosomal protein S23 / Small ribosomal subunit protein uS15m / 28S ribosomal protein S24, mitochondrial / Mitochondrial ribosomal protein S22 / Mitochondrial ribosomal protein S34 / Progonadoliberin / Small ribosomal subunit protein mS38 / Small ribosomal subunit protein bS16m / Mitochondrial ribosomal protein S6 / Mitochondrial ribosomal protein S27 / Mitochondrial ribosomal protein S17 / Small ribosomal subunit protein mS31 / Small ribosomal subunit protein uS10m / 28S ribosomal protein S2, mitochondrial / Small ribosomal subunit protein mS33 / 28S ribosomal protein S35, mitochondrial isoform 1 / Coiled-coil-helix-coiled-coil-helix domain containing 1 / Small ribosomal subunit protein uS5m / Mitochondrial ribosomal protein L55 / Large ribosomal subunit protein bL19m / Small ribosomal subunit protein mS29 / 28S ribosomal protein S9, mitochondrial / Small ribosomal subunit protein mS39 / Peptide chain release factor 1, mitochondrial / Small ribosomal subunit protein mS40
Similarity search - Component
Biological speciesHomo sapiens (human)
Sus scrofa (pig)
MethodELECTRON MICROSCOPY / single particle reconstruction / cryo EM / Resolution: 3.6 Å
AuthorsSaurer, M. / Leibundgut, M. / Scaiola, A. / Schoenhut, T. / Ban, N.
Funding support Switzerland, European Union, 3items
OrganizationGrant numberCountry
Swiss National Science Foundation179190 Switzerland
Swiss National Science Foundation182341 Switzerland
European Molecular Biology Organization (EMBO)1074-2019European Union
CitationJournal: 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
DepositionMar 23, 2023Deposition site: PDBE / Processing site: PDBE
Revision 1.0May 24, 2023Provider: repository / Type: Initial release
Revision 2.0Nov 15, 2023Group: Atomic model / Data collection ...Atomic model / Data collection / Derived calculations / Refinement description
Category: atom_site / chem_comp_atom ...atom_site / chem_comp_atom / chem_comp_bond / em_3d_fitting_list / pdbx_validate_rmsd_angle / pdbx_validate_torsion / struct_conn
Item: _atom_site.auth_atom_id / _atom_site.label_atom_id ..._atom_site.auth_atom_id / _atom_site.label_atom_id / _em_3d_fitting_list.initial_refinement_model_id / _struct_conn.pdbx_leaving_atom_flag / _struct_conn.ptnr1_label_atom_id
Revision 2.1Apr 24, 2024Group: Data collection / Structure summary / Category: chem_comp / chem_comp_atom / chem_comp_bond
Item: _chem_comp.pdbx_synonyms / _chem_comp_atom.atom_id ..._chem_comp.pdbx_synonyms / _chem_comp_atom.atom_id / _chem_comp_bond.atom_id_1 / _chem_comp_bond.atom_id_2 / _chem_comp_bond.pdbx_aromatic_flag / _chem_comp_bond.value_order

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Structure visualization

Structure viewerMolecule:
MolmilJmol/JSmol

Downloads & links

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Assembly

Deposited unit
BX: 39S ribosomal protein L19, mitochondrial
Bd: bL31m
AA: 12S rRNA
AB: 28S ribosomal protein S35, mitochondrial isoform 1
AC: Mitochondrial ribosomal protein S24
AD: Aurora kinase A interacting protein 1
AE: bS6m,Mitochondrial ribosomal protein S6
AF: Mitochondrial ribosomal protein S7
AG: P-site Met-tRNA(fMet)
AH: Mitochondrial ribosomal protein S10
AI: mRNA
AJ: Mitochondrial ribosomal protein S12
AK: Mitochondrial ribosomal protein S14
AL: 28S ribosomal protein S15, mitochondrial
AM: 28S ribosomal protein S16, mitochondrial
AN: uS17m
AO: 28S ribosomal protein S18b, mitochondrial
AP: Mitochondrial ribosomal protein S18C
AQ: 28S ribosomal protein S21, mitochondrial
AR: Mitochondrial ribosomal protein S22
AS: mS23
AT: Mitochondrial ribosomal protein S25
AU: Mitochondrial ribosomal protein S26
AV: Mitochondrial ribosomal protein S27
AW: Mitoribosomal protein ms28, mrps28
AX: Death associated protein 3
AY: 28S ribosomal protein S31, mitochondrial
AZ: Mitochondrial ribosomal protein S33
Aa: Peptide chain release factor 1, mitochondrial
Ab: Mitochondrial ribosomal protein S2
Ac: Coiled-coil-helix-coiled-coil-helix domain containing 1
Ad: 28S ribosomal protein S5, mitochondrial isoform X2
Ae: mS39
Ag: uS9m
Ai: Mitochondrial ribosomal protein S11
Aj: Mitochondrial ribosomal protein S34
hetero molecules


