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- EMDB-22011: hEAAT3-IFS-Na -

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Basic information

Entry
Database: EMDB / ID: EMD-22011
TitlehEAAT3-IFS-Na
Map datainward facing human EAAT3
Sample
  • Complex: inward facing hEAAT3 trimer
    • Complex: inward facing hEAAT3 trimer
      • Protein or peptide: Excitatory amino acid transporter 3
Keywordshumant Excitatory amino acid transporter 3 / TRANSPORT PROTEIN
Function / homology
Function and homology information


D-aspartate transmembrane transport / regulation of protein targeting to membrane / D-aspartate transmembrane transporter activity / Defective SLC1A1 is implicated in schizophrenia 18 (SCZD18) and dicarboxylic aminoaciduria (DCBXA) / distal dendrite / cysteine transport / cysteine transmembrane transporter activity / neurotransmitter receptor transport to plasma membrane / high-affinity L-glutamate transmembrane transporter activity / glutamate:sodium symporter activity ...D-aspartate transmembrane transport / regulation of protein targeting to membrane / D-aspartate transmembrane transporter activity / Defective SLC1A1 is implicated in schizophrenia 18 (SCZD18) and dicarboxylic aminoaciduria (DCBXA) / distal dendrite / cysteine transport / cysteine transmembrane transporter activity / neurotransmitter receptor transport to plasma membrane / high-affinity L-glutamate transmembrane transporter activity / glutamate:sodium symporter activity / response to decreased oxygen levels / L-glutamate import / cellular response to mercury ion / Transport of inorganic cations/anions and amino acids/oligopeptides / retina layer formation / L-glutamate transmembrane transporter activity / cellular response to ammonium ion / L-glutamate transmembrane transport / glutathione biosynthetic process / D-aspartate import across plasma membrane / L-aspartate transmembrane transport / righting reflex / zinc ion transmembrane transport / cellular response to bisphenol A / L-aspartate transmembrane transporter activity / grooming behavior / intracellular glutamate homeostasis / L-aspartate import across plasma membrane / Glutamate Neurotransmitter Release Cycle / proximal dendrite / monoatomic anion channel activity / L-glutamate import across plasma membrane / apical dendrite / transepithelial transport / response to anesthetic / conditioned place preference / intracellular zinc ion homeostasis / cellular response to cocaine / chloride transmembrane transporter activity / blood vessel morphogenesis / motor neuron apoptotic process / response to morphine / motor behavior / G protein-coupled dopamine receptor signaling pathway / glutamate receptor signaling pathway / glutamate binding / neurotransmitter transport / superoxide metabolic process / heart contraction / maintenance of blood-brain barrier / perisynaptic space / dopamine metabolic process / adult behavior / : / asymmetric synapse / glial cell projection / response to axon injury / behavioral fear response / synaptic cleft / postsynaptic modulation of chemical synaptic transmission / transport across blood-brain barrier / positive regulation of heart rate / monoatomic ion transport / neurogenesis / axon terminus / response to amphetamine / chloride transmembrane transport / dendritic shaft / cell periphery / locomotory behavior / synapse organization / cytokine-mediated signaling pathway / brain development / Schaffer collateral - CA1 synapse / memory / long-term synaptic potentiation / recycling endosome membrane / late endosome membrane / presynapse / cellular response to oxidative stress / early endosome membrane / gene expression / chemical synaptic transmission / dendritic spine / perikaryon / negative regulation of neuron apoptotic process / apical plasma membrane / membrane raft / response to xenobiotic stimulus / axon / external side of plasma membrane / neuronal cell body / dendrite / endoplasmic reticulum / extracellular exosome / metal ion binding / identical protein binding / membrane / plasma membrane
Similarity search - Function
: / Sodium:dicarboxylate symporter family signature 2. / Sodium:dicarboxylate symporter / Sodium:dicarboxylate symporter, conserved site / Sodium:dicarboxylate symporter superfamily / Sodium:dicarboxylate symporter family / Sodium:dicarboxylate symporter family signature 1.
Similarity search - Domain/homology
Excitatory amino acid transporter 3
Similarity search - Component
Biological speciesHomo sapiens (human)
Methodsingle particle reconstruction / cryo EM / Resolution: 2.85 Å
AuthorsQiu B / Matthies D
Funding support United States, 1 items
OrganizationGrant numberCountry
National Institutes of Health/National Institute of Neurological Disorders and Stroke (NIH/NINDS)R01NS064357 United States
CitationJournal: Sci Adv / Year: 2021
Title: Cryo-EM structures of excitatory amino acid transporter 3 visualize coupled substrate, sodium, and proton binding and transport.
Authors: Biao Qiu / Doreen Matthies / Eva Fortea / Zhiheng Yu / Olga Boudker /
Abstract: Human excitatory amino acid transporter 3 (hEAAT3) mediates glutamate uptake in neurons, intestine, and kidney. Here, we report cryo-EM structures of hEAAT3 in several functional states where the ...Human excitatory amino acid transporter 3 (hEAAT3) mediates glutamate uptake in neurons, intestine, and kidney. Here, we report cryo-EM structures of hEAAT3 in several functional states where the transporter is empty, bound to coupled sodium ions only, or fully loaded with three sodium ions, a proton, and the substrate aspartate. The structures suggest that hEAAT3 operates by an elevator mechanism involving three functionally independent subunits. When the substrate-binding site is near the cytoplasm, it has a remarkably low affinity for the substrate, perhaps facilitating its release and allowing the rapid transport turnover. The mechanism of the coupled uptake of the sodium ions and the substrate is conserved across evolutionarily distant families and is augmented by coupling to protons in EAATs. The structures further suggest a mechanism by which a conserved glutamate residue mediates proton symport.
History
DepositionMay 20, 2020-
Header (metadata) releaseMar 17, 2021-
Map releaseMar 17, 2021-
UpdateJun 4, 2025-
Current statusJun 4, 2025Processing site: RCSB / Status: Released

