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- EMDB-2685: Density map of GluK2 desensitized state in complex with 2S,4R-4-m... -

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Basic information

Entry
Database: EMDB / ID: EMD-2685
TitleDensity map of GluK2 desensitized state in complex with 2S,4R-4-methylglutamate
Map dataReconstruction of GluK2 desensitized state
Sample
  • Sample: GluAK2 with 2S,4R-4-methylglutamate
  • Protein or peptide: GluK2
  • Ligand: 2S,4R-4-methylglutamate
KeywordsMembrane Protein / Ion Channel / Glutamate Receptor
Function / homology
Function and homology information


mossy fiber rosette / detection of cold stimulus involved in thermoception / Activation of Na-permeable kainate receptors / kainate selective glutamate receptor complex / Activation of Ca-permeable Kainate Receptor / regulation of short-term neuronal synaptic plasticity / glutamate receptor activity / negative regulation of synaptic transmission, glutamatergic / ubiquitin conjugating enzyme binding / regulation of JNK cascade ...mossy fiber rosette / detection of cold stimulus involved in thermoception / Activation of Na-permeable kainate receptors / kainate selective glutamate receptor complex / Activation of Ca-permeable Kainate Receptor / regulation of short-term neuronal synaptic plasticity / glutamate receptor activity / negative regulation of synaptic transmission, glutamatergic / ubiquitin conjugating enzyme binding / regulation of JNK cascade / inhibitory postsynaptic potential / receptor clustering / kainate selective glutamate receptor activity / modulation of excitatory postsynaptic potential / extracellularly glutamate-gated ion channel activity / ionotropic glutamate receptor complex / positive regulation of synaptic transmission / behavioral fear response / neuronal action potential / glutamate-gated receptor activity / glutamate-gated calcium ion channel activity / ligand-gated monoatomic ion channel activity involved in regulation of presynaptic membrane potential / presynaptic modulation of chemical synaptic transmission / dendrite cytoplasm / hippocampal mossy fiber to CA3 synapse / SNARE binding / regulation of membrane potential / excitatory postsynaptic potential / transmitter-gated monoatomic ion channel activity involved in regulation of postsynaptic membrane potential / synaptic transmission, glutamatergic / PDZ domain binding / regulation of long-term neuronal synaptic plasticity / postsynaptic density membrane / modulation of chemical synaptic transmission / intracellular calcium ion homeostasis / terminal bouton / positive regulation of neuron apoptotic process / presynaptic membrane / neuron apoptotic process / scaffold protein binding / perikaryon / chemical synaptic transmission / negative regulation of neuron apoptotic process / postsynaptic membrane / postsynaptic density / axon / neuronal cell body / synapse / dendrite / ubiquitin protein ligase binding / glutamatergic synapse / identical protein binding / membrane / plasma membrane
Similarity search - Function
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
Glutamate receptor ionotropic, kainate 2
Similarity search - Component
Biological speciesRattus norvegicus (Norway rat) / synthetic construct (others)
Methodsingle particle reconstruction / cryo EM / Resolution: 7.6 Å
AuthorsMeyerson JR / Kumar J / Chittori S / Rao P / Pierson J / Bartesaghi A / Mayer ML / Subramaniam S
CitationJournal: Nature / Year: 2014
Title: Structural mechanism of glutamate receptor activation and desensitization.
Authors: Joel R Meyerson / Janesh Kumar / Sagar Chittori / Prashant Rao / Jason Pierson / Alberto Bartesaghi / Mark L Mayer / Sriram Subramaniam /
Abstract: Ionotropic glutamate receptors are ligand-gated ion channels that mediate excitatory synaptic transmission in the vertebrate brain. To gain a better understanding of how structural changes gate ion ...Ionotropic glutamate receptors are ligand-gated ion channels that mediate excitatory synaptic transmission in the vertebrate brain. To gain a better understanding of how structural changes gate ion flux across the membrane, we trapped rat AMPA (α-amino-3-hydroxy-5-methyl-4-isoxazole propionic acid) and kainate receptor subtypes in their major functional states and analysed the resulting structures using cryo-electron microscopy. We show that transition to the active state involves a 'corkscrew' motion of the receptor assembly, driven by closure of the ligand-binding domain. Desensitization is accompanied by disruption of the amino-terminal domain tetramer in AMPA, but not kainate, receptors with a two-fold to four-fold symmetry transition in the ligand-binding domains in both subtypes. The 7.6 Å structure of a desensitized kainate receptor shows how these changes accommodate channel closing. These findings integrate previous physiological, biochemical and structural analyses of glutamate receptors and provide a molecular explanation for key steps in receptor gating.
History
DepositionJun 20, 2014-
Header (metadata) releaseJul 30, 2014-
Map releaseAug 13, 2014-
UpdateOct 29, 2014-
Current statusOct 29, 2014Processing site: PDBe / Status: Released

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Structure visualization

Movie
  • Surface view with section colored by density value
  • Surface level: 0.121
  • Imaged by UCSF Chimera
  • Download
  • Surface view colored by height
  • Surface level: 0.121
  • Imaged by UCSF Chimera
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  • Surface view with fitted model
  • Atomic models: PDB-4uqq
  • Surface level: 0.121
  • Imaged by UCSF Chimera
  • Download
  • Simplified surface model + fitted atomic model
  • Atomic modelsPDB-4uqq
  • Imaged by Jmol
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Movie viewer
Structure viewerEM map:
SurfViewMolmilJmol/JSmol
Supplemental images

EMD-2685-Glu...

