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- PDB-9b64: GluA2 flip Q in complex with TARPgamma2 at pH5, class23, structur... -

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Basic information

Entry
Database: PDB / ID: 9b64
TitleGluA2 flip Q in complex with TARPgamma2 at pH5, class23, structure of LBD-TMD-TARPgamma2
Components
  • Isoform Flip of Glutamate receptor 2
  • Voltage-dependent calcium channel gamma-2 subunit
KeywordsTRANSPORT PROTEIN / AMPA receptor / ionotropic glutamate receptor / ion channel / auxiliary subunit
Function / homology
Function and homology information


Presynaptic depolarization and calcium channel opening / LGI-ADAM interactions / Trafficking of AMPA receptors / eye blink reflex / positive regulation of protein localization to basolateral plasma membrane / cerebellar mossy fiber / postsynaptic neurotransmitter receptor diffusion trapping / regulation of AMPA receptor activity / membrane hyperpolarization / nervous system process ...Presynaptic depolarization and calcium channel opening / LGI-ADAM interactions / Trafficking of AMPA receptors / eye blink reflex / positive regulation of protein localization to basolateral plasma membrane / cerebellar mossy fiber / postsynaptic neurotransmitter receptor diffusion trapping / regulation of AMPA receptor activity / membrane hyperpolarization / nervous system process / protein targeting to membrane / voltage-gated calcium channel complex / spine synapse / dendritic spine neck / dendritic spine cytoplasm / cellular response to amine stimulus / dendritic spine head / neurotransmitter receptor localization to postsynaptic specialization membrane / perisynaptic space / Activation of AMPA receptors / ligand-gated monoatomic cation channel activity / neuromuscular junction development / AMPA glutamate receptor activity / response to lithium ion / Trafficking of GluR2-containing AMPA receptors / transmission of nerve impulse / AMPA glutamate receptor clustering / kainate selective glutamate receptor activity / cellular response to glycine / AMPA glutamate receptor complex / immunoglobulin binding / asymmetric synapse / extracellularly glutamate-gated ion channel activity / regulation of receptor recycling / ionotropic glutamate receptor complex / membrane depolarization / conditioned place preference / glutamate receptor binding / Unblocking of NMDA receptors, glutamate binding and activation / positive regulation of synaptic transmission / regulation of synaptic transmission, glutamatergic / regulation of postsynaptic membrane neurotransmitter receptor levels / response to fungicide / voltage-gated calcium channel activity / glutamate-gated receptor activity / cytoskeletal protein binding / extracellular ligand-gated monoatomic ion channel activity / regulation of long-term synaptic depression / cellular response to brain-derived neurotrophic factor stimulus / glutamate-gated calcium ion channel activity / presynaptic active zone membrane / somatodendritic compartment / dendrite membrane / excitatory synapse / ionotropic glutamate receptor binding / ligand-gated monoatomic ion channel activity involved in regulation of presynaptic membrane potential / ionotropic glutamate receptor signaling pathway / dendrite cytoplasm / synaptic membrane / positive regulation of excitatory postsynaptic potential / hippocampal mossy fiber to CA3 synapse / dendritic shaft / SNARE binding / PDZ domain binding / synaptic transmission, glutamatergic / regulation of membrane potential / protein tetramerization / transmitter-gated monoatomic ion channel activity involved in regulation of postsynaptic membrane potential / establishment of protein localization / response to calcium ion / cerebral cortex development / postsynaptic density membrane / modulation of chemical synaptic transmission / receptor internalization / Schaffer collateral - CA1 synapse / terminal bouton / synaptic vesicle / long-term synaptic potentiation / synaptic vesicle membrane / signaling receptor activity / amyloid-beta binding / presynapse / growth cone / presynaptic membrane / scaffold protein binding / dendritic spine / chemical synaptic transmission / perikaryon / postsynaptic membrane / neuron projection / postsynaptic density / axon / external side of plasma membrane / neuronal cell body / synapse / dendrite / protein kinase binding / protein-containing complex binding / glutamatergic synapse / cell surface
Similarity search - Function
