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- EMDB-44245: GluA2 flip Q in complex with TARPgamma2 at pH5, class23, structur... -

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Basic information

Entry
Database: EMDB / ID: EMD-44245
TitleGluA2 flip Q in complex with TARPgamma2 at pH5, class23, structure of LBD-TMD-TARPgamma2
Map dataGluA2 flip Q isoform in complex with TARPgamma2 at pH5.3, class23 structure of LBD-TMD-TARPgamma2
Sample
  • Complex: GluA2 (flip-Q isoform) in complex with TARPgamma2 at 4:4 stoichiometry
    • Protein or peptide: Voltage-dependent calcium channel gamma-2 subunit
    • Protein or peptide: Isoform Flip of Glutamate receptor 2
KeywordsAMPA receptor / ionotropic glutamate receptor / ion channel / auxiliary subunit / TRANSPORT PROTEIN
Function / homology
Function and homology information


Presynaptic depolarization and calcium channel opening / regulation of postsynaptic neurotransmitter receptor activity / LGI-ADAM interactions / Trafficking of AMPA receptors / eye blink reflex / positive regulation of protein localization to basolateral plasma membrane / cerebellar mossy fiber / neurotransmitter receptor transport, postsynaptic endosome to lysosome / regulation of AMPA receptor activity / neurotransmitter receptor internalization ...Presynaptic depolarization and calcium channel opening / regulation of postsynaptic neurotransmitter receptor activity / LGI-ADAM interactions / Trafficking of AMPA receptors / eye blink reflex / positive regulation of protein localization to basolateral plasma membrane / cerebellar mossy fiber / neurotransmitter receptor transport, postsynaptic endosome to lysosome / regulation of AMPA receptor activity / neurotransmitter receptor internalization / membrane hyperpolarization / postsynaptic neurotransmitter receptor diffusion trapping / nervous system process / protein targeting to membrane / voltage-gated calcium channel complex / neurotransmitter receptor localization to postsynaptic specialization membrane / neuromuscular junction development / spine synapse / dendritic spine neck / dendritic spine head / Activation of AMPA receptors / perisynaptic space / AMPA glutamate receptor activity / transmission of nerve impulse / ligand-gated monoatomic cation channel activity / channel regulator activity / Trafficking of GluR2-containing AMPA receptors / response to lithium ion / immunoglobulin binding / regulation of postsynaptic membrane neurotransmitter receptor levels / AMPA glutamate receptor complex / membrane depolarization / kainate selective glutamate receptor activity / ionotropic glutamate receptor complex / extracellularly glutamate-gated ion channel activity / cellular response to glycine / asymmetric synapse / regulation of receptor recycling / Unblocking of NMDA receptors, glutamate binding and activation / voltage-gated calcium channel activity / positive regulation of synaptic transmission / glutamate receptor binding / extracellular ligand-gated monoatomic ion channel activity / glutamate-gated receptor activity / response to fungicide / glutamate-gated calcium ion channel activity / presynaptic active zone membrane / regulation of synaptic transmission, glutamatergic / somatodendritic compartment / dendrite membrane / cellular response to brain-derived neurotrophic factor stimulus / cytoskeletal protein binding / ligand-gated monoatomic ion channel activity involved in regulation of presynaptic membrane potential / ionotropic glutamate receptor binding / dendrite cytoplasm / ionotropic glutamate receptor signaling pathway / hippocampal mossy fiber to CA3 synapse / positive regulation of synaptic transmission, glutamatergic / regulation of membrane potential / SNARE binding / dendritic shaft / synaptic transmission, glutamatergic / synaptic membrane / PDZ domain binding / transmitter-gated monoatomic ion channel activity involved in regulation of postsynaptic membrane potential / protein tetramerization / postsynaptic density membrane / establishment of protein localization / modulation of chemical synaptic transmission / Schaffer collateral - CA1 synapse / terminal bouton / receptor internalization / cerebral cortex development / synaptic vesicle membrane / response to calcium ion / synaptic vesicle / presynapse / signaling receptor activity / presynaptic membrane / amyloid-beta binding / growth cone / scaffold protein binding / chemical synaptic transmission / perikaryon / postsynaptic membrane / dendritic spine / postsynaptic density / neuron projection / axon / neuronal cell body / glutamatergic synapse / dendrite / synapse / protein-containing complex binding / protein kinase binding / cell surface / endoplasmic reticulum / protein-containing complex / identical protein binding / membrane
Similarity search - Function
Voltage-dependent calcium channel, gamma-2 subunit / : / PMP-22/EMP/MP20/Claudin family / Voltage-dependent calcium channel, gamma subunit / PMP-22/EMP/MP20/Claudin superfamily / 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 ...Voltage-dependent calcium channel, gamma-2 subunit / : / PMP-22/EMP/MP20/Claudin family / Voltage-dependent calcium channel, gamma subunit / PMP-22/EMP/MP20/Claudin superfamily / 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
Voltage-dependent calcium channel gamma-2 subunit / Glutamate receptor 2
Similarity search - Component
Biological speciesMus musculus (house mouse) / Rattus norvegicus (Norway rat)
Methodsingle particle reconstruction / cryo EM / Resolution: 3.56 Å
AuthorsNakagawa T / Greger IH
Funding support United States, United Kingdom, 4 items
OrganizationGrant numberCountry
National Institutes of Health/National Institute of Mental Health (NIH/NIMH)R01MH123474 United States
UK Research and Innovation (UKRI)MC_U105174197 United Kingdom
Biotechnology and Biological Sciences Research Council (BBSRC)BB/N002113/1 United Kingdom
Wellcome Trust223194/Z/21/Z United Kingdom
CitationJournal: Nat Struct Mol Biol / Year: 2024
Title: Proton-triggered rearrangement of the AMPA receptor N-terminal domains impacts receptor kinetics and synaptic localization.
Authors: Josip Ivica / Nejc Kejzar / Hinze Ho / Imogen Stockwell / Viktor Kuchtiak / Alexander M Scrutton / Terunaga Nakagawa / Ingo H Greger /
Abstract: AMPA glutamate receptors (AMPARs) are ion channel tetramers that mediate the majority of fast excitatory synaptic transmission. They are composed of four subunits (GluA1-GluA4); the GluA2 subunit ...AMPA glutamate receptors (AMPARs) are ion channel tetramers that mediate the majority of fast excitatory synaptic transmission. They are composed of four subunits (GluA1-GluA4); the GluA2 subunit dominates AMPAR function throughout the forebrain. Its extracellular N-terminal domain (NTD) determines receptor localization at the synapse, ensuring reliable synaptic transmission and plasticity. This synaptic anchoring function requires a compact NTD tier, stabilized by a GluA2-specific NTD interface. Here we show that low pH conditions, which accompany synaptic activity, rupture this interface. All-atom molecular dynamics simulations reveal that protonation of an interfacial histidine residue (H208) centrally contributes to NTD rearrangement. Moreover, in stark contrast to their canonical compact arrangement at neutral pH, GluA2 cryo-electron microscopy structures exhibit a wide spectrum of NTD conformations under acidic conditions. We show that the consequences of this pH-dependent conformational control are twofold: rupture of the NTD tier slows recovery from desensitized states and increases receptor mobility at mouse hippocampal synapses. Therefore, a proton-triggered NTD switch will shape both AMPAR location and kinetics, thereby impacting synaptic signal transmission.
History
DepositionMar 23, 2024-
Header (metadata) releaseJul 31, 2024-
Map releaseJul 31, 2024-
UpdateNov 6, 2024-
Current statusNov 6, 2024Processing site: RCSB / Status: Released

