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- PDB-8ss3: Structure of LBD-TMD of AMPA receptor GluA2 in complex with auxil... -

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Basic information

Entry
Database: PDB / ID: 8ss3
TitleStructure of LBD-TMD of AMPA receptor GluA2 in complex with auxiliary subunits TARP gamma-5 and cornichon-2 bound to competitive antagonist ZK and channel blocker spermidine (closed state)
Components
  • Glutamate receptor 2, Voltage-dependent calcium channel gamma-5 subunit chimera
  • Protein cornichon homolog 2
KeywordsSIGNALING PROTEIN / AMPA receptor / spermidine / TARP gamma-5 / cornichon-2
Function / homology
Function and homology information


neurotransmitter receptor transport, postsynaptic endosome to lysosome / regulation of AMPA receptor activity / neurotransmitter receptor internalization / channel regulator activity / Cargo concentration in the ER / postsynaptic neurotransmitter receptor diffusion trapping / COPII-mediated vesicle transport / spine synapse / dendritic spine neck / dendritic spine head ...neurotransmitter receptor transport, postsynaptic endosome to lysosome / regulation of AMPA receptor activity / neurotransmitter receptor internalization / channel regulator activity / Cargo concentration in the ER / postsynaptic neurotransmitter receptor diffusion trapping / COPII-mediated vesicle transport / spine synapse / dendritic spine neck / dendritic spine head / Activation of AMPA receptors / response to lithium ion / perisynaptic space / cellular response to glycine / transmission of nerve impulse / AMPA glutamate receptor activity / Trafficking of GluR2-containing AMPA receptors / immunoglobulin binding / AMPA glutamate receptor complex / kainate selective glutamate receptor activity / ionotropic glutamate receptor complex / extracellularly glutamate-gated ion channel activity / asymmetric synapse / regulation of receptor recycling / Unblocking of NMDA receptors, glutamate binding and activation / voltage-gated calcium channel activity / glutamate receptor binding / positive regulation of synaptic transmission / presynaptic active zone membrane / glutamate-gated receptor activity / response to fungicide / regulation of synaptic transmission, glutamatergic / cellular response to brain-derived neurotrophic factor stimulus / vesicle-mediated transport / somatodendritic compartment / dendrite membrane / ligand-gated monoatomic ion channel activity involved in regulation of presynaptic membrane potential / ionotropic glutamate receptor binding / ionotropic glutamate receptor signaling pathway / endoplasmic reticulum-Golgi intermediate compartment membrane / cytoskeletal protein binding / dendrite cytoplasm / positive regulation of synaptic transmission, glutamatergic / SNARE binding / dendritic shaft / transmitter-gated monoatomic ion channel activity involved in regulation of postsynaptic membrane potential / synaptic membrane / synaptic transmission, glutamatergic / PDZ domain binding / protein tetramerization / postsynaptic density membrane / modulation of chemical synaptic transmission / Schaffer collateral - CA1 synapse / establishment of protein localization / ER to Golgi transport vesicle membrane / terminal bouton / receptor internalization / synaptic vesicle membrane / cerebral cortex development / synaptic vesicle / presynapse / signaling receptor activity / presynaptic membrane / amyloid-beta binding / growth cone / perikaryon / chemical synaptic transmission / scaffold protein binding / postsynaptic membrane / dendritic spine / postsynaptic density / neuron projection / axon / neuronal cell body / dendrite / synapse / glutamatergic synapse / protein-containing complex binding / endoplasmic reticulum membrane / protein kinase binding / cell surface / endoplasmic reticulum / protein-containing complex / membrane / identical protein binding / plasma membrane
Similarity search - Function
