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- PDB-7qa8: Structure of the GPCR dimer Ste2 bound to an antagonist -

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Basic information

Entry
Database: PDB / ID: 7qa8
TitleStructure of the GPCR dimer Ste2 bound to an antagonist
Components
  • HIS-ALA-LEU-GLN-LEU-LYS-PRO-GLY-GLN-PRO-NLE-TYR
  • Pheromone alpha factor receptor
KeywordsMEMBRANE PROTEIN / GPCR / dimer / antagonist-bound / fungal
Function / homologymating-type factor pheromone receptor activity / GPCR fungal pheromone mating factor, STE2 / Pheromone alpha factor receptor, double transmembrane domain superfamily / Fungal pheromone mating factor STE2 GPCR / response to pheromone / membrane / CHOLESTEROL HEMISUCCINATE / Pheromone alpha factor receptor
Function and homology information
Biological speciesSaccharomyces cerevisiae (brewer's yeast)
MethodELECTRON MICROSCOPY / single particle reconstruction / cryo EM / Resolution: 2.7 Å
AuthorsVelazhahan, V. / Tate, C.G.
Funding support United Kingdom, 1items
OrganizationGrant numberCountry
Medical Research Council (MRC, United Kingdom)MRC U105197215 United Kingdom
CitationJournal: Nature / Year: 2021
Title: Structure of the class D GPCR Ste2 dimer coupled to two G proteins.
Authors: Vaithish Velazhahan / Ning Ma / Gáspár Pándy-Szekeres / Albert J Kooistra / Yang Lee / David E Gloriam / Nagarajan Vaidehi / Christopher G Tate /
Abstract: G-protein-coupled receptors (GPCRs) are divided phylogenetically into six classes, denoted A to F. More than 370 structures of vertebrate GPCRs (belonging to classes A, B, C and F) have been ...G-protein-coupled receptors (GPCRs) are divided phylogenetically into six classes, denoted A to F. More than 370 structures of vertebrate GPCRs (belonging to classes A, B, C and F) have been determined, leading to a substantial understanding of their function. By contrast, there are no structures of class D GPCRs, which are found exclusively in fungi where they regulate survival and reproduction. Here we determine the structure of a class D GPCR, the Saccharomyces cerevisiae pheromone receptor Ste2, in an active state coupled to the heterotrimeric G protein Gpa1-Ste4-Ste18. Ste2 was purified as a homodimer coupled to two G proteins. The dimer interface of Ste2 is formed by the N terminus, the transmembrane helices H1, H2 and H7, and the first extracellular loop ECL1. We establish a class D1 generic residue numbering system (CD1) to enable comparisons with orthologues and with other GPCR classes. The structure of Ste2 bears similarities in overall topology to class A GPCRs, but the transmembrane helix H4 is shifted by more than 20 Å and the G-protein-binding site is a shallow groove rather than a cleft. The structure provides a template for the design of novel drugs to target fungal GPCRs, which could be used to treat numerous intractable fungal diseases.
History
DepositionNov 16, 2021Deposition site: PDBE / Processing site: PDBE
Revision 1.0Mar 16, 2022Provider: repository / Type: Initial release
Revision 1.1Mar 30, 2022Group: Database references / Category: citation / citation_author
Item: _citation.pdbx_database_id_PubMed / _citation.title / _citation_author.identifier_ORCID
Revision 1.2Apr 6, 2022Group: Database references / Category: citation
Item: _citation.journal_volume / _citation.page_first / _citation.page_last
Revision 1.3Oct 16, 2024Group: Data collection / Structure summary
Category: chem_comp_atom / chem_comp_bond ...chem_comp_atom / chem_comp_bond / em_admin / pdbx_entry_details / pdbx_modification_feature
Item: _em_admin.last_update

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Structure visualization

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Assembly

Deposited unit
A: Pheromone alpha factor receptor
K: HIS-ALA-LEU-GLN-LEU-LYS-PRO-GLY-GLN-PRO-NLE-TYR
I: HIS-ALA-LEU-GLN-LEU-LYS-PRO-GLY-GLN-PRO-NLE-TYR
B: Pheromone alpha factor receptor
hetero molecules


Theoretical massNumber of molelcules
Total (without water)115,93842
Polymers98,5044
Non-polymers17,43438
Water1,26170
1


