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- PDB-5wen: GluA2 bound to GSG1L in digitonin, state 2 -

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Basic information

Entry
Database: PDB / ID: 5wen
TitleGluA2 bound to GSG1L in digitonin, state 2
ComponentsGlutamate receptor 2,Germ cell-specific gene 1-like protein
KeywordsTRANSPORT PROTEIN / Ion channel
Function / homology
Function and homology information


regulation of AMPA receptor activity / regulation of postsynaptic neurotransmitter receptor internalization / AMPA glutamate receptor activity / asymmetric synapse / extracellularly glutamate-gated ion channel activity / kainate selective glutamate receptor activity / AMPA glutamate receptor complex / ionotropic glutamate receptor complex / response to lithium ion / regulation of receptor recycling ...regulation of AMPA receptor activity / regulation of postsynaptic neurotransmitter receptor internalization / AMPA glutamate receptor activity / asymmetric synapse / extracellularly glutamate-gated ion channel activity / kainate selective glutamate receptor activity / AMPA glutamate receptor complex / ionotropic glutamate receptor complex / response to lithium ion / regulation of receptor recycling / immunoglobulin binding / somatodendritic compartment / positive regulation of synaptic transmission / integral component of postsynaptic membrane / ionotropic glutamate receptor activity / SNARE binding / response to fungicide / regulation of synaptic transmission, glutamatergic / integral component of postsynaptic density membrane / synaptic membrane / glutamate receptor activity / transmitter-gated ion channel activity involved in regulation of postsynaptic membrane potential / synaptic vesicle membrane / ionotropic glutamate receptor signaling pathway / synaptic transmission, glutamatergic / cytoskeletal protein binding / dendritic shaft / postsynaptic density membrane / dendrite cytoplasm / integral component of presynaptic membrane / PDZ domain binding / receptor internalization / Schaffer collateral - CA1 synapse / presynaptic membrane / protein tetramerization / terminal bouton / establishment of protein localization / modulation of chemical synaptic transmission / growth cone / postsynaptic membrane / chemical synaptic transmission / synaptic vesicle / amyloid-beta binding / ATPase binding / perikaryon / signaling receptor activity / dendritic spine / postsynaptic density / neuron projection / synapse / cell junction / glutamatergic synapse / dendrite / endoplasmic reticulum membrane / neuronal cell body / protein kinase binding / endoplasmic reticulum / integral component of plasma membrane / cell surface / protein-containing complex / membrane / identical protein binding / plasma membrane
Ligated ion channel L-glutamate- and glycine-binding site / GSG1-like protein / Ionotropic glutamate receptor / Ionotropic glutamate receptor, metazoa / GSG1-like / Ionotropic glutamate receptor, L-glutamate and glycine-binding domain / Periplasmic binding protein-like I / Receptor, ligand binding region / Ligand-gated ion channel / Receptor family ligand binding region
Glutamate receptor 2 / Germ cell-specific gene 1-like protein
Biological speciesRattus norvegicus (Norway rat)
Mus musculus (house mouse)
MethodELECTRON MICROSCOPY / single particle reconstruction / cryo EM / Resolution: 6.8 Å
AuthorsTwomey, E.C. / Yelshanskaya, M.V. / Grassucci, R.A. / Frank, J. / Sobolevsky, A.I.
Funding support United States, 5items
OrganizationGrant numberCountry
National Institutes of Health/National Institute of Neurological Disorders and Stroke (NIH/NINDS)NS093838 United States
National Institutes of Health/National Institute of Neurological Disorders and Stroke (NIH/NINDS)NS083660 United States
National Institutes of Health/National Cancer Institute (NIH/NCI)CA206573 United States
National Institutes of Health/National Institute of General Medical Sciences (NIH/NIGMS)GM029169 United States
Howard Hughes Medical Institute (HHMI) United States
CitationJournal: Nature / Year: 2017
Title: Channel opening and gating mechanism in AMPA-subtype glutamate receptors.
Authors: Edward C Twomey / Maria V Yelshanskaya / Robert A Grassucci / Joachim Frank / Alexander I Sobolevsky /
Abstract: AMPA (α-amino-3-hydroxy-5-methyl-4-isoxazole propionic acid)-subtype ionotropic glutamate receptors mediate fast excitatory neurotransmission throughout the central nervous system. Gated by the ...AMPA (α-amino-3-hydroxy-5-methyl-4-isoxazole propionic acid)-subtype ionotropic glutamate receptors mediate fast excitatory neurotransmission throughout the central nervous system. Gated by the neurotransmitter glutamate, AMPA receptors are critical for synaptic strength, and dysregulation of AMPA receptor-mediated signalling is linked to numerous neurological diseases. Here we use cryo-electron microscopy to solve the structures of AMPA receptor-auxiliary subunit complexes in the apo, antagonist- and agonist-bound states and determine the iris-like mechanism of ion channel opening. The ion channel selectivity filter is formed by the extended portions of the re-entrant M2 loops, while the helical portions of M2 contribute to extensive hydrophobic interfaces between AMPA receptor subunits in the ion channel. We show how the permeation pathway changes upon channel opening and identify conformational changes throughout the entire AMPA receptor that accompany activation and desensitization. Our findings provide a framework for understanding gating across the family of ionotropic glutamate receptors and the role of AMPA receptors in excitatory neurotransmission.
Validation Report
SummaryFull reportAbout validation report
History
DepositionJul 10, 2017Deposition site: RCSB / Processing site: RCSB
Revision 1.0Aug 2, 2017Provider: repository / Type: Initial release
Revision 1.1Sep 13, 2017Group: Author supporting evidence / Database references / Category: citation / pdbx_audit_support
Item: _citation.journal_id_ISSN / _citation.journal_volume ..._citation.journal_id_ISSN / _citation.journal_volume / _citation.page_first / _citation.page_last / _pdbx_audit_support.funding_organization
Revision 1.2Jul 18, 2018Group: Data collection / Experimental preparation / Category: em_sample_support / Item: _em_sample_support.grid_type
Revision 1.3Nov 20, 2019Group: Author supporting evidence / Category: pdbx_audit_support / Item: _pdbx_audit_support.funding_organization