Voltage-dependent calcium channel, gamma-5 subunit / Cornichon / Cornichon, conserved site / Cornichon protein / Cornichon family signature. / Cornichon / PMP-22/EMP/MP20/Claudin tight junction / Voltage-dependent calcium channel, gamma subunit / PMP-22/EMP/MP20/Claudin superfamily / Bacterial extracellular solute-binding proteins, family 3 ...Voltage-dependent calcium channel, gamma-5 subunit / Cornichon / Cornichon, conserved site / Cornichon protein / Cornichon family signature. / Cornichon / PMP-22/EMP/MP20/Claudin tight junction / Voltage-dependent calcium channel, gamma subunit / PMP-22/EMP/MP20/Claudin superfamily / Bacterial extracellular solute-binding proteins, family 3 / Solute-binding protein family 3/N-terminal domain of MltF / Ionotropic glutamate receptor, metazoa / Ligated ion channel L-glutamate- and glycine-binding site / : / Ligand-gated ion channel / Ionotropic glutamate receptor, L-glutamate and glycine-binding domain / Ligated ion channel L-glutamate- and glycine-binding site / Ionotropic glutamate receptor / Eukaryotic homologues of bacterial periplasmic substrate binding proteins. / Receptor, ligand binding region / Receptor family ligand binding region / Periplasmic binding protein-like I
Similarity search - Domain/homology
CHOLESTEROL / 1,2-DIOLEOYL-SN-GLYCERO-3-PHOSPHOCHOLINE / SPERMIDINE / Chem-ZK1 / Glutamate receptor 2 / Protein cornichon homolog 2 / Voltage-dependent calcium channel gamma-5 subunit
Similarity search - Component
Biological speciesRattus norvegicus (Norway rat)
Homo sapiens (human)
MethodELECTRON MICROSCOPY / single particle reconstruction / cryo EM / Resolution: 3.21 Å
AuthorsGangwar, S.P. / Yen, L.Y. / Yelshanskaya, M.V. / Sobolevsky, A.I.
Funding support United States, 4items
OrganizationGrant numberCountry
National Institutes of Health/National Institute of Neurological Disorders and Stroke (NIH/NINDS)NS083660 United States
National Institutes of Health/National Institute of Neurological Disorders and Stroke (NIH/NINDS)NS107253 United States
National Institutes of Health/National Institute of Arthritis and Musculoskeletal and Skin Diseases (NIH/NIAMS)AR078814 United States
National Institutes of Health/National Cancer Institute (NIH/NCI)CA206573 United States
CitationJournal: Nat Struct Mol Biol / Year: 2023
Title: Modulation of GluA2-γ5 synaptic complex desensitization, polyamine block and antiepileptic perampanel inhibition by auxiliary subunit cornichon-2.
Authors: Shanti Pal Gangwar / Laura Y Yen / Maria V Yelshanskaya / Aryeh Korman / Drew R Jones / Alexander I Sobolevsky /
Abstract: Synaptic complexes of α-amino-3-hydroxy-5-methyl-4-isoxazolepropionic acid (AMPA) receptors (AMPARs) with auxiliary subunits mediate most excitatory neurotransmission and can be targeted to treat ...Synaptic complexes of α-amino-3-hydroxy-5-methyl-4-isoxazolepropionic acid (AMPA) receptors (AMPARs) with auxiliary subunits mediate most excitatory neurotransmission and can be targeted to treat neuropsychiatric and neurological disorders, including epilepsy. Here we present cryogenic-electron microscopy structures of rat GluA2 AMPAR complexes with inhibitory mouse γ5 and potentiating human cornichon-2 (CNIH2) auxiliary subunits. CNIH2 appears to destabilize the desensitized state of the complex by reducing the separation of the upper lobes in ligand-binding domain dimers. At the same time, CNIH2 stabilizes binding of polyamine spermidine to the selectivity filter of the closed ion channel. Nevertheless, CNIH2, and to a lesser extent γ5, attenuate polyamine block of the open channel and reduce the potency of the antiepileptic drug perampanel that inhibits the synaptic complex allosterically by binding to sites in the ion channel extracellular collar. These findings illustrate the fine-tuning of synaptic complex structure and function in an auxiliary subunit-dependent manner, which is critical for the study of brain region-specific neurotransmission and design of therapeutics for disease treatment.
History
DepositionMay 8, 2023Deposition site: RCSB / Processing site: RCSB
Revision 1.0Sep 6, 2023Provider: repository / Type: Initial release
Revision 1.1Sep 13, 2023Group: Database references / Category: citation / citation_author / Item: _citation.pdbx_database_id_PubMed / _citation.title
Revision 1.2Nov 1, 2023Group: Database references / Category: citation
Item: _citation.journal_volume / _citation.page_first / _citation.page_last

