+Open data
-Basic information
Entry | Database: EMDB / ID: EMD-23245 | |||||||||
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Title | Integrin AlphaIIbBeta3-PT25-2 Complex | |||||||||
Map data | Cryo-EM map of Human aIIBb3 integrin headpiece with PT25-2 antibody fragment bound | |||||||||
Sample |
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Function / homology | Function and homology information tube development / regulation of serotonin uptake / positive regulation of adenylate cyclase-inhibiting opioid receptor signaling pathway / alpha9-beta1 integrin-ADAM8 complex / regulation of trophoblast cell migration / integrin alphaIIb-beta3 complex / regulation of postsynaptic neurotransmitter receptor diffusion trapping / alphav-beta3 integrin-vitronectin complex / regulation of extracellular matrix organization / positive regulation of glomerular mesangial cell proliferation ...tube development / regulation of serotonin uptake / positive regulation of adenylate cyclase-inhibiting opioid receptor signaling pathway / alpha9-beta1 integrin-ADAM8 complex / regulation of trophoblast cell migration / integrin alphaIIb-beta3 complex / regulation of postsynaptic neurotransmitter receptor diffusion trapping / alphav-beta3 integrin-vitronectin complex / regulation of extracellular matrix organization / positive regulation of glomerular mesangial cell proliferation / platelet alpha granule membrane / negative regulation of lipoprotein metabolic process / integrin alphav-beta3 complex / negative regulation of low-density lipoprotein receptor activity / fibrinogen binding / alphav-beta3 integrin-PKCalpha complex / maintenance of postsynaptic specialization structure / alphav-beta3 integrin-HMGB1 complex / blood coagulation, fibrin clot formation / mesodermal cell differentiation / vascular endothelial growth factor receptor 2 binding / negative regulation of lipid transport / glycinergic synapse / angiogenesis involved in wound healing / Elastic fibre formation / regulation of release of sequestered calcium ion into cytosol / alphav-beta3 integrin-IGF-1-IGF1R complex / cell-substrate junction assembly / platelet-derived growth factor receptor binding / filopodium membrane / extracellular matrix binding / positive regulation of vascular endothelial growth factor receptor signaling pathway / positive regulation of fibroblast migration / regulation of postsynaptic neurotransmitter receptor internalization / apolipoprotein A-I-mediated signaling pathway / regulation of bone resorption / apoptotic cell clearance / wound healing, spreading of epidermal cells / positive regulation of cell adhesion mediated by integrin / heterotypic cell-cell adhesion / integrin complex / positive regulation of leukocyte migration / Molecules associated with elastic fibres / microvillus membrane / smooth muscle cell migration / cell adhesion mediated by integrin / positive regulation of cell-matrix adhesion / negative chemotaxis / Syndecan interactions / cellular response to insulin-like growth factor stimulus / activation of protein kinase activity / p130Cas linkage to MAPK signaling for integrins / protein disulfide isomerase activity / cell-substrate adhesion / positive regulation of smooth muscle cell migration / positive regulation of osteoblast proliferation / cellular response to platelet-derived growth factor stimulus / TGF-beta receptor signaling activates SMADs / PECAM1 interactions / GRB2:SOS provides linkage to MAPK signaling for Integrins / lamellipodium membrane / platelet-derived growth factor receptor signaling pathway / fibronectin binding / negative regulation of macrophage derived foam cell differentiation / negative regulation of lipid storage / ECM proteoglycans / positive regulation of bone resorption / positive regulation of T cell migration / Integrin cell surface interactions / negative regulation of endothelial cell apoptotic process / coreceptor activity / positive regulation of substrate adhesion-dependent cell spreading / cell adhesion molecule binding / positive regulation of endothelial cell proliferation / embryo implantation / Integrin signaling / positive regulation of endothelial cell migration / cell-matrix adhesion / substrate adhesion-dependent cell spreading / platelet aggregation / response to activity / Signal transduction by L1 / integrin-mediated signaling pathway / regulation of actin cytoskeleton organization / protein kinase C binding / positive regulation of smooth muscle cell proliferation / wound healing / Signaling by high-kinase activity BRAF mutants / MAP2K and MAPK activation / RUNX1 regulates genes involved in megakaryocyte differentiation and platelet function / platelet activation / VEGFA-VEGFR2 Pathway / cell-cell adhesion / ruffle membrane / cellular response to mechanical stimulus / positive regulation of angiogenesis / positive regulation of peptidyl-tyrosine phosphorylation / Signaling by RAF1 mutants / Signaling by moderate kinase activity BRAF mutants / Paradoxical activation of RAF signaling by kinase inactive BRAF Similarity search - Function | |||||||||
Biological species | Mus musculus (house mouse) / Homo sapiens (human) | |||||||||
Method | single particle reconstruction / cryo EM / Resolution: 3.3 Å | |||||||||
Authors | Bush MW / Walz T / Coller B / Filizola M / Spasic A / Nesic D / Li J | |||||||||
Funding support | United States, 1 items
