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- PDB-8bst: Crystal structure of the kainate receptor GluK3-H523A ligand bind... -
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Open data
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Basic information
Entry | Database: PDB / ID: 8bst | ||||||
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Title | Crystal structure of the kainate receptor GluK3-H523A ligand binding domain in complex with kainate at 2.7A resolution | ||||||
![]() | Glutamate receptor ionotropic, kainate 3 | ||||||
![]() | MEMBRANE PROTEIN / IONOTROPIC GLUTAMATE RECEPTOR / GLUK3 LIGAND-BINDING DOMAIN / GluK3-M523A-LBD | ||||||
Function / homology | ![]() Presynaptic function of Kainate receptors / cochlear hair cell ribbon synapse / adenylate cyclase inhibiting G protein-coupled glutamate receptor activity / kainate selective glutamate receptor complex / G protein-coupled glutamate receptor signaling pathway / Activation of Ca-permeable Kainate Receptor / negative regulation of synaptic transmission, glutamatergic / glutamate receptor activity / glutamate receptor signaling pathway / kainate selective glutamate receptor activity ...Presynaptic function of Kainate receptors / cochlear hair cell ribbon synapse / adenylate cyclase inhibiting G protein-coupled glutamate receptor activity / kainate selective glutamate receptor complex / G protein-coupled glutamate receptor signaling pathway / Activation of Ca-permeable Kainate Receptor / negative regulation of synaptic transmission, glutamatergic / glutamate receptor activity / glutamate receptor signaling pathway / kainate selective glutamate receptor activity / glutamate-gated receptor activity / ligand-gated monoatomic ion channel activity involved in regulation of presynaptic membrane potential / dendrite cytoplasm / regulation of membrane potential / transmitter-gated monoatomic ion channel activity involved in regulation of postsynaptic membrane potential / synaptic transmission, glutamatergic / postsynaptic density membrane / modulation of chemical synaptic transmission / terminal bouton / presynaptic membrane / chemical synaptic transmission / postsynaptic membrane / perikaryon / axon / glutamatergic synapse / dendrite / plasma membrane Similarity search - Function | ||||||
Biological species | ![]() ![]() | ||||||
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![]() | Venskutonyte, R. / Frydenvang, K. / Kastrup, J.S. | ||||||
Funding support | 1items
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![]() | ![]() Title: Small-molecule positive allosteric modulation of homomeric kainate receptors GluK1-3: development of screening assays and insight into GluK3 structure. Authors: Yasmin Bay / Raminta Venskutonytė / Stine M Frantsen / Thor S Thorsen / Maria Musgaard / Karla Frydenvang / Pierre Francotte / Bernard Pirotte / Philip C Biggin / Anders S Kristensen / ...Authors: Yasmin Bay / Raminta Venskutonytė / Stine M Frantsen / Thor S Thorsen / Maria Musgaard / Karla Frydenvang / Pierre Francotte / Bernard Pirotte / Philip C Biggin / Anders S Kristensen / Thomas Boesen / Darryl S Pickering / Michael Gajhede / Jette S Kastrup / ![]() ![]() ![]() Abstract: The kainate receptors GluK1-3 (glutamate receptor ionotropic, kainate receptors 1-3) belong to the family of ionotropic glutamate receptors and are essential for fast excitatory neurotransmission in ...The kainate receptors GluK1-3 (glutamate receptor ionotropic, kainate receptors 1-3) belong to the family of ionotropic glutamate receptors and are essential for fast excitatory neurotransmission in the brain, and are associated with neurological and psychiatric diseases. How these receptors can be modulated by small-molecule agents is not well understood, especially for GluK3. We show that the positive allosteric modulator BPAM344 can be used to establish robust calcium-sensitive fluorescence-based assays to test agonists, antagonists, and positive allosteric modulators of GluK1-3. The half-maximal effective concentration (EC) of BPAM344 for potentiating the response of 100 μm kainate was determined to be 26.3 μm for GluK1, 75.4 μm for GluK2, and 639 μm for GluK3. Domoate was found to be a potent agonist for GluK1 and GluK2, with an EC of 0.77 and 1.33 μm, respectively, upon co-application of 150 μm BPAM344. At GluK3, domoate acts as a very weak agonist or antagonist with a half-maximal inhibitory concentration (IC) of 14.5 μm, in presence of 500 μm BPAM344 and 100 μm kainate for competition binding. Using H523A-mutated GluK3, we determined the first dimeric structure of the ligand-binding domain by X-ray crystallography, allowing location of BPAM344, as well as zinc-, sodium-, and chloride-ion binding sites at the dimer interface. Molecular dynamics simulations support the stability of the ion sites as well as the involvement of Asp761, Asp790, and Glu797 in the binding of zinc ions. Using electron microscopy, we show that, in presence of glutamate and BPAM344, full-length GluK3 adopts a dimer-of-dimers arrangement. | ||||||