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Structure visualization

Structure viewerMolecule:
MolmilJmol/JSmol

Downloads & links

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Assembly

Deposited unit
A: Glutamate receptor 2, Voltage-dependent calcium channel gamma-5 subunit chimera
B: Glutamate receptor 2, Voltage-dependent calcium channel gamma-5 subunit chimera
C: Glutamate receptor 2, Voltage-dependent calcium channel gamma-5 subunit chimera
D: Glutamate receptor 2, Voltage-dependent calcium channel gamma-5 subunit chimera
E: Protein cornichon homolog 2
F: Protein cornichon homolog 2
hetero molecules


Theoretical massNumber of molelcules
Total (without water)547,87371
Polymers497,8126
Non-polymers50,06165
Water0
1


  • Idetical with deposited unit
  • defined by author
  • Evidence: electron microscopy
TypeNameSymmetry operationNumber
identity operation1_5551

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Components

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Protein , 2 types, 6 molecules ABCDEF

#1: Protein
Glutamate receptor 2, Voltage-dependent calcium channel gamma-5 subunit chimera / / GluR-2 / AMPA-selective glutamate receptor 2 / GluR-B / GluR-K2 / Glutamate receptor ionotropic / ...GluR-2 / AMPA-selective glutamate receptor 2 / GluR-B / GluR-K2 / Glutamate receptor ionotropic / AMPA 2 / GluA2 / Neuronal voltage-gated calcium channel gamma-5 subunit / Transmembrane AMPAR regulatory protein gamma-5 / TARP gamma-5


Mass: 114978.859 Da / Num. of mol.: 4
Source method: isolated from a genetically manipulated source
Source: (gene. exp.) Rattus norvegicus (Norway rat) / Gene: Gria2, Glur2, Cacng5 / Plasmid: pEG BacMam / Cell line (production host): HEK293S-GnTi / Production host: Homo sapiens (human) / References: UniProt: P19491, UniProt: Q8VHW8
#2: Protein Protein cornichon homolog 2 / CNIH-2 / Cornichon family AMPA receptor auxiliary protein 2 / Cornichon-like protein


Mass: 18948.420 Da / Num. of mol.: 2
Source method: isolated from a genetically manipulated source
Source: (gene. exp.) Homo sapiens (human) / Cell line: HEK293S-GnTi / Gene: CNIH2, CNIL / Cell line (production host): HEK293S-GnTi / Production host: Homo sapiens (human) / References: UniProt: Q6PI25

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Non-polymers , 5 types, 65 molecules

#3: Chemical
ChemComp-ZK1 / {[7-morpholin-4-yl-2,3-dioxo-6-(trifluoromethyl)-3,4-dihydroquinoxalin-1(2H)-yl]methyl}phosphonic acid / [[3,4-Dihydro-7-(4-morpholinyl)-2,3-dioxo-6-(trifluorom ethyl)-1(2H)-quinoxalinyl]methyl]phosphonic acid / Fanapanel


Mass: 409.254 Da / Num. of mol.: 4 / Source method: obtained synthetically / Formula: C14H15F3N3O6P / Feature type: SUBJECT OF INVESTIGATION / Comment: antagonist, medication*YM
#4: Chemical...
ChemComp-PCW / 1,2-DIOLEOYL-SN-GLYCERO-3-PHOSPHOCHOLINE / (Z,Z)-4-HYDROXY-N,N,N-TRIMETHYL-10-OXO-7-[(1-OXO-9-OCTADECENYL)OXY]-3,5,9-TRIOXA-4-PHOSPHAHEPTACOS-18-EN-1-AMINIUM-4-OXIDE


