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- PDB-6irf: Structure of the human GluN1/GluN2A NMDA receptor in the glutamat... -

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Entry
Database: PDB / ID: 6irf
TitleStructure of the human GluN1/GluN2A NMDA receptor in the glutamate/glycine-bound state at pH 6.3, Class I
Components
  • Glutamate receptor ionotropic, NMDA 1
  • Glutamate receptor ionotropic, NMDA 2A
KeywordsMEMBRANE PROTEIN / ionotropic glutamate receptors / NMDA receptors / synaptic protein
Function / homologyIonotropic glutamate receptor, metazoa / CREB1 phosphorylation through the activation of CaMKII/CaMKK/CaMKIV cascasde / N-methyl D-aspartate receptor 2B3 C-terminus / Calmodulin-binding domain C0 of NMDA receptor NR1 subunit / Ligated ion channel L-glutamate- and glycine-binding site / EPHB-mediated forward signaling / Receptor, ligand binding region / Unblocking of NMDA receptors, glutamate binding and activation / Receptor family ligand binding region / Ligand-gated ion channel ...Ionotropic glutamate receptor, metazoa / CREB1 phosphorylation through the activation of CaMKII/CaMKK/CaMKIV cascasde / N-methyl D-aspartate receptor 2B3 C-terminus / Calmodulin-binding domain C0 of NMDA receptor NR1 subunit / Ligated ion channel L-glutamate- and glycine-binding site / EPHB-mediated forward signaling / Receptor, ligand binding region / Unblocking of NMDA receptors, glutamate binding and activation / Receptor family ligand binding region / Ligand-gated ion channel / Periplasmic binding protein-like I / Ionotropic glutamate receptor, L-glutamate and glycine-binding domain / Glutamate [NMDA] receptor, epsilon subunit, C-terminal / Ras activation upon Ca2+ influx through NMDA receptor / Synaptic adhesion-like molecules / Ionotropic glutamate receptor / RAF/MAP kinase cascade / MECP2 regulates neuronal receptors and channels / Calmodulin-binding domain C0, NMDA receptor, NR1 subunit / Neurexins and neuroligins / response to glycine / propylene metabolic process / pons maturation / protein localization to postsynaptic membrane / directional locomotion / excitatory chemical synaptic transmission / regulation of respiratory gaseous exchange / conditioned taste aversion / olfactory learning / male mating behavior / serotonin metabolic process / glutamate receptor signaling pathway / startle response / regulation of synapse assembly / sleep / glutamate-gated calcium ion channel activity / neurotransmitter binding / calcium ion transmembrane import into cytosol / cation transport / neurogenesis / respiratory gaseous exchange / calcium ion homeostasis / activation of cysteine-type endopeptidase activity / glycine binding / NMDA glutamate receptor activity / glutamate binding / NMDA selective glutamate receptor complex / regulation of dendrite morphogenesis / dopamine metabolic process / suckling behavior / synaptic membrane / positive regulation of cysteine-type endopeptidase activity / social behavior / positive regulation of reactive oxygen species biosynthetic process / regulation of axonogenesis / synaptic transmission, glutamatergic / postsynaptic density membrane / positive regulation of calcium ion transport into cytosol / response to morphine / excitatory synapse / long-term memory / excitatory postsynaptic potential / synaptic cleft / positive regulation of excitatory postsynaptic potential / sensory perception of pain / prepulse inhibition / adult locomotory behavior / regulation of membrane potential / cerebral cortex development / integral component of postsynaptic density membrane / regulation of sensory perception of pain / transmitter-gated ion channel activity involved in regulation of postsynaptic membrane potential / response to amphetamine / regulation of synaptic plasticity / visual learning / long-term synaptic potentiation / ionotropic glutamate receptor signaling pathway / memory / regulation of long-term neuronal synaptic plasticity / positive regulation of long-term synaptic potentiation / presynaptic membrane / terminal bouton / synaptic vesicle / ephrin receptor signaling pathway / negative regulation of protein catabolic process / protein heterotetramerization / brain development / postsynaptic membrane / response to wounding / learning or memory / Ras guanyl-nucleotide exchange factor activity / chemical synaptic transmission / amyloid-beta binding / negative regulation of neuron apoptotic process / calmodulin binding / response to ethanol / dendritic spine / MAPK cascade / postsynaptic density / protein-containing complex binding
Function and homology information
Specimen sourceHomo sapiens (human)
MethodELECTRON MICROSCOPY / single particle reconstruction / cryo EM / 5.1 Å resolution
AuthorsZhang, J. / Chang, S. / Zhang, X. / Zhu, S.
CitationJournal: Cell Rep / Year: 2018
Title: Structural Basis of the Proton Sensitivity of Human GluN1-GluN2A NMDA Receptors.
Authors: Jin-Bao Zhang / Shenghai Chang / Pan Xu / Miao Miao / Hangjun Wu / Youyi Zhang / Tongtong Zhang / Han Wang / Jilin Zhang / Chun Xie / Nan Song / Cheng Luo / Xing Zhang / Shujia Zhu
Abstract: N-methyl-D-aspartate (NMDA) receptors are critical for synaptic development and plasticity. While glutamate is the primary agonist, protons can modulate NMDA receptor activity at synapses during ...N-methyl-D-aspartate (NMDA) receptors are critical for synaptic development and plasticity. While glutamate is the primary agonist, protons can modulate NMDA receptor activity at synapses during vesicle exocytosis by mechanisms that are unknown. We used cryo-electron microscopy to solve the structures of the human GluN1-GluN2A NMDA receptor at pH 7.8 and pH 6.3. Our structures demonstrate that the proton sensor predominantly resides in the N-terminal domain (NTD) of the GluN2A subunit and reveal the allosteric coupling mechanism between the proton sensor and the channel gate. Under high-pH conditions, the GluN2A-NTD adopts an "open-and-twisted" conformation. However, upon protonation at the lower pH, the GluN2A-NTD transits from an open- to closed-cleft conformation, causing rearrangements between the tetrameric NTDs and agonist-binding domains. The conformational mobility observed in our structures (presumably from protonation) is supported by molecular dynamics simulation. Our findings reveal the structural mechanisms by which protons allosterically inhibit human GluN1-GluN2A receptor activity.
Validation Report
SummaryFull reportAbout validation report
DateDeposition: Nov 12, 2018 / Release: Jan 16, 2019