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Structure visualization

Supplemental images

emd_44245.png

Downloads & links

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Map

FileDownload / File: emd_44245.map.gz / Format: CCP4 / Size: 178 MB / Type: IMAGE STORED AS FLOATING POINT NUMBER (4 BYTES)
AnnotationGluA2 flip Q isoform in complex with TARPgamma2 at pH5.3, class23 structure of LBD-TMD-TARPgamma2
Projections & slices

Image control

Size
Brightness
Contrast
Others
AxesZ (Sec.)Y (Row.)X (Col.)
0.82 Å/pix.
x 360 pix.
= 295.2 Å		0.82 Å/pix.
x 360 pix.
= 295.2 Å		0.82 Å/pix.
x 360 pix.
= 295.2 Å

Surface

Projections

Slices (1/3)

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Slices (2/3)

Images are generated by Spider.

Voxel sizeX=Y=Z: 0.82 Å
Density

Histogram

Histogram (log scale)
Contour LevelBy AUTHOR: 0.00934
Minimum - Maximum-0.032694776 - 0.055235602
Average (Standard dev.)0.000029864996 (±0.0021393339)
SymmetrySpace group: 1
Details

EMDB XML:

Map geometry
Axis orderXYZ
Origin000
Dimensions360360360
Spacing360360360
CellA=B=C: 295.2 Å
α=β=γ: 90.0 °

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Supplemental data

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Additional map: Focused refinement of the NTDs

Fileemd_44245_additional_1.map
AnnotationFocused refinement of the NTDs
Projections & Slices
AxesZYX

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Histogram

Histogram (log scale)

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Additional map: Un-sharpenned map: GluA2 flip Q isoform in complex...

Fileemd_44245_additional_2.map
AnnotationUn-sharpenned map: GluA2 flip Q isoform in complex with TARPgamma2 at pH5.3, class23 structure of LBD-TMD-TARPgamma2
Projections & Slices
AxesZYX

Projections

Slices (1/2)
Density Histograms

Histogram

Histogram (log scale)

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Half map: Half1 map: GluA2 flip Q isoform in complex...