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Structure visualization

Movie
  • Surface view with section colored by density value
  • Surface level: 0.025
  • Imaged by UCSF Chimera
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  • Surface view colored by cylindrical radius
  • Surface level: 0.025
  • Imaged by UCSF Chimera
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  • Surface view with fitted model
  • Atomic models: PDB-6x2l
  • Surface level: 0.025
  • Imaged by UCSF Chimera
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Movie viewer
Structure viewerEM map:
SurfViewMolmilJmol/JSmol
Supplemental images

emd_22011.png

Downloads & links

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Map

FileDownload / File: emd_22011.map.gz / Format: CCP4 / Size: 91.1 MB / Type: IMAGE STORED AS FLOATING POINT NUMBER (4 BYTES)
Annotationinward facing human EAAT3
Projections & slices

Image control

Size
Brightness
Contrast
Others
AxesZ (Sec.)Y (Row.)X (Col.)
0.83 Å/pix.
x 288 pix.
= 239.616 Å		0.83 Å/pix.
x 288 pix.
= 239.616 Å		0.83 Å/pix.
x 288 pix.
= 239.616 Å

Surface

Projections

Slices (1/3)

Slices (1/2)

Slices (2/3)

Images are generated by Spider.

Voxel sizeX=Y=Z: 0.832 Å
Density

Histogram

Histogram (log scale)
Contour LevelBy AUTHOR: 0.025 / Movie #1: 0.025
Minimum - Maximum-0.072760314 - 0.12378639
Average (Standard dev.)0.00029491424 (±0.0036509547)
SymmetrySpace group: 1
Details

EMDB XML:

Map geometry
Axis orderXYZ
Origin000
Dimensions288288288
Spacing288288288
CellA=B=C: 239.616 Å
α=β=γ: 90.0 °

CCP4 map header:

modeImage stored as Reals
Å/pix. X/Y/Z0.8320.8320.832
M x/y/z288288288
origin x/y/z0.0000.0000.000
length x/y/z239.616239.616239.616
α/β/γ90.00090.00090.000
MAP C/R/S123
start NC/NR/NS000
NC/NR/NS288288288
D min/max/mean-0.0730.1240.000

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Supplemental data

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Sample components

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Entire : inward facing hEAAT3 trimer

EntireName: inward facing hEAAT3 trimer
Components
  • Complex: inward facing hEAAT3 trimer
    • Complex: inward facing hEAAT3 trimer
      • Protein or peptide: Excitatory amino acid transporter 3

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Supramolecule #1: inward facing hEAAT3 trimer

SupramoleculeName: inward facing hEAAT3 trimer / type: complex / ID: 1 / Parent: 0 / Macromolecule list: all
Source (natural)Organism: Homo sapiens (human)

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Supramolecule #2: inward facing hEAAT3 trimer

SupramoleculeName: inward facing hEAAT3 trimer / type: complex / ID: 2 / Parent: 1 / Macromolecule list: all
Source (natural)Organism: Homo sapiens (human)

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Macromolecule #1: Excitatory amino acid transporter 3

