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- PDB-7n8y: Oxidized PheRS G318W from Salmonella enterica serovar Typhimurium -

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Basic information

Entry
Database: PDB / ID: 7n8y
TitleOxidized PheRS G318W from Salmonella enterica serovar Typhimurium
Components
  • Phenylalanine--tRNA ligase alpha subunit
  • Phenylalanine--tRNA ligase beta subunit
KeywordsRNA BINDING PROTEIN / LIGASE / synthetase / tRNA-binding protein / tetrameric / oxidized
Function / homology
Function and homology information


phenylalanine-tRNA ligase / phenylalanine-tRNA ligase activity / phenylalanyl-tRNA aminoacylation / tRNA binding / magnesium ion binding / ATP binding / cytoplasm
Similarity search - Function
Phenylalanyl-tRNA synthetase, class IIc, mitochondrial / Phenylalanine-tRNA ligase, class II, N-terminal / Phenylalanine-tRNA ligase alpha chain 1, bacterial / Aminoacyl tRNA synthetase class II, N-terminal domain / Phenylalanine-tRNA ligase, class IIc, beta subunit, bacterial type / Phenylalanyl-tRNA synthetase, class IIc, alpha subunit / Phenylalanly tRNA synthetase, tRNA-binding-domain / tRNA synthetase, B5-domain / tRNA synthetase B5 domain / B5 domain profile. ...Phenylalanyl-tRNA synthetase, class IIc, mitochondrial / Phenylalanine-tRNA ligase, class II, N-terminal / Phenylalanine-tRNA ligase alpha chain 1, bacterial / Aminoacyl tRNA synthetase class II, N-terminal domain / Phenylalanine-tRNA ligase, class IIc, beta subunit, bacterial type / Phenylalanyl-tRNA synthetase, class IIc, alpha subunit / Phenylalanly tRNA synthetase, tRNA-binding-domain / tRNA synthetase, B5-domain / tRNA synthetase B5 domain / B5 domain profile. / tRNA synthetase B5 domain / B3/4 domain / Ferrodoxin-fold anticodon-binding domain / Ferrodoxin-fold anticodon-binding domain superfamily / Ferredoxin-fold anticodon binding domain / Ferredoxin-fold anticodon binding (FDX-ACB) domain profile. / Ferredoxin-fold anticodon binding domain / B3/B4 tRNA-binding domain / Phenylalanyl-tRNA synthetase-like, B3/B4 / Phenylalanyl tRNA synthetase beta chain, core domain / Phenylalanyl tRNA synthetase beta chain CLM domain / B3/4 domain / Class I and II aminoacyl-tRNA synthetase, tRNA-binding arm / Phenylalanyl-tRNA synthetase / tRNA synthetases class II core domain (F) / tRNA-binding domain / Putative tRNA binding domain / tRNA-binding domain profile. / Putative DNA-binding domain superfamily / Aminoacyl-tRNA synthetase, class II / Aminoacyl-transfer RNA synthetases class-II family profile. / Nucleic acid-binding, OB-fold
Similarity search - Domain/homology
Phenylalanine--tRNA ligase beta subunit / Phenylalanine--tRNA ligase alpha subunit
Similarity search - Component
Biological speciesSalmonella enterica subsp. enterica serovar Typhimurium (bacteria)
MethodELECTRON MICROSCOPY / single particle reconstruction / cryo EM / Resolution: 3.65 Å
AuthorsSrinivas, P. / Dunham, C.M.
Funding support United States, 3items
OrganizationGrant numberCountry
National Institutes of Health/National Institute of General Medical Sciences (NIH/NIGMS)R01GM065183 United States
National Institutes of Health/National Institute Of Allergy and Infectious Diseases (NIH/NIAID)T32AI106699 United States
National Institutes of Health/National Institute of General Medical Sciences (NIH/NIGMS)T32GM008602 United States
CitationJournal: Nucleic Acids Res / Year: 2021
Title: Oxidation alters the architecture of the phenylalanyl-tRNA synthetase editing domain to confer hyperaccuracy.
Authors: Pooja Srinivas / Rebecca E Steiner / Ian J Pavelich / Ricardo Guerrero-Ferreira / Puneet Juneja / Michael Ibba / Christine M Dunham /
Abstract: High fidelity during protein synthesis is accomplished by aminoacyl-tRNA synthetases (aaRSs). These enzymes ligate an amino acid to a cognate tRNA and have proofreading and editing capabilities that ...High fidelity during protein synthesis is accomplished by aminoacyl-tRNA synthetases (aaRSs). These enzymes ligate an amino acid to a cognate tRNA and have proofreading and editing capabilities that ensure high fidelity. Phenylalanyl-tRNA synthetase (PheRS) preferentially ligates a phenylalanine to a tRNAPhe over the chemically similar tyrosine, which differs from phenylalanine by a single hydroxyl group. In bacteria that undergo exposure to oxidative stress such as Salmonella enterica serovar Typhimurium, tyrosine isomer levels increase due to phenylalanine oxidation. Several residues are oxidized in PheRS and contribute to hyperactive editing, including against mischarged Tyr-tRNAPhe, despite these oxidized residues not being directly implicated in PheRS activity. Here, we solve a 3.6 Å cryo-electron microscopy structure of oxidized S. Typhimurium PheRS. We find that oxidation results in widespread structural rearrangements in the β-subunit editing domain and enlargement of its editing domain. Oxidization also enlarges the phenylalanyl-adenylate binding pocket but to a lesser extent. Together, these changes likely explain why oxidation leads to hyperaccurate editing and decreased misincorporation of tyrosine. Taken together, these results help increase our understanding of the survival of S. Typhimurium during human infection.
History
DepositionJun 16, 2021Deposition site: RCSB / Processing site: RCSB
Revision 1.0Oct 6, 2021Provider: repository / Type: Initial release
Revision 1.1Oct 13, 2021Group: Database references / Category: citation / Item: _citation.pdbx_database_id_PubMed / _citation.title
Revision 1.2Dec 1, 2021Group: Database references / Category: citation
Item: _citation.journal_volume / _citation.page_first / _citation.page_last
Revision 1.3Jun 5, 2024Group: Data collection / Category: chem_comp_atom / chem_comp_bond