Fileemd_44245_half_map_1.map
AnnotationHalf1 map: GluA2 flip Q isoform in complex with TARPgamma2 at pH5.3, class23 structure of LBD-TMD-TARPgamma2
Projections & Slices
AxesZYX

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Density Histograms

Histogram

Histogram (log scale)

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Half map: Half2 map: GluA2 flip Q isoform in complex...

Fileemd_44245_half_map_2.map
AnnotationHalf2 map: GluA2 flip Q isoform in complex with TARPgamma2 at pH5.3, class23 structure of LBD-TMD-TARPgamma2
Projections & Slices
AxesZYX

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Slices (1/2)
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Histogram

Histogram (log scale)

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Sample components

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Entire : GluA2 (flip-Q isoform) in complex with TARPgamma2 at 4:4 stoichiometry

EntireName: GluA2 (flip-Q isoform) in complex with TARPgamma2 at 4:4 stoichiometry
Components
  • Complex: GluA2 (flip-Q isoform) in complex with TARPgamma2 at 4:4 stoichiometry
    • Protein or peptide: Voltage-dependent calcium channel gamma-2 subunit
    • Protein or peptide: Isoform Flip of Glutamate receptor 2

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Supramolecule #1: GluA2 (flip-Q isoform) in complex with TARPgamma2 at 4:4 stoichiometry

SupramoleculeName: GluA2 (flip-Q isoform) in complex with TARPgamma2 at 4:4 stoichiometry
type: complex / ID: 1 / Parent: 0 / Macromolecule list: all
Molecular weightTheoretical: 500 KDa

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Macromolecule #1: Voltage-dependent calcium channel gamma-2 subunit

MacromoleculeName: Voltage-dependent calcium channel gamma-2 subunit / type: protein_or_peptide / ID: 1 / Number of copies: 4 / Enantiomer: LEVO
Source (natural)Organism: Mus musculus (house mouse)
Molecular weightTheoretical: 35.938746 KDa
Recombinant expressionOrganism: Homo sapiens (human)
SequenceString: MGLFDRGVQM LLTTVGAFAA FSLMTIAVGT DYWLYSRGVC KTKSVSENET SKKNEEVMTH SGLWRTCCLE GNFKGLCKQI DHFPEDADY EADTAEYFLR AVRASSIFPI LSVILLFMGG LCIAASEFYK TRHNIILSAG IFFVSAGLSN IIGIIVYISA N AGDPSKSD ...String:
MGLFDRGVQM LLTTVGAFAA FSLMTIAVGT DYWLYSRGVC KTKSVSENET SKKNEEVMTH SGLWRTCCLE GNFKGLCKQI DHFPEDADY EADTAEYFLR AVRASSIFPI LSVILLFMGG LCIAASEFYK TRHNIILSAG IFFVSAGLSN IIGIIVYISA N AGDPSKSD SKKNSYSYGW SFYFGALSFI IAEMVGVLAV HMFIDRHKQL RATARATDYL QASAITRIPS YRYRYQRRSR SS SRSTEPS HSRDASPVGV KGFNTLPSTE ISMYTLSRDP LKAATTPTAT YNSDRDNSFL QVHNCIQKDS KDSLHANTAN RRT TPV

UniProtKB: Voltage-dependent calcium channel gamma-2 subunit

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Macromolecule #2: Isoform Flip of Glutamate receptor 2

