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- EMDB-47298: Ligand-binding and transmembrane domains of kainate receptor GluK... -

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Entry
Database: EMDB / ID: EMD-47298
TitleLigand-binding and transmembrane domains of kainate receptor GluK2 in complex with positive allosteric modulator BPAM-344 and channel blocker Kukoamine-A
Map dataLigand-binding and transmembrane domains of kainate receptor GluK2 in complex with positive allosteric modulator BPAM-344 and channel blocker Kukoamine-A
Sample
  • Complex: Ligand-binding and transmembrane domains of kainate receptor GluK2 in complex with positive allosteric modulator BPAM-344 and channel blocker Kukoamine-A
    • Protein or peptide: Glutamate receptor ionotropic, kainate 2
  • Ligand: 4-cyclopropyl-7-fluoro-3,4-dihydro-2H-1,2,4-benzothiadiazine 1,1-dioxide
  • Ligand: (2S)-3-(hexadecanoyloxy)-2-[(9Z)-octadec-9-enoyloxy]propyl 2-(trimethylammonio)ethyl phosphate
  • Ligand: 2-acetamido-2-deoxy-beta-D-glucopyranose
  • Ligand: CHOLESTEROL
  • Ligand: 1-deoxy-alpha-D-mannopyranose
  • Ligand: N,N'-[butane-1,4-diylbis(azanediylpropane-3,1-diyl)]bis[3-(3,4-dihydroxyphenyl)propanamide]
KeywordsKainate receptor / GluK2 / Positive allosteric modulator / BPAM344 / Channel blocker / Kukoamine-A / membrane protein
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 / negative regulation of synaptic transmission, glutamatergic / regulation of short-term neuronal synaptic plasticity / inhibitory postsynaptic potential / regulation of JNK cascade / glutamate receptor activity ...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 / negative regulation of synaptic transmission, glutamatergic / regulation of short-term neuronal synaptic plasticity / inhibitory postsynaptic potential / regulation of JNK cascade / glutamate receptor activity / ubiquitin conjugating enzyme binding / receptor clustering / modulation of excitatory postsynaptic potential / kainate selective glutamate receptor activity / ionotropic glutamate receptor complex / extracellularly glutamate-gated ion channel activity / behavioral fear response / positive regulation of synaptic transmission / 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 / dendrite cytoplasm / presynaptic modulation of chemical synaptic transmission / hippocampal mossy fiber to CA3 synapse / SNARE binding / excitatory postsynaptic potential / synaptic transmission, glutamatergic / transmitter-gated monoatomic ion channel activity involved in regulation of postsynaptic membrane potential / regulation of membrane potential / PDZ domain binding / regulation of long-term neuronal synaptic plasticity / modulation of chemical synaptic transmission / postsynaptic density membrane / terminal bouton / intracellular calcium ion homeostasis / positive regulation of neuron apoptotic process / presynaptic membrane / scaffold protein binding / perikaryon / chemical synaptic transmission / neuron apoptotic process / postsynaptic membrane / negative regulation of neuron apoptotic process / postsynaptic density / axon / neuronal cell body / dendrite / synapse / 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)
Methodsingle particle reconstruction / cryo EM / Resolution: 3.75 Å
AuthorsYen LY / Newton TP / Gangwar SP / Yelshanskaya MV / Sobolevsky AI
Funding support United States, 4 items
OrganizationGrant numberCountry
National Institutes of Health/National Institute of Neurological Disorders and Stroke (NIH/NINDS)R01 NS083660 United States
National Institutes of Health/National Institute of Neurological Disorders and Stroke (NIH/NINDS)R01 NS107253 United States
National Institutes of Health/National Institute of Arthritis and Musculoskeletal and Skin Diseases (NIH/NIAMS)R01 AR078814 United States
National Institutes of Health/National Cancer Institute (NIH/NCI)R01 CA206573 United States
CitationJournal: Nat Commun / Year: 2024
Title: Trapping of spermine, Kukoamine A, and polyamine toxin blockers in GluK2 kainate receptor channels.
Authors: Shanti Pal Gangwar / Maria V Yelshanskaya / Muhammed Aktolun / Laura Y Yen / Thomas P Newton / Kristian Strømgaard / Maria G Kurnikova / Alexander I Sobolevsky /
Abstract: Kainate receptors (KARs) are a subtype of ionotropic glutamate receptor (iGluR) channels, a superfamily of ligand-gated ion channels which mediate the majority of excitatory neurotransmission in the ...Kainate receptors (KARs) are a subtype of ionotropic glutamate receptor (iGluR) channels, a superfamily of ligand-gated ion channels which mediate the majority of excitatory neurotransmission in the central nervous system. KARs modulate neuronal circuits and plasticity during development and are implicated in neurological disorders, including epilepsy, depression, schizophrenia, anxiety, and autism. Calcium-permeable KARs undergo ion channel block, but the therapeutic potential of channel blockers remains underdeveloped, mainly due to limited structural knowledge. Here, we present closed-state structures of GluK2 KAR homotetramers in complex with ion channel blockers NpTx-8, PhTx-74, Kukoamine A, and spermine. We find that blockers reside inside the GluK2 ion channel pore, intracellular to the closed M3 helix bundle-crossing gate, with their hydrophobic heads filling the central cavity and positively charged polyamine tails spanning the selectivity filter. Molecular dynamics (MD) simulations of our structures illuminate interactions responsible for different affinity and binding poses of the blockers. Our structures elucidate the trapping mechanism of KAR channel block and provide a template for designing new blockers that can selectively target calcium-permeable KARs in neuropathologies.
History
DepositionOct 11, 2024-
Header (metadata) releaseDec 11, 2024-
Map releaseDec 11, 2024-
UpdateDec 11, 2024-
Current statusDec 11, 2024Processing site: RCSB / Status: Released