Theoretical massNumber of molelcules
Total (without water)1,343,831181
Polymers1,338,28036
Non-polymers5,551145
Water543
1


  • Idetical with deposited unit
  • defined by author
  • Evidence: electron microscopy
TypeNameSymmetry operationNumber
identity operation1_5551

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Components

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Protein , 12 types, 12 molecules BXBdADAEANASAWAXAaAcAeAg

#1: Protein 39S ribosomal protein L19, mitochondrial / Mitochondrial ribosomal protein L19


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
#2: Protein bL31m


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 Aurora kinase A interacting protein 1


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 bS6m,Mitochondrial ribosomal protein S6


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 uS17m


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 mS23


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 Mitoribosomal protein ms28, mrps28


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 Death associated protein 3


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 Peptide chain release factor 1, mitochondrial / MRF-1 / MtRF-1


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 Coiled-coil-helix-coiled-coil-helix domain containing 1


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 mS39


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 uS9m


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

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RNA chain , 3 types, 3 molecules AAAGAI

#3: RNA chain 12S rRNA


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
#9: RNA chain P-site Met-tRNA(fMet)


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 mRNA


Mass: 2902.831 Da / Num. of mol.: 1 / Source method: obtained synthetically / Details: Synthetic mRNA / Source: (synth.) Homo sapiens (human)

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28S ribosomal protein ... , 7 types, 7 molecules ABALAMAOAQAYAd

#4: Protein 28S ribosomal protein S35, mitochondrial isoform 1 / Mitochondrial ribosomal protein S35


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
#14: Protein 28S ribosomal protein S15, mitochondrial


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 28S ribosomal protein S16, mitochondrial / Mitochondrial ribosomal protein S16


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 28S ribosomal protein S18b, mitochondrial / S18mt-b / 28S ribosomal protein S18-2 / mitochondrial / MRP-S18-2


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 28S ribosomal protein S21, mitochondrial


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 28S ribosomal protein S31, mitochondrial


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 28S ribosomal protein S5, mitochondrial isoform X2 / Mitochondrial ribosomal protein S5


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

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Mitochondrial ribosomal protein ... , 14 types, 14 molecules ACAFAHAJAKAPARATAUAVAZAbAiAj

#5: Protein Mitochondrial ribosomal protein S24


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
#8: Protein Mitochondrial ribosomal protein S7


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 Mitochondrial ribosomal protein S10


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 Mitochondrial ribosomal protein S12 / Mitoribosomal protein us12m / mrps12


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 Mitochondrial ribosomal protein S14 / Mitoribosomal protein us14m / mrps14


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 Mitochondrial ribosomal protein S18C / Mitoribosomal protein bs18m / mrps18c


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 Mitochondrial ribosomal protein S22


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 Mitochondrial ribosomal protein S25 / Mitoribosomal protein ms25 / mrps25


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 Mitochondrial ribosomal protein S26


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 Mitochondrial ribosomal protein S27


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 Mitochondrial ribosomal protein S33


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 Mitochondrial ribosomal protein S2


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 Mitochondrial ribosomal protein S11


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 Mitochondrial ribosomal protein S34


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

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Non-polymers , 9 types, 148 molecules

#37: Chemical...
ChemComp-MG / MAGNESIUM ION


Mass: 24.305 Da / Num. of mol.: 127 / Source method: obtained synthetically / Formula: Mg
#38: Chemical ChemComp-SPM / SPERMINE


Mass: 202.340 Da / Num. of mol.: 1 / Source method: obtained synthetically / Formula: C10H26N4
#39: Chemical ChemComp-SPD / SPERMIDINE / N-(2-AMINO-PROPYL)-1,4-DIAMINOBUTANE / PA(34)


Mass: 145.246 Da / Num. of mol.: 1 / Source method: obtained synthetically / Formula: C7H19N3
#40: Chemical
ChemComp-K / POTASSIUM ION