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Structure visualization

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Structure viewerMolecule:
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Assembly

Deposited unit
A: Phenylalanine--tRNA ligase alpha subunit
B: Phenylalanine--tRNA ligase beta subunit
C: Phenylalanine--tRNA ligase alpha subunit
D: Phenylalanine--tRNA ligase beta subunit


Theoretical massNumber of molelcules
Total (without water)248,5694
Polymers248,5694
Non-polymers00
Water00
1


  • Idetical with deposited unit
  • defined by author
TypeNameSymmetry operationNumber
identity operation1_5551

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Components

#1: Protein Phenylalanine--tRNA ligase alpha subunit / Phenylalanyl-tRNA synthetase alpha subunit / PheRS


Mass: 36799.645 Da / Num. of mol.: 2
Source method: isolated from a genetically manipulated source
Source: (gene. exp.) Salmonella enterica subsp. enterica serovar Typhimurium (bacteria)
Gene: pheS / Production host: Escherichia coli BL21(DE3) (bacteria) / References: UniProt: A0A0F7J874, phenylalanine-tRNA ligase
#2: Protein Phenylalanine--tRNA ligase beta subunit / Phenylalanyl-tRNA synthetase beta subunit / PheRS


Mass: 87484.883 Da / Num. of mol.: 2 / Mutation: G318W
Source method: isolated from a genetically manipulated source
Source: (gene. exp.) Salmonella enterica subsp. enterica serovar Typhimurium (bacteria)
Gene: pheT / Production host: Escherichia coli BL21(DE3) (bacteria) / References: UniProt: A0A0D6GBX6, phenylalanine-tRNA ligase

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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: apo-PheRS tetramer structure / Type: COMPLEX / Entity ID: all / Source: RECOMBINANT
Source (natural)Organism: Salmonella enterica subsp. enterica serovar Typhimurium (bacteria)
Source (recombinant)Organism: Escherichia coli BL21(DE3) (bacteria)
Buffer solutionpH: 7.5
SpecimenEmbedding applied: NO / Shadowing applied: NO / Staining applied: NO / Vitrification applied: YES
Specimen supportGrid material: COPPER / Grid mesh size: 300 divisions/in. / Grid type: Quantifoil R1.2/1.3
VitrificationInstrument: FEI VITROBOT MARK IV / Cryogen name: ETHANE / Humidity: 100 % / Chamber temperature: 277 K

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Electron microscopy imaging

Experimental equipment
Model: Talos Arctica / Image courtesy: FEI Company
MicroscopyModel: FEI TALOS ARCTICA
Electron gunElectron source: FIELD EMISSION GUN / Accelerating voltage: 200 kV / Illumination mode: FLOOD BEAM
Electron lensMode: BRIGHT FIELD
Specimen holderCryogen: NITROGEN
Image recordingAverage exposure time: 12 sec. / Electron dose: 64.67 e/Å2 / Film or detector model: GATAN K2 SUMMIT (4k x 4k)

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Processing

SoftwareName: PHENIX / Version: 1.17.1_3660: / Classification: refinement
EM software
IDNameVersionCategory
4GctfCTF correction
13RELION33D reconstruction
CTF correctionType: PHASE FLIPPING AND AMPLITUDE CORRECTION
3D reconstructionResolution: 3.65 Å / Resolution method: FSC 0.143 CUT-OFF / Num. of particles: 204902 / Symmetry type: POINT
Refine LS restraints
Refine-IDTypeDev idealNumber
ELECTRON MICROSCOPYf_bond_d0.00816441
ELECTRON MICROSCOPYf_angle_d0.88722297
ELECTRON MICROSCOPYf_dihedral_angle_d18.3642252
ELECTRON MICROSCOPYf_chiral_restr0.0512530
ELECTRON MICROSCOPYf_plane_restr0.0052943

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