Mass: 787.121 Da / Num. of mol.: 50 / Source method: obtained synthetically / Formula: C44H85NO8P / Comment: DOPC, phospholipid*YM
#5: Chemical
ChemComp-CLR / CHOLESTEROL / Cholesterol


Mass: 386.654 Da / Num. of mol.: 4 / Source method: obtained synthetically / Formula: C27H46O
#6: Chemical
ChemComp-AJP / Digitonin / Digitonin


Mass: 1229.312 Da / Num. of mol.: 6 / Source method: obtained synthetically / Formula: C56H92O29 / Comment: detergent*YM
#7: Chemical ChemComp-SPD / SPERMIDINE / N-(2-AMINO-PROPYL)-1,4-DIAMINOBUTANE / PA(34) / Spermidine


Mass: 145.246 Da / Num. of mol.: 1 / Source method: obtained synthetically / Formula: C7H19N3 / Feature type: SUBJECT OF INVESTIGATION

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Details

Has ligand of interestY

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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: GluA2-TARP gamma5-cornichon-2 / Type: COMPLEX
Details: Structure of LBD-TMD of AMPA receptor GluA2 in complex with auxiliary subunits TARP gamma-5 and cornichon-2 bound to competitive antagonist ZK and channel blocker spermidine (closed state)
Entity ID: #1-#2 / Source: MULTIPLE SOURCES
Source (natural)
IDEntity assembly-IDOrganismNcbi tax-ID
21Rattus norvegicus (Norway rat)10116
31Homo sapiens (human)9606
Source (recombinant)
IDEntity assembly-IDOrganismNcbi tax-IDCellPlasmid
21Homo sapiens (human)9606HEK293S-GnTipEG BacMam
31Homo sapiens (human)9606HEK293S-GnTipEG BacMam
Buffer solutionpH: 8
Buffer component
IDConc.NameFormulaBuffer-ID
120 mMTrisC4H11NO31
2150 mMSodium chlorideNaClSodium chloride1
30.05 %DigitoninC56H92O291
40.005 %Cholesteryl hemisuccinateC31H50O41
SpecimenConc.: 4.5 mg/ml / Embedding applied: NO / Shadowing applied: NO / Staining applied: NO / Vitrification applied: YES
Details: Protein extracted and purified in detergent micelle
Specimen supportGrid material: GOLD / Grid mesh size: 300 divisions/in. / Grid type: UltrAuFoil
VitrificationInstrument: FEI VITROBOT MARK IV / Cryogen name: ETHANE / Humidity: 100 % / Chamber temperature: 277 K / Details: vitrification carried out in nitrogen atmosphere

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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: FLOOD BEAM
Electron lensMode: BRIGHT FIELDBright-field microscopy / Nominal defocus max: 2000 nm / Nominal defocus min: 1000 nm / Cs: 2.7 mm / C2 aperture diameter: 100 µm
Specimen holderCryogen: NITROGEN / Specimen holder model: FEI TITAN KRIOS AUTOGRID HOLDER
Image recordingElectron dose: 45 e/Å2 / Film or detector model: GATAN K3 (6k x 4k)

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Processing

EM software
IDNameVersionCategory
7Cootmodel fitting
9PHENIXmodel refinement
13cryoSPARC4.0.33D reconstruction
CTF correctionType: PHASE FLIPPING AND AMPLITUDE CORRECTION
SymmetryPoint symmetry: C2 (2 fold cyclic)
3D reconstructionResolution: 3.21 Å / Resolution method: FSC 0.143 CUT-OFF / Num. of particles: 81723 / Symmetry type: POINT
Refine LS restraints
Refine-IDTypeDev idealNumber
ELECTRON MICROSCOPYf_bond_d0.01120315
ELECTRON MICROSCOPYf_angle_d1.41827278
ELECTRON MICROSCOPYf_dihedral_angle_d14.82811733
ELECTRON MICROSCOPYf_chiral_restr0.0782954
ELECTRON MICROSCOPYf_plane_restr0.0073166

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