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- EMDB-8823: Activated GluA2 complex bound to glutamate, cyclothiazide, and ST... -

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Basic information

Entry
Database: EMDB / ID: EMD-8823
TitleActivated GluA2 complex bound to glutamate, cyclothiazide, and STZ in digitonin
Map data
SampleActivated GluA2 complex bound to glutamate, cyclothiazide, and STZ in digitonin
  • Glutamate receptor 2,Voltage-dependent calcium channel gamma-2 subunit chimera
  • (ligand) x 2
Function / homology
Function and homology information


eye blink reflex / positive regulation of protein localization to basolateral plasma membrane / neurotransmitter receptor transport, postsynaptic endosome to lysosome / neurotransmitter receptor internalization / regulation of postsynaptic membrane neurotransmitter receptor levels / cerebellar mossy fiber / neurotransmitter receptor localization to postsynaptic specialization membrane / regulation of postsynaptic neurotransmitter receptor activity / positive regulation of AMPA receptor activity / regulation of AMPA receptor activity ...eye blink reflex / positive regulation of protein localization to basolateral plasma membrane / neurotransmitter receptor transport, postsynaptic endosome to lysosome / neurotransmitter receptor internalization / regulation of postsynaptic membrane neurotransmitter receptor levels / cerebellar mossy fiber / neurotransmitter receptor localization to postsynaptic specialization membrane / regulation of postsynaptic neurotransmitter receptor activity / positive regulation of AMPA receptor activity / regulation of AMPA receptor activity / postsynaptic neurotransmitter receptor diffusion trapping / membrane hyperpolarization / protein targeting to membrane / voltage-gated calcium channel complex / somatodendritic compartment / neuromuscular junction development / membrane depolarization / transmission of nerve impulse / nervous system process / voltage-gated calcium channel activity / AMPA glutamate receptor activity / asymmetric synapse / ionotropic glutamate receptor binding / kainate selective glutamate receptor activity / AMPA glutamate receptor complex / ionotropic glutamate receptor complex / response to lithium ion / regulation of receptor recycling / immunoglobulin binding / positive regulation of synaptic transmission / integral component of postsynaptic membrane / regulation of membrane potential / SNARE binding / ionotropic glutamate receptor activity / response to fungicide / regulation of synaptic transmission, glutamatergic / integral component of postsynaptic density membrane / glutamate receptor activity / transmitter-gated ion channel activity involved in regulation of postsynaptic membrane potential / ionotropic glutamate receptor signaling pathway / synaptic transmission, glutamatergic / cytoskeletal protein binding / synaptic vesicle membrane / postsynaptic density membrane / dendritic shaft / hippocampal mossy fiber to CA3 synapse / receptor internalization / dendrite cytoplasm / integral component of presynaptic membrane / Schaffer collateral - CA1 synapse / PDZ domain binding / presynaptic membrane / terminal bouton / response to calcium ion / protein tetramerization / establishment of protein localization / modulation of chemical synaptic transmission / growth cone / chemical synaptic transmission / amyloid-beta binding / postsynaptic membrane / ATPase binding / perikaryon / signaling receptor activity / dendritic spine / postsynaptic density / synapse / cell junction / glutamatergic synapse / dendrite / neuronal cell body / endoplasmic reticulum membrane / protein kinase binding / integral component of plasma membrane / cell surface / protein-containing complex / identical protein binding / plasma membrane / cytosol
PMP-22/EMP/MP20/Claudin superfamily / Ionotropic glutamate receptor, L-glutamate and glycine-binding domain / Receptor, ligand binding region / Ionotropic glutamate receptor, metazoa / Ionotropic glutamate receptor / Voltage-dependent calcium channel, gamma subunit / Voltage-dependent calcium channel, gamma-2 subunit / Periplasmic binding protein-like I
Voltage-dependent calcium channel gamma-2 subunit / Glutamate receptor 2
Biological speciesRattus norvegicus (Norway rat) / Mus musculus (house mouse)
Methodsingle particle reconstruction / cryo EM / Resolution: 4.2 Å
AuthorsTwomey EC / Yelshanskaya MV / Grassucci RA / Frank J / Sobolevsky AI
Funding support United States, 5 items
OrganizationGrant numberCountry
National Institutes of Health/National Institute of Neurological Disorders and Stroke (NIH/NINDS)NS093838 United States
National Institutes of Health/National Cancer Institute (NIH/NCI)CA206573 United States
National Institutes of Health/National Institute of Neurological Disorders and Stroke (NIH/NINDS)NS083660 United States
Howard Hughes Medical Institute (HHMI) United States
National Institutes of Health/National Institute of General Medical Sciences (NIH/NIGMS)GM029169 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 ReportPDB-ID: 5weo