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Structure visualization

Supplemental images

emd_47298.png

Downloads & links

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Map

FileDownload / File: emd_47298.map.gz / Format: CCP4 / Size: 178 MB / Type: IMAGE STORED AS FLOATING POINT NUMBER (4 BYTES)
AnnotationLigand-binding and transmembrane domains of kainate receptor GluK2 in complex with positive allosteric modulator BPAM-344 and channel blocker Kukoamine-A
Projections & slices

Image control

Size
Brightness
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Others
AxesX (Sec.)Y (Row.)Z (Col.)
1.07 Å/pix.
x 360 pix.
= 384.84 Å		1.07 Å/pix.
x 360 pix.
= 384.84 Å		1.07 Å/pix.
x 360 pix.
= 384.84 Å

Surface

Projections

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Images are generated by Spider.

Voxel sizeX=Y=Z: 1.069 Å
Density

Histogram

Histogram (log scale)
Contour LevelBy AUTHOR: 0.132
Minimum - Maximum-0.5958187 - 1.2153387
Average (Standard dev.)0.0005073188 (±0.01866883)
SymmetrySpace group: 1
Details

EMDB XML:

Map geometry
Axis orderZYX
Origin000
Dimensions360360360
Spacing360360360
CellA=B=C: 384.84 Å
α=β=γ: 90.0 °

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Supplemental data

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Half map: #2

Fileemd_47298_half_map_1.map
Projections & Slices
AxesZYX

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Slices (1/2)
Density Histograms

Histogram

Histogram (log scale)

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Half map: #1

Fileemd_47298_half_map_2.map
Projections & Slices
AxesZYX

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Histogram (log scale)

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Sample components

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Entire : Ligand-binding and transmembrane domains of kainate receptor GluK...

EntireName: Ligand-binding and transmembrane domains of kainate receptor GluK2 in complex with positive allosteric modulator BPAM-344 and channel blocker Kukoamine-A
Components
  • Complex: Ligand-binding and transmembrane domains of kainate receptor GluK2 in complex with positive allosteric modulator BPAM-344 and channel blocker Kukoamine-A
    • Protein or peptide: Glutamate receptor ionotropic, kainate 2
  • Ligand: 4-cyclopropyl-7-fluoro-3,4-dihydro-2H-1,2,4-benzothiadiazine 1,1-dioxide
  • Ligand: (2S)-3-(hexadecanoyloxy)-2-[(9Z)-octadec-9-enoyloxy]propyl 2-(trimethylammonio)ethyl phosphate
  • Ligand: 2-acetamido-2-deoxy-beta-D-glucopyranose
  • Ligand: CHOLESTEROL
  • Ligand: 1-deoxy-alpha-D-mannopyranose
  • Ligand: N,N'-[butane-1,4-diylbis(azanediylpropane-3,1-diyl)]bis[3-(3,4-dihydroxyphenyl)propanamide]

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Supramolecule #1: Ligand-binding and transmembrane domains of kainate receptor GluK...

