+Open data
-Basic information
Entry | Database: PDB / ID: 8f6q | |||||||||
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Title | CryoEM structure of designed modular protein oligomer C8-71 | |||||||||
Components | C8-71 | |||||||||
Keywords | DE NOVO PROTEIN / Synthetic / Self-assembling / Oligomeric / Helical repeats | |||||||||
Biological species | synthetic construct (others) | |||||||||
Method | ELECTRON MICROSCOPY / single particle reconstruction / cryo EM / Resolution: 3.6 Å | |||||||||
Authors | Redler, R.L. / Edman, N.I. / Baker, D. / Ekiert, D. / Bhabha, G. | |||||||||
Funding support | United States, 2items
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Citation | Journal: bioRxiv / Year: 2023 Title: Modulation of FGF pathway signaling and vascular differentiation using designed oligomeric assemblies. Authors: Natasha I Edman / Rachel L Redler / Ashish Phal / Thomas Schlichthaerle / Sanjay R Srivatsan / Ali Etemadi / Seong J An / Andrew Favor / Devon Ehnes / Zhe Li / Florian Praetorius / Max ...Authors: Natasha I Edman / Rachel L Redler / Ashish Phal / Thomas Schlichthaerle / Sanjay R Srivatsan / Ali Etemadi / Seong J An / Andrew Favor / Devon Ehnes / Zhe Li / Florian Praetorius / Max Gordon / Wei Yang / Brian Coventry / Derrick R Hicks / Longxing Cao / Neville Bethel / Piper Heine / Analisa Murray / Stacey Gerben / Lauren Carter / Marcos Miranda / Babak Negahdari / Sangwon Lee / Cole Trapnell / Lance Stewart / Damian C Ekiert / Joseph Schlessinger / Jay Shendure / Gira Bhabha / Hannele Ruohola-Baker / David Baker / Abstract: Growth factors and cytokines signal by binding to the extracellular domains of their receptors and drive association and transphosphorylation of the receptor intracellular tyrosine kinase domains, ...Growth factors and cytokines signal by binding to the extracellular domains of their receptors and drive association and transphosphorylation of the receptor intracellular tyrosine kinase domains, initiating downstream signaling cascades. To enable systematic exploration of how receptor valency and geometry affects signaling outcomes, we designed cyclic homo-oligomers with up to 8 subunits using repeat protein building blocks that can be modularly extended. By incorporating a designed fibroblast growth-factor receptor (FGFR) binding module into these scaffolds, we generated a series of synthetic signaling ligands that exhibit potent valency- and geometry-dependent Ca2+ release and MAPK pathway activation. The high specificity of the designed agonists reveal distinct roles for two FGFR splice variants in driving endothelial and mesenchymal cell fates during early vascular development. The ability to incorporate receptor binding domains and repeat extensions in a modular fashion makes our designed scaffolds broadly useful for probing and manipulating cellular signaling pathways. #1: Journal: Cell(Cambridge,Mass.) / Year: 2024 Title: Modulation of FGF pathway signaling and vascular differentiation using designed oligomeric assemblies Authors: Edman, N.I. / Redler, R.L. / Phal, A. / Schlichthaerle, T. / Srivatsan, S.R. / Etemadi, A. / An, S.J. / Favor, A. / Ehnes, D. / Li, Z. / Praetorius, F. / Gordon, M. / Yang, W. / Coventry, B. ...Authors: Edman, N.I. / Redler, R.L. / Phal, A. / Schlichthaerle, T. / Srivatsan, S.R. / Etemadi, A. / An, S.J. / Favor, A. / Ehnes, D. / Li, Z. / Praetorius, F. / Gordon, M. / Yang, W. / Coventry, B. / Hicks, D.R. / Cao, L. / Bethel, N. / Heine, P. / Murray, A. / Gerben, S. / Carter, L. / Miranda, M. / Negahdari, B. / Lee, S. / Trapnell, C. / Stewart, L. / Ekiert, D.C. / Schlessinger, J. / Shendure, J. / Bhabha, G. / Ruohola-Baker, H. / Baker, D. | |||||||||
History |
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-Structure visualization
Structure viewer | Molecule: MolmilJmol/JSmol |
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-Downloads & links
-Download
PDBx/mmCIF format | 8f6q.cif.gz | 270.3 KB | Display | PDBx/mmCIF format |