Voltage-dependent calcium channel, gamma-2 subunit / : / PMP-22/EMP/MP20/Claudin family / Voltage-dependent calcium channel, gamma subunit / PMP-22/EMP/MP20/Claudin superfamily / Ionotropic glutamate receptor, metazoa / Ligated ion channel L-glutamate- and glycine-binding site / Ionotropic glutamate receptor, L-glutamate and glycine-binding domain / Ligated ion channel L-glutamate- and glycine-binding site / Ligand-gated ion channel ...Voltage-dependent calcium channel, gamma-2 subunit / : / PMP-22/EMP/MP20/Claudin family / Voltage-dependent calcium channel, gamma subunit / PMP-22/EMP/MP20/Claudin superfamily / Ionotropic glutamate receptor, metazoa / Ligated ion channel L-glutamate- and glycine-binding site / Ionotropic glutamate receptor, L-glutamate and glycine-binding domain / Ligated ion channel L-glutamate- and glycine-binding site / Ligand-gated ion channel / : / 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
Voltage-dependent calcium channel gamma-2 subunit / Glutamate receptor 2
Similarity search - Component
Biological speciesMus musculus (house mouse)
Rattus norvegicus (Norway rat)
MethodELECTRON MICROSCOPY / single particle reconstruction / cryo EM / Resolution: 3.56 Å
AuthorsNakagawa, T. / Greger, I.H.
Funding support United States, United Kingdom, 4items
OrganizationGrant numberCountry
National Institutes of Health/National Institute of Mental Health (NIH/NIMH)R01MH123474 United States
UK Research and Innovation (UKRI)MC_U105174197 United Kingdom
Biotechnology and Biological Sciences Research Council (BBSRC)BB/N002113/1 United Kingdom
Wellcome Trust223194/Z/21/Z United Kingdom
CitationJournal: Nat Struct Mol Biol / Year: 2024
Title: Proton-triggered rearrangement of the AMPA receptor N-terminal domains impacts receptor kinetics and synaptic localization.
Authors: Josip Ivica / Nejc Kejzar / Hinze Ho / Imogen Stockwell / Viktor Kuchtiak / Alexander M Scrutton / Terunaga Nakagawa / Ingo H Greger /
Abstract: AMPA glutamate receptors (AMPARs) are ion channel tetramers that mediate the majority of fast excitatory synaptic transmission. They are composed of four subunits (GluA1-GluA4); the GluA2 subunit ...AMPA glutamate receptors (AMPARs) are ion channel tetramers that mediate the majority of fast excitatory synaptic transmission. They are composed of four subunits (GluA1-GluA4); the GluA2 subunit dominates AMPAR function throughout the forebrain. Its extracellular N-terminal domain (NTD) determines receptor localization at the synapse, ensuring reliable synaptic transmission and plasticity. This synaptic anchoring function requires a compact NTD tier, stabilized by a GluA2-specific NTD interface. Here we show that low pH conditions, which accompany synaptic activity, rupture this interface. All-atom molecular dynamics simulations reveal that protonation of an interfacial histidine residue (H208) centrally contributes to NTD rearrangement. Moreover, in stark contrast to their canonical compact arrangement at neutral pH, GluA2 cryo-electron microscopy structures exhibit a wide spectrum of NTD conformations under acidic conditions. We show that the consequences of this pH-dependent conformational control are twofold: rupture of the NTD tier slows recovery from desensitized states and increases receptor mobility at mouse hippocampal synapses. Therefore, a proton-triggered NTD switch will shape both AMPAR location and kinetics, thereby impacting synaptic signal transmission.
History
DepositionMar 23, 2024Deposition site: RCSB / Processing site: RCSB
Revision 1.0Jul 31, 2024Provider: repository / Type: Initial release
Revision 1.1Aug 21, 2024Group: Data collection / Database references / Category: citation / em_admin
Item: _citation.country / _citation.journal_abbrev ..._citation.country / _citation.journal_abbrev / _citation.journal_id_CSD / _citation.journal_id_ISSN / _citation.pdbx_database_id_DOI / _citation.year / _em_admin.last_update
Revision 1.2Sep 4, 2024Group: Data collection / Database references / Category: citation / em_admin
Item: _citation.pdbx_database_id_PubMed / _citation.title / _em_admin.last_update
Revision 1.3Oct 30, 2024Group: Data collection / Database references / Structure summary
Category: citation / em_admin ...citation / em_admin / pdbx_entry_details / pdbx_modification_feature
Item: _citation.journal_volume / _citation.page_first ..._citation.journal_volume / _citation.page_first / _citation.page_last / _em_admin.last_update
Revision 1.4Nov 6, 2024Group: Data collection / Category: em_admin / Item: _em_admin.last_update