SupramoleculeName: Ligand-binding and transmembrane domains of kainate receptor GluK2 in complex with positive allosteric modulator BPAM-344 and channel blocker Kukoamine-A
type: complex / ID: 1 / Parent: 0 / Macromolecule list: #1
Source (natural)Organism: Rattus norvegicus (Norway rat)

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Macromolecule #1: Glutamate receptor ionotropic, kainate 2

MacromoleculeName: Glutamate receptor ionotropic, kainate 2 / type: protein_or_peptide / ID: 1 / Number of copies: 4 / Enantiomer: LEVO
Source (natural)Organism: Rattus norvegicus (Norway rat)
Molecular weightTheoretical: 102.509977 KDa
Recombinant expressionOrganism: Homo sapiens (human)
SequenceString: MKIISPVLSN LVFSRSIKVL LCLLWIGYSQ GTTHVLRFGG IFEYVESGPM GAEELAFRFA VNTINRNRTL LPNTTLTYDT QKINLYDSF EASKKACDQL SLGVAAIFGP SHSSSANAVQ SICNALGVPH IQTRWKHQVS DNKDSFYVSL YPDFSSLSRA I LDLVQFFK ...String:
MKIISPVLSN LVFSRSIKVL LCLLWIGYSQ GTTHVLRFGG IFEYVESGPM GAEELAFRFA VNTINRNRTL LPNTTLTYDT QKINLYDSF EASKKACDQL SLGVAAIFGP SHSSSANAVQ SICNALGVPH IQTRWKHQVS DNKDSFYVSL YPDFSSLSRA I LDLVQFFK WKTVTVVYDD STGLIRLQEL IKAPSRYNLR LKIRQLPADT KDAKPLLKEM KRGKEFHVIF DCSHEMAAGI LK QALAMGM MTEYYHYIFT TLDLFALDVE PYRYSGVNMT GFRILNTENT QVSSIIEKWS MERLQAPPKP DSGLLDGFMT TDA ALMYDA VHVVSVAVQQ FPQMTVSSLQ CNRHKPWRFG TRFMSLIKEA HWEGLTGRIT FNKTNGLRTD FDLDVISLKE EGLE KIGTW DPASGLNMTE SQKGKPANIT DSLSNRSLIV TTILEEPYVL FKKSDKPLYG NDRFEGYCID LLRELSTILG FTYEI RLVE DGKYGAQDDV NGQWNGMVRE LIDHKADLAV APLAITYVRE KVIDFSKPFM TLGISILYRK PNGTNPGVFS FLNPLS PDI WMYVLLACLG VSCVLFVIAR FSPYEWYNPH PCNPDSDVVE NNFTLLNSFW FGVGALMQQG SELMPKALST RIVGGIW WF FTLIIISSYT ANLAAFLTVE RMESPIDSAD DLAKQTKIEY GAVEDGATMT FFKKSKISTY DKMWAFMSSR RQSVLVKS N EEGIQRVLTS DYAFLMESTT IEFVTQRNCN LTQIGGLIDS KGYGVGTPMG SPYRDKITIA ILQLQEEGKL HMMKEKWWR GNGCPEEESK EASALGVQNI GGIFIVLAAG LVLSVFVAVG EFLYKSKKNA QLEKRSFCSA MVEELRMSLK CQRRLKHKPQ APVIVKTEE VINMHTFNDR RLPGKETMA

UniProtKB: Glutamate receptor ionotropic, kainate 2

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Macromolecule #2: 4-cyclopropyl-7-fluoro-3,4-dihydro-2H-1,2,4-benzothiadiazine 1,1-...