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PDB format | pdb8f6q.ent.gz | 219.5 KB | Display | PDB format |
PDBx/mmJSON format | 8f6q.json.gz | Tree view | PDBx/mmJSON format | |
Others | Other downloads |
-Validation report
Arichive directory | https://data.pdbj.org/pub/pdb/validation_reports/f6/8f6q ftp://data.pdbj.org/pub/pdb/validation_reports/f6/8f6q | HTTPS FTP |
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-Related structure data
-Links
-Assembly
Deposited unit |
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1 |
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-Components
#1: Protein | Mass: 23435.004 Da / Num. of mol.: 8 Source method: isolated from a genetically manipulated source Source: (gene. exp.) synthetic construct (others) / Production host: Escherichia coli BL21 (bacteria) |
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-Experimental details
-Experiment
Experiment | Method: ELECTRON MICROSCOPY |
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EM experiment | Aggregation state: PARTICLE / 3D reconstruction method: single particle reconstruction |
-Sample preparation
Component | Name: Self-assembled homo-octamer of de novo designed protein C8-71 Type: COMPLEX / Entity ID: all / Source: RECOMBINANT | ||||||||||||||||||
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Source (natural) | Organism: synthetic construct (others) | ||||||||||||||||||
Source (recombinant) | Organism: Escherichia coli BL21 (bacteria) | ||||||||||||||||||
Buffer solution |
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Specimen | Experiment-ID: 1 / Conc.: 0.9 mg/ml / Embedding applied: NO / Shadowing applied: NO / Staining applied: NO / Vitrification applied: YES / Details: Sample present within the ice layer as both isolated homo-octameric rings and fibrils of variable length
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Specimen support |
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Vitrification | Chamber temperature: 295 K / Cryogen name: ETHANE / Details: blot time = 4s; blot force = 0 / Entry-ID: 8F6Q / Humidity: 100 % / Instrument: FEI VITROBOT MARK IV
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-Electron microscopy imaging
Experimental equipment | Model: Titan Krios / Image courtesy: FEI Company |
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Microscopy | Model: FEI TITAN KRIOS |
Electron gun | Electron source: FIELD EMISSION GUN / Accelerating voltage: 300 kV / Illumination mode: FLOOD BEAM |
Electron lens | Mode: BRIGHT FIELD / Nominal magnification: 81000 X / Nominal defocus max: 2500 nm / Nominal defocus min: 800 nm |
Specimen holder | Specimen holder model: FEI TITAN KRIOS AUTOGRID HOLDER |
Image recording | Average exposure time: 2.5 sec. / Electron dose: 61.3 e/Å2 / Film or detector model: GATAN K3 (6k x 4k) / Num. of grids imaged: 2 / Num. of real images: 3092 |
-Processing
Software |
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EM software |
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CTF correction | Type: PHASE FLIPPING AND AMPLITUDE CORRECTION | |||||||||||||||||||||||||||||||||||||||||||||
Symmetry | Point symmetry: C8 (8 fold cyclic) | |||||||||||||||||||||||||||||||||||||||||||||
3D reconstruction | Resolution: 3.6 Å / Resolution method: FSC 0.143 CUT-OFF / Num. of particles: 132391 / Symmetry type: POINT | |||||||||||||||||||||||||||||||||||||||||||||
Atomic model building | Protocol: AB INITIO MODEL / Space: REAL / Target criteria: Correlation coefficients Details: Designed oligomer was used as initial model and was initially fit into map using UCSF Chimera | |||||||||||||||||||||||||||||||||||||||||||||
Refinement | Cross valid method: NONE Stereochemistry target values: GeoStd + Monomer Library + CDL v1.2 | |||||||||||||||||||||||||||||||||||||||||||||
Displacement parameters | Biso mean: 102.29 Å2 | |||||||||||||||||||||||||||||||||||||||||||||
Refine LS restraints |
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