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Assembly

Deposited unit
A: Glutamate receptor ionotropic, NMDA 1
B: Glutamate receptor ionotropic, NMDA 2A
C: Glutamate receptor ionotropic, NMDA 1
D: Glutamate receptor ionotropic, NMDA 2A


Theoretical massNumber of molelcules
Total (without water)379,0574
Polyers379,0574
Non-polymers00
Water0
1


TypeNameSymmetry operationNumber
identity operation1_5551
Buried area (Å2)22060
ΔGint (kcal/M)-167
Surface area (Å2)152730

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Components

#1: Protein/peptide Glutamate receptor ionotropic, NMDA 1 / N-methyl-D-aspartate receptor subunit NR1 / NMD-R1


Mass: 95336.219 Da / Num. of mol.: 2 / Details: 2 mM Glycine / Mutation: G612R / Source: (gene. exp.) Homo sapiens (human) / Gene: Grin1, NMDAR1 / Cell line (production host): HEK293S GnTl- / Production host: Homo sapiens (human) / References: UniProt: Q05586
#2: Protein/peptide Glutamate receptor ionotropic, NMDA 2A / N-methyl D-aspartate receptor subtype 2A / hNR2A


Mass: 94192.172 Da / Num. of mol.: 2 / Details: 2 mM L-Glutamate / Mutation: E656R, E657R / Source: (gene. exp.) Homo sapiens (human) / Gene: Grin2a / Cell line (production host): HEK293S GnTl- / Production host: Homo sapiens (human) / References: UniProt: Q12879

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Experimental details

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Experiment

ExperimentMethod: ELECTRON MICROSCOPY
EM experimentAggregation state: PARTICLE / Reconstruction method: single particle reconstruction

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Sample preparation

ComponentName: Human GluN1/GluN2A NMDA receptors in the glutamate/glycine bound state at pH 6.3, Class I
Type: COMPLEX / Details: with the presence of Glycine,L-glutamate and EDTA / Entity ID: 1, 2 / Source: RECOMBINANT
Molecular weightValue: 0.38 MDa / Experimental value: NO
Source (natural)Organism: Homo sapiens (human)
Source (recombinant)Cell: HEK293S GnTl- / Organism: Homo sapiens (human) / Plasmid: pEG-Bacmam
Buffer solutionDetails: Solutions were made fresh. / pH: 6.3
Buffer component
IDConc.NameFormulaBuffer ID
1150 mMsodium chlorideNaCl1
220 mMMESC6H13NO4S1
30.05 mMEDTA(HO2CCH2)2NCH2CH2N(CH2CO2H)21
40.005 mMCholesteryl Hemisuccinate Tris SaltC31H50O4(C4H11NO3)1
50.001 g/mLDigitoninC56H92O291
60.1 mMCHAPSOC32H58N2O8S1
72 mMGlycineNH2CH2COOH1
82 mML-Glutamic acid monosodium salt hydrateC5H8NNaO4(xH2O)1
SpecimenConc.: 3.5 mg/ml / Details: Tetrameric GluN1/GluN2A NMDA receptors / Embedding applied: NO / Shadowing applied: NO / Staining applied: NO / Vitrification applied: YES
Specimen supportDetails: 15 mA / Grid material: GOLD / Grid mesh size: 200 / Grid type: Quantifoil R1.2/1.3
VitrificationInstrument: FEI VITROBOT MARK II / Cryogen name: ETHANE / Humidity: 100 % / Chamber temperature: 291 kelvins
Details: blot for 2 seconds before plunging in liquid ethane

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Electron microscopy imaging

Experimental equipment
Model: Titan Krios / Image courtesy: FEI Company
MicroscopyMicroscope model: FEI TITAN KRIOS
Electron gunElectron source: FIELD EMISSION GUN / Accelerating voltage: 300 kV / Illumination mode: FLOOD BEAM
Electron lensMode: BRIGHT FIELDBright-field microscopy
Image recordingAverage exposure time: 12 sec. / Electron dose: 56 e/Å2 / Film or detector model: GATAN K2 SUMMIT (4k x 4k) / Number of grids imaged: 4
Image scansMovie frames/image: 40 / Used frames/image: 1-40

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Processing

EM software
IDNameCategory
1Gautomatchparticle selection
4GctfCTF correction
7UCSF Chimeramodel fitting
13PHENIXmodel refinement
CTF correctionType: PHASE FLIPPING AND AMPLITUDE CORRECTION
Particle selectionNumber of particles selected: 722287
3D reconstructionResolution: 5.1 Å / Resolution method: FSC 0.143 CUT-OFF / Number of particles: 193878 / Symmetry type: POINT
Atomic model buildingRef protocol: RIGID BODY FIT
Atomic model building
IDPDB-IDPdb chain ID 3D fitting ID
14PE5

4pe5

A,B,C,D1
25TQ0

5tq0

B1
35H8F

5h8f

A,B1

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