History |
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Structure visualization
Structure viewer | Molecule: ![]() ![]() |
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PDBx/mmCIF format | ![]() | 417.4 KB | Display | ![]() |
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PDB format | ![]() | 344.7 KB | Display | ![]() |
PDBx/mmJSON format | ![]() | Tree view | ![]() | |
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-Validation report
Arichive directory | ![]() ![]() | HTTPS FTP |
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-Related structure data
Related structure data | ![]() 8bsuC C: citing same article ( |
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Similar structure data | Similarity search - Function & homology ![]() |
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Links
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Assembly
Deposited unit | ![]()
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Unit cell |
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Components
-Protein , 1 types, 4 molecules ABCD
#1: Protein | Mass: 29025.383 Da / Num. of mol.: 4 / Mutation: M523A Source method: isolated from a genetically manipulated source Details: THE PROTEIN CRYSTALLIZED IS THE EXTRACELLULAR LIGAND BINDING DOMAIN OF GLUK3-H523A. TRANSMEMBRANE REGIONS WERE GENETICALLY REMOVED AND REPLACED WITH A GLY-THR LINKER (RESIDUES 547 AND 548 OF ...Details: THE PROTEIN CRYSTALLIZED IS THE EXTRACELLULAR LIGAND BINDING DOMAIN OF GLUK3-H523A. TRANSMEMBRANE REGIONS WERE GENETICALLY REMOVED AND REPLACED WITH A GLY-THR LINKER (RESIDUES 547 AND 548 OF THE STRUCTURE). THEREFORE, THE SEQUENCE MATCHES DISCONTINUOUSLY WITH THE REFERENCE DATABASE (432-546, 669-806),THE PROTEIN CRYSTALLIZED IS THE EXTRACELLULAR LIGAND BINDING DOMAIN OF GLUK3-H523A. TRANSMEMBRANE REGIONS WERE GENETICALLY REMOVED AND REPLACED WITH A GLY-THR LINKER (RESIDUES 547 AND 548 OF THE STRUCTURE). THEREFORE, THE SEQUENCE MATCHES DISCONTINUOUSLY WITH THE REFERENCE DATABASE (432-546, 669-806) Source: (gene. exp.) ![]() ![]() ![]() ![]() |
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-Non-polymers , 8 types, 143 molecules ![](data/chem/img/ACT.gif)
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![](data/chem/img/KAI.gif)
![](data/chem/img/GOL.gif)
![](data/chem/img/HOH.gif)
#2: Chemical | ChemComp-ACT / #3: Chemical | ChemComp-ZN / #4: Chemical | ChemComp-CL / #5: Chemical | ChemComp-SO4 / #6: Chemical | ChemComp-PEG / | #7: Chemical | ChemComp-KAI / #8: Chemical | ChemComp-GOL / #9: Water | ChemComp-HOH / | |
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-Details
Has ligand of interest | Y |
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-Experimental details
-Experiment
Experiment | Method: ![]() |
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Sample preparation
Crystal | Density Matthews: 2.35 Å3/Da / Density % sol: 47.63 % |
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Crystal grow | Temperature: 293 K / Method: vapor diffusion, hanging drop / pH: 8.5 Details: PEG4000, Lithium sulfate, zinc acetate, Tris buffer pH 8.5 |
-Data collection
Diffraction | Mean temperature: 100 K / Serial crystal experiment: N |
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Diffraction source | Source: ![]() ![]() ![]() |
Detector | Type: MARMOSAIC 225 mm CCD / Detector: CCD / Date: Oct 28, 2014 |
Radiation | Protocol: SINGLE WAVELENGTH / Monochromatic (M) / Laue (L): M / Scattering type: x-ray |
Radiation wavelength | Wavelength: 1 Å / Relative weight: 1 |
Reflection | Resolution: 2.7→42.46 Å / Num. obs: 29504 / % possible obs: 99.7 % / Redundancy: 4.3 % / CC1/2: 0.98 / Rmerge(I) obs: 0.13 / Rpim(I) all: 0.072 / Rrim(I) all: 0.149 / Χ2: 0.94 / Net I/σ(I): 8.6 / Num. measured all: 126006 |
Reflection shell | Resolution: 2.7→2.83 Å / % possible obs: 99.5 % / Redundancy: 4.3 % / Rmerge(I) obs: 0.804 / Num. measured all: 16674 / Num. unique obs: 3891 / CC1/2: 0.723 / Rpim(I) all: 0.439 / Rrim(I) all: 0.917 / Χ2: 0.98 / Net I/σ(I) obs: 1.9 |
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Processing
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Refinement | Method to determine structure: ![]()
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Solvent computation | Shrinkage radii: 0.9 Å / VDW probe radii: 1.11 Å / Solvent model: FLAT BULK SOLVENT MODEL | |||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||
Refinement step | Cycle: LAST / Resolution: 2.7→39.88 Å
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Refine LS restraints |
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LS refinement shell |
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Refinement TLS params. | Method: refined / Refine-ID: X-RAY DIFFRACTION
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Refinement TLS group |
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