  • Idetical with deposited unit
  • defined by author
  • Evidence: gel filtration
TypeNameSymmetry operationNumber
identity operation1_5551
Buried area12470 Å2
ΔGint-66 kcal/mol
Surface area33820 Å2

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Components

#1: Protein Pheromone alpha factor receptor


Mass: 47885.402 Da / Num. of mol.: 2
Source method: isolated from a genetically manipulated source
Source: (gene. exp.) Saccharomyces cerevisiae (brewer's yeast)
Gene: STE2 / Production host: Trichoplusia ni (cabbage looper) / References: UniProt: P0CI39
#2: Protein/peptide HIS-ALA-LEU-GLN-LEU-LYS-PRO-GLY-GLN-PRO-NLE-TYR


Mass: 1366.606 Da / Num. of mol.: 2 / Source method: obtained synthetically / Source: (synth.) Saccharomyces cerevisiae (brewer's yeast)
#3: Sugar
ChemComp-NAG / 2-acetamido-2-deoxy-beta-D-glucopyranose / N-acetyl-beta-D-glucosamine / 2-acetamido-2-deoxy-beta-D-glucose / 2-acetamido-2-deoxy-D-glucose / 2-acetamido-2-deoxy-glucose / N-ACETYL-D-GLUCOSAMINE


Type: D-saccharide, beta linking / Mass: 221.208 Da / Num. of mol.: 4 / Source method: obtained synthetically / Formula: C8H15NO6
IdentifierTypeProgram
DGlcpNAcbCONDENSED IUPAC CARBOHYDRATE SYMBOLGMML 1.0
N-acetyl-b-D-glucopyranosamineCOMMON NAMEGMML 1.0
b-D-GlcpNAcIUPAC CARBOHYDRATE SYMBOLPDB-CARE 1.0
GlcNAcSNFG CARBOHYDRATE SYMBOLGMML 1.0
#4: Chemical...
ChemComp-Y01 / CHOLESTEROL HEMISUCCINATE


Mass: 486.726 Da / Num. of mol.: 34 / Source method: obtained synthetically / Formula: C31H50O4
#5: Water ChemComp-HOH / water


Mass: 18.015 Da / Num. of mol.: 70 / Source method: isolated from a natural source / Formula: H2O
Has protein modificationY

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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: GPCR dimer Ste2 bound to an antagonist / Type: COMPLEX / Entity ID: #1-#2 / Source: RECOMBINANT
Molecular weightExperimental value: NO
Source (natural)Organism: Saccharomyces cerevisiae (brewer's yeast)
Source (recombinant)Organism: Trichoplusia ni (cabbage looper)
Buffer solutionpH: 7.5
SpecimenConc.: 3.1 mg/ml / Embedding applied: NO / Shadowing applied: NO / Staining applied: NO / Vitrification applied: YES
VitrificationInstrument: FEI VITROBOT MARK IV / 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
MicroscopyModel: TFS KRIOS
Electron gunElectron source: FIELD EMISSION GUN / Accelerating voltage: 300 kV / Illumination mode: FLOOD BEAM
Electron lensMode: BRIGHT FIELD / Cs: 2.7 mm / C2 aperture diameter: 50 µm
Image recordingElectron dose: 57 e/Å2 / Film or detector model: GATAN K3 (6k x 4k)

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Processing

SoftwareName: PHENIX / Version: 1.19.2_4158: / Classification: refinement
EM software
IDNameVersionCategory
10RELION3.1initial Euler assignment
11RELION3.1final Euler assignment
CTF correctionType: NONE
SymmetryPoint symmetry: C2 (2 fold cyclic)
3D reconstructionResolution: 2.7 Å / Resolution method: FSC 0.143 CUT-OFF / Num. of particles: 136877 / Symmetry type: POINT
Refine LS restraints
Refine-IDTypeDev idealNumber
ELECTRON MICROSCOPYf_bond_d0.0056243
ELECTRON MICROSCOPYf_angle_d0.9188682
ELECTRON MICROSCOPYf_dihedral_angle_d15.5671688
ELECTRON MICROSCOPYf_chiral_restr0.0551156
ELECTRON MICROSCOPYf_plane_restr0.009913

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