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Citation | Journal: Blood Adv / Year: 2021 Title: Electron microscopy shows that binding of monoclonal antibody PT25-2 primes integrin αIIbβ3 for ligand binding. Authors: Dragana Nešić / Martin Bush / Aleksandar Spasic / Jihong Li / Tetsuji Kamata / Makoto Handa / Marta Filizola / Thomas Walz / Barry S Coller / Abstract: The murine monoclonal antibody (mAb) PT25-2 induces αIIbβ3 to bind ligand and initiate platelet aggregation. The underlying mechanism is unclear, because previous mutagenesis studies suggested that ...The murine monoclonal antibody (mAb) PT25-2 induces αIIbβ3 to bind ligand and initiate platelet aggregation. The underlying mechanism is unclear, because previous mutagenesis studies suggested that PT25-2 binds to the αIIb β propeller, a site distant from the Arg-Gly-Asp-binding pocket. To elucidate the mechanism, we studied the αIIbβ3-PT25-2 Fab complex by negative-stain and cryo-electron microscopy (EM). We found that PT25-2 binding results in αIIbβ3 partially exposing multiple ligand-induced binding site epitopes and adopting extended conformations without swing-out of the β3 hybrid domain. The cryo-EM structure showed PT25-2 binding to the αIIb residues identified by mutagenesis but also to 2 additional regions. Overlay of the cryo-EM structure with the bent αIIbβ3 crystal structure showed that binding of PT25-2 creates clashes with the αIIb calf-1/calf-2 domains, suggesting that PT25-2 selectively binds to partially or fully extended receptor conformations and prevents a return to its bent conformation. Kinetic studies of the binding of PT25-2 compared with mAbs 10E5 and 7E3 support this hypothesis. We conclude that PT25-2 induces αIIbβ3 ligand binding by binding to extended conformations and by preventing the interactions between the αIIb and β3 leg domains and subsequently the βI and β3 leg domains required for the bent-closed conformation. | |||||||||
History |
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-Structure visualization
Movie |
Movie viewer |
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Structure viewer | EM map: SurfViewMolmilJmol/JSmol |
Supplemental images |
-Downloads & links
-EMDB archive
Map data | emd_23245.map.gz | 117.2 MB | EMDB map data format | |
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Header (meta data) | emd-23245-v30.xml emd-23245.xml | 17.8 KB 17.8 KB | Display Display | EMDB header |
Images | emd_23245.png | 173.2 KB | ||
Archive directory | http://ftp.pdbj.org/pub/emdb/structures/EMD-23245 ftp://ftp.pdbj.org/pub/emdb/structures/EMD-23245 | HTTPS FTP |
-Validation report
Summary document | emd_23245_validation.pdf.gz | 461.9 KB | Display | EMDB validaton report |
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Full document | emd_23245_full_validation.pdf.gz | 461.4 KB | Display | |
Data in XML | emd_23245_validation.xml.gz | 5.3 KB | Display | |
Data in CIF | emd_23245_validation.cif.gz | 6.3 KB | Display | |
Arichive directory | https://ftp.pdbj.org/pub/emdb/validation_reports/EMD-23245 ftp://ftp.pdbj.org/pub/emdb/validation_reports/EMD-23245 | HTTPS FTP |
-Related structure data
Related structure data | 7la4MC M: atomic model generated by this map C: citing same article (ref.) |
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Similar structure data |
-Links
EMDB pages | EMDB (EBI/PDBe) / EMDataResource |
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Related items in Molecule of the Month |
-Map
File | Download / File: emd_23245.map.gz / Format: CCP4 / Size: 125 MB / Type: IMAGE STORED AS FLOATING POINT NUMBER (4 BYTES) | ||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||
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Annotation | Cryo-EM map of Human aIIBb3 integrin headpiece with PT25-2 antibody fragment bound | ||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||
Projections & slices | Image control
Images are generated by Spider. | ||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||
Voxel size | X=Y=Z: 1 Å | ||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||
Density |
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Symmetry | Space group: 1 | ||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||
Details | EMDB XML:
CCP4 map header:
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-Supplemental data
-Sample components
+Entire : Human aIIBb3 integrin headpiece with PT25-2 antibody fragment bound
+Supramolecule #1: Human aIIBb3 integrin headpiece with PT25-2 antibody fragment bound
+Supramolecule #2: PT25-2 heavy and light chains
+Supramolecule #3: aIIBb3 integrin headpiece
+Macromolecule #1: PT25-2 heavy chain
+Macromolecule #2: PT25-2 light chain
+Macromolecule #3: Integrin alpha-IIb
+Macromolecule #4: Integrin beta-3
+Macromolecule #8: CALCIUM ION
+Macromolecule #9: 2-acetamido-2-deoxy-beta-D-glucopyranose
+Macromolecule #10: MAGNESIUM ION
+Macromolecule #11: water
-Experimental details
-Structure determination
Method | cryo EM |
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Processing | single particle reconstruction |
Aggregation state | particle |
-Sample preparation
Buffer | pH: 7.4 |
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Grid | Model: Quantifoil R1.2/1.3 / Material: COPPER / Support film - Material: CARBON / Support film - topology: HOLEY / Pretreatment - Type: GLOW DISCHARGE / Pretreatment - Atmosphere: AIR |
Vitrification | Cryogen name: ETHANE / Chamber humidity: 100 % / Chamber temperature: 277 K / Instrument: FEI VITROBOT MARK IV |
-Electron microscopy
Microscope | FEI TITAN KRIOS |
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Image recording | Film or detector model: GATAN K2 SUMMIT (4k x 4k) / Detector mode: SUPER-RESOLUTION / Average electron dose: 80.0 e/Å2 |
Electron beam | Acceleration voltage: 300 kV / Electron source: FIELD EMISSION GUN |
Electron optics | Illumination mode: FLOOD BEAM / Imaging mode: BRIGHT FIELD |
Sample stage | Specimen holder model: FEI TITAN KRIOS AUTOGRID HOLDER / Cooling holder cryogen: NITROGEN |
Experimental equipment | Model: Titan Krios / Image courtesy: FEI Company |
-Image processing
Final reconstruction | Resolution.type: BY AUTHOR / Resolution: 3.3 Å / Resolution method: FSC 0.143 CUT-OFF / Number images used: 166124 |
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Initial angle assignment | Type: MAXIMUM LIKELIHOOD |
Final angle assignment | Type: MAXIMUM LIKELIHOOD |