MacromoleculeName: Isoform Flip of Glutamate receptor 2 / type: protein_or_peptide / ID: 2 / Details: A FLAG epitope tag is inserted near the C-terminus / Number of copies: 4 / Enantiomer: LEVO
Source (natural)Organism: Rattus norvegicus (Norway rat)
Molecular weightTheoretical: 99.617492 KDa
Recombinant expressionOrganism: Homo sapiens (human)
SequenceString: MQKIMHISVL LSPVLWGLIF GVSSNSIQIG GLFPRGADQE YSAFRVGMVQ FSTSEFRLTP HIDNLEVANS FAVTNAFCSQ FSRGVYAIF GFYDKKSVNT ITSFCGTLHV SFITPSFPTD GTHPFVIQMR PDLKGALLSL IEYYQWDKFA YLYDSDRGLS T LQAVLDSA ...String:
MQKIMHISVL LSPVLWGLIF GVSSNSIQIG GLFPRGADQE YSAFRVGMVQ FSTSEFRLTP HIDNLEVANS FAVTNAFCSQ FSRGVYAIF GFYDKKSVNT ITSFCGTLHV SFITPSFPTD GTHPFVIQMR PDLKGALLSL IEYYQWDKFA YLYDSDRGLS T LQAVLDSA AEKKWQVTAI NVGNINNDKK DETYRSLFQD LELKKERRVI LDCERDKVND IVDQVITIGK HVKGYHYIIA NL GFTDGDL LKIQFGGANV SGFQIVDYDD SLVSKFIERW STLEEKEYPG AHTATIKYTS ALTYDAVQVM TEAFRNLRKQ RIE ISRRGN AGDCLANPAV PWGQGVEIER ALKQVQVEGL SGNIKFDQNG KRINYTINIM ELKTNGPRKI GYWSEVDKMV VTLT ELPSG NDTSGLENKT VVVTTILESP YVMMKKNHEM LEGNERYEGY CVDLAAEIAK HCGFKYKLTI VGDGKYGARD ADTKI WNGM VGELVYGKAD IAIAPLTITL VREEVIDFSK PFMSLGISIM IKKPQKSKPG VFSFLDPLAY EIWMCIVFAY IGVSVV LFL VSRFSPYEWH TEEFEDGRET QSSESTNEFG IFNSLWFSLG AFMQQGCDIS PRSLSGRIVG GVWWFFTLII ISSYTAN LA AFLTVERMVS PIESAEDLSK QTEIAYGTLD SGSTKEFFRR SKIAVFDKMW TYMRSAEPSV FVRTTAEGVA RVRKSKGK Y AYLLESTMNE YIEQRKPCDT MKVGGNLDSK GYDIATPKGS SLGTPVNLAV LKLSEQGVLD KLKNKWWYDK GECGAKDSG SKEKTSALSL SNVAGVFYIL VGGLGLAMLV ALIEFCYKSR AEAKRMKVAK NPQNINPSSS QNSQNFATDY KDDDDKEGYN VYGIESVKI

UniProtKB: Glutamate receptor 2

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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

Concentration10 mg/mL
BufferpH: 8
Component:
ConcentrationFormulaName
150.0 mMNaClsodium chloride
20.0 mMTris
0.03 %GDN

Details: Tris adjusted to pH 8 using HCl. 4 micro litter of protein in buffer was mixed with 1 micro litter of 50mM citric acid buffer (the 50mM citric acid buffer was prepared by diluting 0.5M ...Details: Tris adjusted to pH 8 using HCl. 4 micro litter of protein in buffer was mixed with 1 micro litter of 50mM citric acid buffer (the 50mM citric acid buffer was prepared by diluting 0.5M citric acid/sodium citrate buffer at pH4.0) immediately before applying the sample to the grid.
GridModel: UltrAuFoil R1.2/1.3 / Material: GOLD / Mesh: 300 / Support film - Material: CARBON / Support film - topology: HOLEY ARRAY / Pretreatment - Type: GLOW DISCHARGE / Pretreatment - Time: 120 sec.
VitrificationCryogen name: ETHANE / Chamber humidity: 100 % / Chamber temperature: 277.15 K / Instrument: FEI VITROBOT MARK IV

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Electron microscopy

MicroscopeTFS KRIOS
Image recordingFilm or detector model: GATAN K3 BIOQUANTUM (6k x 4k) / Number grids imaged: 1 / Number real images: 19684 / Average electron dose: 55.6 e/Å2
Electron beamAcceleration voltage: 300 kV / Electron source: FIELD EMISSION GUN
Electron opticsIllumination mode: FLOOD BEAM / Imaging mode: BRIGHT FIELD / Nominal defocus max: 2.4 µm / Nominal defocus min: 1.0 µm
Sample stageCooling holder cryogen: NITROGEN
Experimental equipment
Model: Titan Krios / Image courtesy: FEI Company

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Image processing

Particle selectionDetails: Relion Autopick
Startup modelType of model: EMDB MAP
EMDB ID:

29386

Final reconstructionNumber classes used: 1 / Applied symmetry - Point group: C1 (asymmetric) / Algorithm: FOURIER SPACE / Resolution.type: BY AUTHOR / Resolution: 3.56 Å / Resolution method: FSC 0.143 CUT-OFF / Software - Name: RELION (ver. 4) / Number images used: 29945
Initial angle assignmentType: MAXIMUM LIKELIHOOD / Software - Name: RELION (ver. 4)
Final angle assignmentType: MAXIMUM LIKELIHOOD / Software - Name: RELION (ver. 4)
Final 3D classificationNumber classes: 40 / Software - Name: RELION (ver. 4)
FSC plot (resolution estimation)

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Atomic model buiding 1

Initial model
PDB IDChain

8fqf

source_name: PDB, initial_model_type: experimental model

8fqg

source_name: PDB, initial_model_type: experimental model
RefinementSpace: REAL
Output model

PDB-9b64:
GluA2 flip Q in complex with TARPgamma2 at pH5, class23, structure of LBD-TMD-TARPgamma2

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