MacromoleculeName: Excitatory amino acid transporter 3 / type: protein_or_peptide / ID: 1
Details: The Glycine and Proline at the N terminal are the residues left after PreScission Protease treatment
Number of copies: 3 / Enantiomer: LEVO
Source (natural)Organism: Homo sapiens (human)
Molecular weightTheoretical: 57.275168 KDa
Recombinant expressionOrganism: Homo sapiens (human)
SequenceString: GPMGKPARKG CEWKRFLKNN WVLLSTVAAV VLGITTGVLV REHSNLSTLE KFYFAFPGEI LMRMLKLIIL PLIISSMITG VAALDSNVS GKIGLRAVVY YFCTTLIAVI LGIVLVVSIK PGVTQKVGEI ARTGSTPEVS TVDAMLDLIR NMFPENLVQA C FQQYKTKR ...String:
GPMGKPARKG CEWKRFLKNN WVLLSTVAAV VLGITTGVLV REHSNLSTLE KFYFAFPGEI LMRMLKLIIL PLIISSMITG VAALDSNVS GKIGLRAVVY YFCTTLIAVI LGIVLVVSIK PGVTQKVGEI ARTGSTPEVS TVDAMLDLIR NMFPENLVQA C FQQYKTKR EEVKPPSDPE MTMTEESFTA VMTTAISKTK TKEYKIVGMY SDGINVLGLI VFCLVFGLVI GKMGEKGQIL VD FFNALSD ATMKIVQIIM CYMPLGILFL IAGKIIEVED WEIFRKLGLY MATVLTGLAI HSIVILPLIY FIVVRKNPFR FAM GMAQAL LTALMISSSS ATLPVTFRCA EENNQVDKRI TRFVLPVGAT INMDGTALYE AVAAVFIAQL NDLDLGIGQI ITIS ITATS ASIGAAGVPQ AGLVTMVIVL SAVGLPAEDV TLIIAVDWLL DRFRTMVNVL GDAFGTGIVE KLSKKELEQM DVSSE VNIV NPFALESTIL DNEDSDTKKS YVNGGFAVDK SDTISFTQTS QF

UniProtKB: Excitatory amino acid transporter 3

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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

Concentration6 mg/mL
BufferpH: 8
Component:
ConcentrationFormulaName
20.0 mMC4H11NO3Tris
200.0 mMNaClSodium Chloride
1.0 mMC4H7NO4L-Aspartate
1.0 mMC9H15O6PTCEP
0.086 mMC56H92O25Digitonin Glyco Diosgenin
GridModel: Quantifoil R1.2/1.3 / Material: COPPER / Mesh: 400 / Support film - Material: CARBON / Support film - topology: HOLEY / Pretreatment - Type: GLOW DISCHARGE / Pretreatment - Time: 60 sec. / Pretreatment - Atmosphere: AIR
VitrificationCryogen name: ETHANE / Chamber humidity: 100 % / Chamber temperature: 277 K / Instrument: FEI VITROBOT MARK IV / Details: blot 3s.
DetailsThis sample was mono disperse

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Electron microscopy

MicroscopeFEI TITAN KRIOS
Image recordingFilm or detector model: GATAN K3 BIOQUANTUM (6k x 4k) / Average electron dose: 60.0 e/Å2
Electron beamAcceleration voltage: 300 kV / Electron source: FIELD EMISSION GUN
Electron opticsC2 aperture diameter: 100.0 µm / Illumination mode: FLOOD BEAM / Imaging mode: BRIGHT FIELD / Cs: 2.7 mm / Nominal defocus max: 1.5 µm / Nominal defocus min: 0.5 µm / Nominal magnification: 105000
Sample stageSpecimen holder model: FEI TITAN KRIOS AUTOGRID HOLDER / Cooling holder cryogen: NITROGEN
Experimental equipment
Model: Titan Krios / Image courtesy: FEI Company

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Image processing

Particle selectionNumber selected: 1717667
CTF correctionSoftware - Name: RELION (ver. 3.0) / Type: PHASE FLIPPING AND AMPLITUDE CORRECTION
Startup modelType of model: OTHER / Details: ab-initio
Final reconstructionNumber classes used: 1 / Applied symmetry - Point group: C3 (3 fold cyclic) / Resolution.type: BY AUTHOR / Resolution: 2.85 Å / Resolution method: FSC 0.143 CUT-OFF / Software - Name: RELION (ver. 3.0) / Number images used: 554920
Initial angle assignmentType: MAXIMUM LIKELIHOOD / Software - Name: RELION (ver. 3.0)
Final angle assignmentType: MAXIMUM LIKELIHOOD / Software - Name: RELION (ver. 3.0)
Final 3D classificationNumber classes: 8
FSC plot (resolution estimation)

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