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Assembly

Deposited unit
A: Glutamate receptor 2,Germ cell-specific gene 1-like protein
B: Glutamate receptor 2,Germ cell-specific gene 1-like protein
C: Glutamate receptor 2,Germ cell-specific gene 1-like protein
D: Glutamate receptor 2,Germ cell-specific gene 1-like protein
hetero molecules


Theoretical massNumber of molelcules
Total (without water)472,3436
Polymers469,8854
Non-polymers2,4592
Water0
1


TypeNameSymmetry operationNumber
identity operation1_5551
Buried area26160 Å2
ΔGint-184 kcal/mol
Surface area161020 Å2

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Components

#1: Protein
Glutamate receptor 2,Germ cell-specific gene 1-like protein / 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 / GSG1-like protein


Mass: 117471.211 Da / Num. of mol.: 4
Fragment: UNP residues 25-847 and UNP residues 2-238 linked via LINKER GTG,UNP residues 25-847 and UNP residues 2-238 linked via LINKER GTG
Mutation: N241E, V382L, G384E, N385D, N392Q, V1151L,N241E, V382L, G384E, N385D, N392Q, V1151L
Source method: isolated from a genetically manipulated source
Source: (gene. exp.) Rattus norvegicus (Norway rat), (gene. exp.) Mus musculus (house mouse)
Gene: Gria2, Glur2, Gsg1l / Cell line (production host): HEK293 gnti-, HEK gnti- / Production host: Homo sapiens (human) / References: UniProt: P19491, UniProt: D3Z7H4
#2: Chemical ChemComp-AJP / Digitonin / Digitonin


Mass: 1229.312 Da / Num. of mol.: 2 / Source method: obtained synthetically / Formula: C56H92O29 / Comment: detergent*YM

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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 bound to GSG1L in digitonin, state 2 / Type: COMPLEX / Entity ID: 1 / Source: RECOMBINANT
Molecular weightExperimental value: NO
Source (natural)Organism: Rattus norvegicus (Norway rat)
Source (recombinant)Organism: Homo sapiens (human)
Buffer solutionpH: 8
SpecimenConc.: 5 mg/ml / Details: GluA2 bound to GSG1L in digitonin, state 2 / Embedding applied: NO / Shadowing applied: NO / Staining applied: NO / Vitrification applied: YES
Specimen supportDetails: Gold-gold grids, hydrogen and oxygen glow discharge (20s, 10 watts, 6.4 sccm H2, 27.5 sccm O2)
Grid material: GOLD / Grid mesh size: 200 divisions/in. / Grid type: C-flat-1.2/1.3
VitrificationInstrument: FEI VITROBOT MARK IV / Cryogen name: ETHANE / Humidity: 100 % / Chamber temperature: 295 K

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Electron microscopy imaging

Experimental equipment
Model: Tecnai Polara / Image courtesy: FEI Company
MicroscopyModel: FEI POLARA 300
Electron gunElectron source: FIELD EMISSION GUN / Accelerating voltage: 300 kV / Illumination mode: SPOT SCAN
Electron lensMode: BRIGHT FIELDBright-field microscopy
Image recordingElectron dose: 67 e/Å2 / Detector mode: COUNTING / Film or detector model: GATAN K2 SUMMIT (4k x 4k)
Image scansMovie frames/image: 40

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Processing

SoftwareName: PHENIX / Version: 1.11.1_2575 / Classification: refinement
CTF correctionType: PHASE FLIPPING AND AMPLITUDE CORRECTION
SymmetryPoint symmetry: C2 (2 fold cyclic)
3D reconstructionResolution: 6.8 Å / Resolution method: FSC 0.143 CUT-OFF / Num. of particles: 18926 / Symmetry type: POINT
Atomic model buildingSpace: REAL
Refine LS restraints
Refinement-IDTypeDev idealNumber
ELECTRON MICROSCOPYf_bond_d0.00327952
ELECTRON MICROSCOPYf_angle_d0.80337796
ELECTRON MICROSCOPYf_dihedral_angle_d10.07616522
ELECTRON MICROSCOPYf_chiral_restr0.0474220
ELECTRON MICROSCOPYf_plane_restr0.0074752

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