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- PDB-9bfs: Cryo-EM structure of Sevenless extracellular domain (dimer, pH 6.6) -

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Basic information

Entry
Database: PDB / ID: 9bfs
TitleCryo-EM structure of Sevenless extracellular domain (dimer, pH 6.6)
ComponentsProtein sevenless
KeywordsSIGNALING PROTEIN / sevenless / receptor tyrosine kinase / RTK / drosophila / eye development / vision / photoreceptor / ROS1
Function / homology
Function and homology information


germ-line stem-cell niche homeostasis / transmembrane receptor protein kinase activity / R7 cell fate commitment / sevenless signaling pathway / regulation of TOR signaling / transmembrane receptor protein tyrosine kinase activity / visual perception / cell surface receptor protein tyrosine kinase signaling pathway / placental growth factor receptor activity / insulin receptor activity ...germ-line stem-cell niche homeostasis / transmembrane receptor protein kinase activity / R7 cell fate commitment / sevenless signaling pathway / regulation of TOR signaling / transmembrane receptor protein tyrosine kinase activity / visual perception / cell surface receptor protein tyrosine kinase signaling pathway / placental growth factor receptor activity / insulin receptor activity / hepatocyte growth factor receptor activity / macrophage colony-stimulating factor receptor activity / platelet-derived growth factor alpha-receptor activity / platelet-derived growth factor beta-receptor activity / stem cell factor receptor activity / boss receptor activity / protein tyrosine kinase collagen receptor activity / brain-derived neurotrophic factor receptor activity / transmembrane-ephrin receptor activity / GPI-linked ephrin receptor activity / epidermal growth factor receptor activity / fibroblast growth factor receptor activity / insulin-like growth factor receptor activity / receptor protein-tyrosine kinase / receptor complex / ATP binding / plasma membrane
Similarity search - Function
LDLR class B repeat / Low-density lipoprotein-receptor YWTD domain / Tyrosine-protein kinase, receptor class II, conserved site / Receptor tyrosine kinase class II signature. / Six-bladed beta-propeller, TolB-like / Fibronectin type III domain / : / Fibronectin type 3 domain / Fibronectin type-III domain profile. / Fibronectin type III ...LDLR class B repeat / Low-density lipoprotein-receptor YWTD domain / Tyrosine-protein kinase, receptor class II, conserved site / Receptor tyrosine kinase class II signature. / Six-bladed beta-propeller, TolB-like / Fibronectin type III domain / : / Fibronectin type 3 domain / Fibronectin type-III domain profile. / Fibronectin type III / Fibronectin type III superfamily / Tyrosine-protein kinase, catalytic domain / Tyrosine kinase, catalytic domain / Tyrosine protein kinases specific active-site signature. / Tyrosine-protein kinase, active site / Serine-threonine/tyrosine-protein kinase, catalytic domain / Protein tyrosine and serine/threonine kinase / Protein kinase, ATP binding site / Protein kinases ATP-binding region signature. / Immunoglobulin-like fold / Protein kinase domain profile. / Protein kinase domain / Protein kinase-like domain superfamily
Similarity search - Domain/homology
Biological speciesDrosophila melanogaster (fruit fly)
MethodELECTRON MICROSCOPY / single particle reconstruction / cryo EM / Resolution: 3.06 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
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Revision 1.0Jan 29, 2025Data content type: Image / Data content type: Image / Provider: repository / Type: Initial release
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Revision 1.1May 21, 2025Group: Data collection / Category: em_admin / em_software / Item: _em_admin.last_update / _em_software.name
Revision 1.1May 21, 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

Downloads & links

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Assembly

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


Theoretical massNumber of molelcules
Total (without water)456,11522
Polymers450,5862
Non-polymers5,52920
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 sevenless


Mass: 225292.922 Da / Num. of mol.: 2
Source method: isolated from a genetically manipulated source
Source: (gene. exp.) Drosophila melanogaster (fruit fly) / Gene: sev, HD-265, CG18085 / Production host: Drosophila melanogaster (fruit fly)
References: UniProt: P13368, receptor protein-tyrosine kinase
#2: Polysaccharide 2-acetamido-2-deoxy-beta-D-glucopyranose-(1-4)-[alpha-L-fucopyranose-(1-6)]2-acetamido-2-deoxy-beta- ...2-acetamido-2-deoxy-beta-D-glucopyranose-(1-4)-[alpha-L-fucopyranose-(1-6)]2-acetamido-2-deoxy-beta-D-glucopyranose


Type: oligosaccharide / Mass: 570.542 Da / Num. of mol.: 2
Source method: isolated from a genetically manipulated source
DescriptorTypeProgram
DGlcpNAcb1-4[LFucpa1-6]DGlcpNAcb1-ROHGlycam Condensed SequenceGMML 1.0
WURCS=2.0/2,3,2/[a2122h-1b_1-5_2*NCC/3=O][a1221m-1a_1-5]/1-1-2/a4-b1_a6-c1WURCSPDB2Glycan 1.1.0
[][D-1-deoxy-GlcpNAc]{[(4+1)][b-D-GlcpNAc]{}[(6+1)][a-L-Fucp]{}}LINUCSPDB-CARE
#3: Polysaccharide 2-acetamido-2-deoxy-beta-D-glucopyranose-(1-4)-2-acetamido-2-deoxy-beta-D-glucopyranose


Type: oligosaccharide / Mass: 424.401 Da / Num. of mol.: 2
Source method: isolated from a genetically manipulated source
DescriptorTypeProgram
DGlcpNAcb1-4DGlcpNAcb1-ROHGlycam Condensed SequenceGMML 1.0
WURCS=2.0/1,2,1/[a2122h-1b_1-5_2*NCC/3=O]/1-1/a4-b1WURCSPDB2Glycan 1.1.0
[][D-1-deoxy-GlcpNAc]{[(4+1)][b-D-GlcpNAc]{}}LINUCSPDB-CARE
#4: 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.: 16 / 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: Sevenless extracellular domain / Type: ORGANELLE OR CELLULAR COMPONENT / Entity ID: #1 / Source: RECOMBINANT
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: 3.06 Å / Resolution method: FSC 0.143 CUT-OFF / Num. of particles: 68141 / Symmetry type: POINT

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