MacromoleculeName: 4-cyclopropyl-7-fluoro-3,4-dihydro-2H-1,2,4-benzothiadiazine 1,1-dioxide
type: ligand / ID: 2 / Number of copies: 4 / Formula: 2J9
Molecular weightTheoretical: 242.27 Da
Chemical component information

ChemComp-2J9:
4-cyclopropyl-7-fluoro-3,4-dihydro-2H-1,2,4-benzothiadiazine 1,1-dioxide

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Macromolecule #3: (2S)-3-(hexadecanoyloxy)-2-[(9Z)-octadec-9-enoyloxy]propyl 2-(tri...

MacromoleculeName: (2S)-3-(hexadecanoyloxy)-2-[(9Z)-octadec-9-enoyloxy]propyl 2-(trimethylammonio)ethyl phosphate
type: ligand / ID: 3 / Number of copies: 8 / Formula: POV
Molecular weightTheoretical: 760.076 Da
Chemical component information

ChemComp-POV:
(2S)-3-(hexadecanoyloxy)-2-[(9Z)-octadec-9-enoyloxy]propyl 2-(trimethylammonio)ethyl phosphate / phospholipid*YM

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Macromolecule #4: 2-acetamido-2-deoxy-beta-D-glucopyranose

MacromoleculeName: 2-acetamido-2-deoxy-beta-D-glucopyranose / type: ligand / ID: 4 / Number of copies: 16 / Formula: NAG
Molecular weightTheoretical: 221.208 Da
Chemical component information

ChemComp-NAG:
2-acetamido-2-deoxy-beta-D-glucopyranose

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Macromolecule #5: CHOLESTEROL

MacromoleculeName: CHOLESTEROL / type: ligand / ID: 5 / Number of copies: 8 / Formula: CLR
Molecular weightTheoretical: 386.654 Da
Chemical component information

ChemComp-CLR:
CHOLESTEROL

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Macromolecule #6: 1-deoxy-alpha-D-mannopyranose

MacromoleculeName: 1-deoxy-alpha-D-mannopyranose / type: ligand / ID: 6 / Number of copies: 4 / Formula: AH2
Molecular weightTheoretical: 164.156 Da
Chemical component information

ChemComp-AH2:
1-deoxy-alpha-D-mannopyranose

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Macromolecule #7: N,N'-[butane-1,4-diylbis(azanediylpropane-3,1-diyl)]bis[3-(3,4-di...

MacromoleculeName: N,N'-[butane-1,4-diylbis(azanediylpropane-3,1-diyl)]bis[3-(3,4-dihydroxyphenyl)propanamide]
type: ligand / ID: 7 / Number of copies: 1 / Formula: A1BDT
Molecular weightTheoretical: 530.656 Da

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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

BufferpH: 8
Component:
ConcentrationName
150.0 mMSodium Chloride
20.0 mMTris(hydroxymethyl)aminomethane
0.05 %Digitonin
5.0 mMBeta-Mercaptoethanol
GridModel: UltrAuFoil R1.2/1.3 / Material: GOLD / Support film - Material: GOLD / Support film - topology: HOLEY
VitrificationCryogen name: ETHANE

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Electron microscopy

MicroscopeTFS KRIOS
Image recordingFilm or detector model: GATAN K3 (6k x 4k) / Average electron dose: 47.25 e/Å2
Electron beamAcceleration voltage: 300 kV / Electron source: FIELD EMISSION GUN
Electron opticsIllumination mode: FLOOD BEAM / Imaging mode: BRIGHT FIELD / Nominal defocus max: 2.0 µm / Nominal defocus min: 1.0 µm
Experimental equipment
Model: Titan Krios / Image courtesy: FEI Company

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Image processing

Startup modelType of model: OTHER
Final reconstructionResolution.type: BY AUTHOR / Resolution: 3.75 Å / Resolution method: FSC 0.143 CUT-OFF / Number images used: 211509
Initial angle assignmentType: OTHER
Final angle assignmentType: OTHER
FSC plot (resolution estimation)

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Atomic model buiding 1

RefinementSpace: REAL
Output model

PDB-9dxt:
Ligand-binding and transmembrane domains of kainate receptor GluK2 in complex with positive allosteric modulator BPAM-344 and channel blocker Kukoamine-A

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