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Structure visualization

Structure viewerMolecule:
MolmilJmol/JSmol

Downloads & links

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Assembly

Deposited unit
H: Voltage-dependent calcium channel gamma-2 subunit
F: Voltage-dependent calcium channel gamma-2 subunit
C: Isoform Flip of Glutamate receptor 2
B: Isoform Flip of Glutamate receptor 2
A: Isoform Flip of Glutamate receptor 2
D: Isoform Flip of Glutamate receptor 2
E: Voltage-dependent calcium channel gamma-2 subunit
G: Voltage-dependent calcium channel gamma-2 subunit


Theoretical massNumber of molelcules
Total (without water)542,2258
Polymers542,2258
Non-polymers00
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
Voltage-dependent calcium channel gamma-2 subunit / Neuronal voltage-gated calcium channel gamma-2 subunit / Stargazin / Transmembrane AMPAR regulatory ...Neuronal voltage-gated calcium channel gamma-2 subunit / Stargazin / Transmembrane AMPAR regulatory protein gamma-2 / TARP gamma-2


Mass: 35938.746 Da / Num. of mol.: 4
Source method: isolated from a genetically manipulated source
Source: (gene. exp.) Mus musculus (house mouse) / Gene: Cacng2, Stg / Cell line (production host): HEK / Production host: Homo sapiens (human) / References: UniProt: O88602
#2: Protein
Isoform Flip of Glutamate receptor 2 / 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


Mass: 99617.492 Da / Num. of mol.: 4
Source method: isolated from a genetically manipulated source
Details: A FLAG epitope tag is inserted near the C-terminus / Source: (gene. exp.) Rattus norvegicus (Norway rat) / Gene: Gria2, Glur2 / Cell line (production host): HEK / Production host: Homo sapiens (human) / References: UniProt: P19491
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: GluA2 (flip-Q isoform) in complex with TARPgamma2 at 4:4 stoichiometry
Type: COMPLEX / Entity ID: all / Source: MULTIPLE SOURCES
Molecular weightValue: 0.5 MDa / Experimental value: NO
Buffer solutionpH: 8
Details: Tris adjusted to pH 8 using HCl. 4 micro litter of protein in buffer was mixed with 1 micro litter of 50mM citric acid buffer (the 50mM citric acid buffer was prepared by diluting 0.5M ...Details: Tris adjusted to pH 8 using HCl. 4 micro litter of protein in buffer was mixed with 1 micro litter of 50mM citric acid buffer (the 50mM citric acid buffer was prepared by diluting 0.5M citric acid/sodium citrate buffer at pH4.0) immediately before applying the sample to the grid.
Buffer component
IDConc.NameFormulaBuffer-ID
1150 mMsodium chlorideNaCl1
220 mMTris1
30.03 %GDN1
SpecimenConc.: 10 mg/ml / Embedding applied: NO / Shadowing applied: NO / Staining applied: NO / Vitrification applied: YES
Specimen supportGrid material: GOLD / Grid mesh size: 300 divisions/in. / Grid type: UltrAuFoil R1.2/1.3
VitrificationInstrument: FEI VITROBOT MARK IV / Cryogen name: ETHANE / Humidity: 100 % / Chamber temperature: 277.15 K

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Electron microscopy imaging

Experimental equipment
Model: Titan Krios / Image courtesy: FEI Company
MicroscopyModel: TFS KRIOS
Electron gunElectron source: FIELD EMISSION GUN / Accelerating voltage: 300 kV / Illumination mode: FLOOD BEAM
Electron lensMode: BRIGHT FIELD / Nominal defocus max: 2400 nm / Nominal defocus min: 1000 nm / Alignment procedure: COMA FREE
Specimen holderCryogen: NITROGEN
Image recordingElectron dose: 55.6 e/Å2 / Film or detector model: GATAN K3 BIOQUANTUM (6k x 4k) / Num. of grids imaged: 1 / Num. of real images: 19684

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Processing

EM software
IDNameVersionCategory
1RELION4particle selection
2EPUimage acquisition
4CTFFIND4CTF correction
7Cootmodel fitting
9PHENIX1.20.1_4487:model refinement
10RELION4initial Euler assignment
11RELION4final Euler assignment
12RELION4classification
13RELION43D reconstruction
CTF correctionType: PHASE FLIPPING AND AMPLITUDE CORRECTION
Particle selectionDetails: Relion Autopick
SymmetryPoint symmetry: C1 (asymmetric)
3D reconstructionResolution: 3.56 Å / Resolution method: FSC 0.143 CUT-OFF / Num. of particles: 29945 / Algorithm: FOURIER SPACE / Num. of class averages: 1 / Symmetry type: POINT
Atomic model buildingSpace: REAL
Atomic model building
IDPDB-ID 3D fitting-IDAccession codeInitial refinement model-IDSource nameType
18FQF

8fqf

18FQF1PDBexperimental model
28FQG

8fqg

18FQG2PDBexperimental model
Refine LS restraints
Refine-IDTypeDev idealNumber
ELECTRON MICROSCOPYf_bond_d0.00318913
ELECTRON MICROSCOPYf_angle_d0.60825573
ELECTRON MICROSCOPYf_dihedral_angle_d3.6282567
ELECTRON MICROSCOPYf_chiral_restr0.0392877
ELECTRON MICROSCOPYf_plane_restr0.0053159

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