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- PDB-9bfr: Cryo-EM structure of Bride of Sevenless extracellular domain (dim... -

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Basic information

Entry
Database: PDB / ID: 9bfr
TitleCryo-EM structure of Bride of Sevenless extracellular domain (dimer, Sevenless-bound form)
ComponentsProtein bride of sevenless
KeywordsSIGNALING PROTEIN / bride of sevenless / boss / GPCR / G-protein coupled receptor / drosophila / eye development / vision / photoreceptor / glucose metabolism
Function / homology
Function and homology information


R8 cell-mediated photoreceptor organization / rhabdomere microvillus membrane / sevenless binding / germ-line stem-cell niche homeostasis / R7 cell fate commitment / sevenless signaling pathway / adenylate cyclase inhibiting G protein-coupled glutamate receptor activity / lipid homeostasis / response to glucose / visual perception ...R8 cell-mediated photoreceptor organization / rhabdomere microvillus membrane / sevenless binding / germ-line stem-cell niche homeostasis / R7 cell fate commitment / sevenless signaling pathway / adenylate cyclase inhibiting G protein-coupled glutamate receptor activity / lipid homeostasis / response to glucose / visual perception / glucose homeostasis / receptor ligand activity / plasma membrane
Similarity search - Function
Bride of sevenless protein / : / GPCR family 3, C-terminal / 7 transmembrane sweet-taste receptor of 3 GCPR
Similarity search - Domain/homology
Protein bride of sevenless
Similarity search - Component
Biological speciesDrosophila melanogaster (fruit fly)
MethodELECTRON MICROSCOPY / single particle reconstruction / cryo EM / Resolution: 3.19 Å
AuthorsCerutti, G. / Shapiro, L.
Funding support United States, 1items
OrganizationGrant numberCountry
Other private United States
CitationJournal: Mol Cell / Year: 2024
Title: Structures and pH-dependent dimerization of the sevenless receptor tyrosine kinase.
Authors: Gabriele Cerutti / Ronald Arias / Fabiana Bahna / Seetha Mannepalli / Phinikoula S Katsamba / Goran Ahlsen / Brian Kloss / Renato Bruni / Andrew Tomlinson / Lawrence Shapiro /
Abstract: Sevenless (Sev) is a Drosophila receptor tyrosine kinase (RTK) required for the specification of the R7 photoreceptor. It is cleaved into α and β subunits and binds the ectodomain of the G-protein- ...Sevenless (Sev) is a Drosophila receptor tyrosine kinase (RTK) required for the specification of the R7 photoreceptor. It is cleaved into α and β subunits and binds the ectodomain of the G-protein-coupled receptor bride of sevenless (Boss). Previous work showed that the Boss ectodomain could bind but not activate Sev; rather, the whole seven-pass transmembrane Boss was required. Here, we show that Sev does not need to be cleaved to function and that a single-pass transmembrane form of Boss activates Sev. We use cryo-electron microscopy and biophysical methods to determine the structural basis of ligand binding and pH-dependent dimerization of Sev, and we discuss the implications in the process of Sev activation. The Sev human homolog, receptor oncogene from sarcoma 1 (ROS1), is associated with oncogenic transformations, and we discuss their structural similarities.
History
DepositionApr 18, 2024Deposition site: RCSB / Processing site: RCSB
Revision 1.0Jan 29, 2025Provider: repository / Type: Initial release
Revision 1.0Jan 29, 2025Data content type: EM metadata / Data content type: EM metadata / Provider: repository / Type: Initial release
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Revision 1.0Jan 29, 2025Data content type: Image / Data content type: Image / Provider: repository / Type: Initial release
Revision 1.0Jan 29, 2025Data content type: Primary map / Data content type: Primary map / Provider: repository / Type: Initial release
Revision 1.1May 14, 2025Group: Data collection / Category: em_admin / em_software / Item: _em_admin.last_update / _em_software.name
Revision 1.1May 14, 2025Data content type: EM metadata / Data content type: EM metadata / EM metadata / Group: Data processing / Experimental summary / Data content type: EM metadata / EM metadata / Category: em_admin / em_software / Data content type: EM metadata / EM metadata / Item: _em_admin.last_update / _em_software.name

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Structure visualization

Structure viewerMolecule:
MolmilJmol/JSmol

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Assembly

Deposited unit
A: Protein bride of sevenless
B: Protein bride of sevenless
hetero molecules


Theoretical massNumber of molelcules
Total (without water)115,8858
Polymers114,5572
Non-polymers1,3276
Water00
1


  • Idetical with deposited unit
  • defined by author
  • Evidence: electron microscopy, not applicable
TypeNameSymmetry operationNumber
identity operation1_5551

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Components

#1: Protein Protein bride of sevenless


Mass: 57278.723 Da / Num. of mol.: 2
Source method: isolated from a genetically manipulated source
Source: (gene. exp.) Drosophila melanogaster (fruit fly) / Gene: boss, CG8285 / Production host: Homo sapiens (human) / References: UniProt: P22815
#2: 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.: 6 / Source method: obtained synthetically / Formula: C8H15NO6 / Feature type: SUBJECT OF INVESTIGATION
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
Has ligand of interestY
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: Bride of Sevenless extracellular domain / Type: ORGANELLE OR CELLULAR COMPONENT / Entity ID: #1 / Source: MULTIPLE SOURCES
Molecular weightExperimental value: NO
Source (natural)Organism: Drosophila melanogaster (fruit fly)
Source (recombinant)Organism: Drosophila melanogaster (fruit fly)
Buffer solutionpH: 6.6
SpecimenEmbedding applied: NO / Shadowing applied: NO / Staining applied: NO / Vitrification applied: YES
Specimen supportGrid material: GOLD
VitrificationCryogen name: ETHANE

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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: OTHER
Electron lensMode: BRIGHT FIELD / Nominal defocus max: 2000 nm / Nominal defocus min: 800 nm
Image recordingElectron dose: 51.8 e/Å2 / Film or detector model: GATAN K3 BIOQUANTUM (6k x 4k)

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Processing

EM softwareName: PHENIX / Category: model refinement
CTF correctionType: PHASE FLIPPING AND AMPLITUDE CORRECTION
SymmetryPoint symmetry: C2 (2 fold cyclic)
3D reconstructionResolution: 3.19 Å / Resolution method: FSC 0.143 CUT-OFF / Num. of particles: 187282 / Symmetry type: POINT
Refine LS restraints
Refine-IDTypeDev idealNumber
ELECTRON MICROSCOPYf_bond_d0.0066382
ELECTRON MICROSCOPYf_angle_d1.0178636
ELECTRON MICROSCOPYf_dihedral_angle_d7.467936
ELECTRON MICROSCOPYf_chiral_restr0.061012
ELECTRON MICROSCOPYf_plane_restr0.0081086

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