Mass: 39.098 Da / Num. of mol.: 11 / Source method: obtained synthetically / Formula: K
#41: Chemical ChemComp-ZN / ZINC ION


Mass: 65.409 Da / Num. of mol.: 1 / Source method: obtained synthetically / Formula: Zn
#42: Chemical ChemComp-FS2 / FE-S-O HYBRID CLUSTER


Mass: 335.508 Da / Num. of mol.: 2 / Source method: obtained synthetically / Formula: Fe4O3S2
#43: Chemical ChemComp-ATP / ADENOSINE-5'-TRIPHOSPHATE


Mass: 507.181 Da / Num. of mol.: 1 / Source method: obtained synthetically / Formula: C10H16N5O13P3 / Comment: ATP, energy-carrying molecule*YM
#44: Chemical ChemComp-GDP / GUANOSINE-5'-DIPHOSPHATE


Type: RNA linking / Mass: 443.201 Da / Num. of mol.: 1 / Source method: obtained synthetically / Formula: C10H15N5O11P2 / Comment: GDP, energy-carrying molecule*YM
#45: Water ChemComp-HOH / water


Mass: 18.015 Da / Num. of mol.: 3 / Source method: isolated from a natural source / Formula: H2O

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Details

Has ligand of interestN

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Experimental details

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Experiment

ExperimentMethod: ELECTRON MICROSCOPY
EM experimentAggregation state: PARTICLE / 3D reconstruction method: single particle reconstruction

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Sample preparation

ComponentName: 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
Molecular weightValue: 1.1 MDa / Experimental value: NO
Source (natural)Organism: Sus scrofa (pig) / Cellular location: Mitochondrial matrix / Organelle: Mitochondria / Tissue: Liver
Buffer solutionpH: 7.5
Details: 20mM HEPES-KOH pH=7.5, 100mM KCl, 20mM MgCl2, ~1% sucrose (from mtRF1 storage buffer)
Buffer component
IDConc.NameFormulaBuffer-ID
120 mMHEPESC8H18N2O4S1
2100 mMpotassium chlorideKCl1
320 mMmagnesium chlorideMgCl21
SpecimenEmbedding applied: NO / Shadowing applied: NO / Staining applied: NO / Vitrification applied: YES
Details: In vitro reconstituted complex (55S). Focused refinement on small subunit.
Specimen supportGrid material: COPPER / Grid type: Quantifoil R2/2
VitrificationInstrument: FEI VITROBOT MARK IV / Cryogen name: ETHANE-PROPANE / Humidity: 100 % / Chamber temperature: 277.15 K

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Electron microscopy imaging

Experimental equipment
Model: Titan Krios / Image courtesy: FEI Company
MicroscopyModel: FEI TITAN KRIOS
Electron gunElectron source: FIELD EMISSION GUN / Accelerating voltage: 300 kV / Illumination mode: FLOOD BEAM
Electron lensMode: 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 holderCryogen: NITROGEN / Specimen holder model: FEI TITAN KRIOS AUTOGRID HOLDER
Image recordingElectron 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 opticsEnergyfilter slit width: 20 eV

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Processing

EM software
IDNameVersionCategory
1cryoSPARC4particle selection
2EPUimage acquisition
4cryoSPARC4CTF correction
7Coot0.9.8.4model fitting
9cryoSPARC4initial Euler assignment
10cryoSPARC4final Euler assignment
12cryoSPARC43D reconstruction
13PHENIX1.20.1model refinement
Image processingDetails: Micrographs were motion corrected and CTF parameters were estimated using Cryosparc live v4.
CTF correctionType: PHASE FLIPPING AND AMPLITUDE CORRECTION
Particle selectionNum. of particles selected: 1471587
Details: Particles were picked with blob picker in Cryosparc live v4.
3D reconstructionResolution: 3.6 Å / Resolution method: FSC 0.143 CUT-OFF / Num. of particles: 50622 / Symmetry type: POINT
Atomic model buildingProtocol: FLEXIBLE FIT / Space: REAL
Atomic model building

3D fitting-ID: 1

IDPDB-IDAccession codeDetailsInitial refinement model-IDSource nameType
17NQH

7nqh

7NQHSus scrofa mitoribosome1PDBexperimental model
27QI4

7qi4

7QI4High-res mammalian mitoribosome2PDBexperimental model
3UniProt AF-O75570-F1mtRF1 model prediction3AlphaFoldin silico model

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