SummaryFull reportAbout validation report
History
DepositionJul 10, 2017-
Header (metadata) releaseAug 2, 2017-
Map releaseAug 2, 2017-
UpdateNov 20, 2019-
Current statusNov 20, 2019Processing site: RCSB / Status: Released

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Structure visualization

Movie
  • Surface view with section colored by density value
  • Surface level: 0.04
  • Imaged by UCSF Chimera
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  • Surface view colored by cylindrical radius
  • Surface level: 0.04
  • Imaged by UCSF Chimera
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  • Surface view with fitted model
  • Atomic models: : PDB-5weo
  • Surface level: 0.04
  • Imaged by UCSF Chimera
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Movie viewer
Structure viewerEM map:
SurfViewMolmilJmol/JSmol
Supplemental images

Downloads & links

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Map

FileDownload / File: emd_8823.map.gz / Format: CCP4 / Size: 103 MB / Type: IMAGE STORED AS FLOATING POINT NUMBER (4 BYTES)
Projections & slices

Image control

Size
Brightness
Contrast
Others
AxesZ (Sec.)Y (Row.)X (Col.)
1.08 Å/pix.
x 300 pix.
= 324. Å
1.08 Å/pix.
x 300 pix.
= 324. Å
1.08 Å/pix.
x 300 pix.
= 324. Å

Surface

Projections

Slices (1/3)

Slices (1/2)

Slices (2/3)

Images are generated by Spider.

Voxel sizeX=Y=Z: 1.08 Å
Density
Contour LevelBy AUTHOR: 0.04 / Movie #1: 0.04
Minimum - Maximum-0.09160006 - 0.16419825
Average (Standard dev.)0.0006511989 (±0.0055219787)
SymmetrySpace group: 1
Details

EMDB XML:

Map geometry
Axis orderXYZ
Origin000
Dimensions300300300
Spacing300300300
CellA=B=C: 324.0 Å
α=β=γ: 90.0 °

CCP4 map header:

modeImage stored as Reals
Å/pix. X/Y/Z1.081.081.08
M x/y/z300300300
origin x/y/z0.0000.0000.000
length x/y/z324.000324.000324.000
α/β/γ90.00090.00090.000
MAP C/R/S123
start NC/NR/NS000
NC/NR/NS300300300
D min/max/mean-0.0920.1640.001

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Supplemental data

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Sample components

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Entire Activated GluA2 complex bound to glutamate, cyclothiazide, and ST...

EntireName: Activated GluA2 complex bound to glutamate, cyclothiazide, and STZ in digitonin
Number of components: 4

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Component #1: protein, Activated GluA2 complex bound to glutamate, cyclothiazid...

ProteinName: Activated GluA2 complex bound to glutamate, cyclothiazide, and STZ in digitonin
Recombinant expression: No
SourceSpecies: Rattus norvegicus (Norway rat)
Source (engineered)Expression System: Homo sapiens (human) / Cell of expression system: HEK293 gnti-

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Component #2: protein, Glutamate receptor 2,Voltage-dependent calcium channel g...

ProteinName: Glutamate receptor 2,Voltage-dependent calcium channel gamma-2 subunit chimera
Number of Copies: 4 / Recombinant expression: No
MassTheoretical: 115.501969 kDa
SourceSpecies: Mus musculus (house mouse)
Source (engineered)Expression System: Homo sapiens (human)

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Component #3: ligand, GLUTAMIC ACID

LigandName: GLUTAMIC ACID / Number of Copies: 4 / Recombinant expression: No
MassTheoretical: 0.147129 kDa

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Component #4: ligand, CYCLOTHIAZIDE

LigandName: CYCLOTHIAZIDE / Number of Copies: 4 / Recombinant expression: No
MassTheoretical: 0.389878 kDa

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Experimental details

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Sample preparation

SpecimenSpecimen state: Particle / Method: cryo EM
Sample solutionSpecimen conc.: 4 mg/mL / pH: 8
VitrificationInstrument: FEI VITROBOT MARK IV / Cryogen name: ETHANE / Temperature: 295 K / Humidity: 100 %

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Electron microscopy imaging

Experimental equipment
Model: Titan Krios / Image courtesy: FEI Company
ImagingMicroscope: FEI TITAN KRIOS
Electron gunElectron source: FIELD EMISSION GUN / Accelerating voltage: 300 kV / Electron dose: 55 e/Å2 / Illumination mode: SPOT SCAN
LensImaging mode: BRIGHT FIELD
Specimen HolderModel: OTHER
CameraDetector: GATAN K2 SUMMIT (4k x 4k)

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Image processing

ProcessingMethod: single particle reconstruction / Applied symmetry: C2 (2 fold cyclic) / Number of projections: 69207
3D reconstructionResolution: 4.2 Å / Resolution method: FSC 0.143 CUT-OFF
Details: TMD map uploaded under additional files is at 4.0 angstrom resolution.

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Atomic model buiding

Modeling #1Refinement space: REAL
Output model

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