Downloads & links

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Map

FileDownload / File: emd_2685.map.gz / Format: CCP4 / Size: 29.8 MB / Type: IMAGE STORED AS FLOATING POINT NUMBER (4 BYTES)
AnnotationReconstruction of GluK2 desensitized state
Projections & slices

Image control

Size
Brightness
Contrast
Others
AxesZ (Sec.)Y (Row.)X (Col.)
1.41 Å/pix.
x 200 pix.
= 281.2 Å		1.41 Å/pix.
x 200 pix.
= 281.2 Å		1.41 Å/pix.
x 200 pix.
= 281.2 Å

Surface

Projections

Slices (1/3)

Slices (1/2)

Slices (2/3)

Images are generated by Spider.

Voxel sizeX=Y=Z: 1.406 Å
Density

Histogram

Histogram (log scale)
Contour LevelBy AUTHOR: 0.121 / Movie #1: 0.121
Minimum - Maximum-0.27725154 - 0.47987753
Average (Standard dev.)0.00528497 (±0.02724039)
SymmetrySpace group: 1
Details

EMDB XML:

Map geometry
Axis orderXYZ
Origin100100100
Dimensions200200200
Spacing200200200
CellA=B=C: 281.2 Å
α=β=γ: 90.0 °

CCP4 map header:

modeImage stored as Reals
Å/pix. X/Y/Z1.4061.4061.406
M x/y/z200200200
origin x/y/z0.0000.0000.000
length x/y/z281.200281.200281.200
α/β/γ90.00090.00090.000
start NX/NY/NZ-180-180-179
NX/NY/NZ360360360
MAP C/R/S123
start NC/NR/NS100100100
NC/NR/NS200200200
D min/max/mean-0.2770.4800.005

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Supplemental data

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Sample components

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Entire : GluAK2 with 2S,4R-4-methylglutamate

EntireName: GluAK2 with 2S,4R-4-methylglutamate
Components
  • Sample: GluAK2 with 2S,4R-4-methylglutamate
  • Protein or peptide: GluK2
  • Ligand: 2S,4R-4-methylglutamate

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Supramolecule #1000: GluAK2 with 2S,4R-4-methylglutamate

SupramoleculeName: GluAK2 with 2S,4R-4-methylglutamate / type: sample / ID: 1000 / Number unique components: 2
Molecular weightExperimental: 400 KDa / Theoretical: 400 KDa

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Macromolecule #1: GluK2

MacromoleculeName: GluK2 / type: protein_or_peptide / ID: 1 / Oligomeric state: Tetramer / Recombinant expression: Yes
Source (natural)Organism: Rattus norvegicus (Norway rat) / synonym: Rat
Molecular weightExperimental: 400 KDa / Theoretical: 400 KDa
Recombinant expressionOrganism: Spodoptera frugiperda (fall armyworm) / Recombinant cell: Sf9 / Recombinant plasmid: pFastBac1
SequenceUniProtKB: Glutamate receptor ionotropic, kainate 2

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Macromolecule #2: 2S,4R-4-methylglutamate

MacromoleculeName: 2S,4R-4-methylglutamate / type: ligand / ID: 2 / Recombinant expression: No
Source (natural)Organism: synthetic construct (others)
Chemical component information

ChemComp-SYM:
2S,4R-4-METHYLGLUTAMATE

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Experimental details

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Structure determination

Methodcryo EM
Processingsingle particle reconstruction
Aggregation stateparticle

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Sample preparation

Concentration1.8 mg/mL
BufferpH: 8
Details: 150 mM NaCl, 20 mM Tris pH 8.0, 0.75 mM DDM, 1 mM 2S,4R-4-methylglutamate
GridDetails: Vitrified specimens were prepared by adding 2.5 uL of liganded protein at 1.8 mg/ml to R2/2 holey carbon grids (Quantifoil, Jena, Germany) rendered hydrophilic by chemical treatment to ...Details: Vitrified specimens were prepared by adding 2.5 uL of liganded protein at 1.8 mg/ml to R2/2 holey carbon grids (Quantifoil, Jena, Germany) rendered hydrophilic by chemical treatment to enable particle distribution into the holes (Meyerson JR, Rao P, Kumar K, Chittori S, Banerjee S, Pierson J, Mayer ML, and Subramaniam S, manuscript in preparation).
VitrificationCryogen name: ETHANE / Chamber humidity: 100 % / Chamber temperature: 120 K / Instrument: FEI VITROBOT MARK IV / Method: Blot for 2 seconds before plunging

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Electron microscopy

MicroscopeFEI TITAN KRIOS
DateAug 1, 2013
Image recordingCategory: CCD / Film or detector model: FEI FALCON II (4k x 4k) / Number real images: 4837 / Average electron dose: 25 e/Å2
Electron beamAcceleration voltage: 300 kV / Electron source: FIELD EMISSION GUN
Electron opticsCalibrated magnification: 47000 / Illumination mode: FLOOD BEAM / Imaging mode: BRIGHT FIELD / Nominal defocus max: 3.5 µm / Nominal defocus min: 2.0 µm / Nominal magnification: 47000
Sample stageSpecimen holder model: FEI TITAN KRIOS AUTOGRID HOLDER
Experimental equipment
Model: Titan Krios / Image courtesy: FEI Company

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Image processing

DetailsParticles were selected interactively at the computer terminal
CTF correctionDetails: Each particle
Final reconstructionApplied symmetry - Point group: C2 (2 fold cyclic) / Resolution.type: BY AUTHOR / Resolution: 7.6 Å / Resolution method: OTHER / Software - Name: Relion / Number images used: 21360

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