[English] 日本語
Yorodumi
- PDB-7awp: Structure of the thermostabilized EAAT1 cryst-II mutant in comple... -

+
Open data


ID or keywords:

Loading...

-
Basic information

Entry
Database: PDB / ID: 7awp
TitleStructure of the thermostabilized EAAT1 cryst-II mutant in complex with rubidium and barium ions and the allosteric inhibitor UCPH101
ComponentsExcitatory amino acid transporter 1,Neutral amino acid transporter B(0),Excitatory amino acid transporter 1
KeywordsTRANSPORT PROTEIN / excitatory amino acid transporter 1 / human glutamate transporter / SLC1A3 / ion-coupling mechanism
Function / homology
Function and homology information


Defective SLC1A3 causes episodic ataxia 6 (EA6) / Astrocytic Glutamate-Glutamine Uptake And Metabolism / cranial nerve development / glutamine secretion / gamma-aminobutyric acid biosynthetic process / neurotransmitter uptake / L-glutamine import across plasma membrane / cell morphogenesis involved in neuron differentiation / L-glutamine transmembrane transporter activity / glutamine transport ...Defective SLC1A3 causes episodic ataxia 6 (EA6) / Astrocytic Glutamate-Glutamine Uptake And Metabolism / cranial nerve development / glutamine secretion / gamma-aminobutyric acid biosynthetic process / neurotransmitter uptake / L-glutamine import across plasma membrane / cell morphogenesis involved in neuron differentiation / L-glutamine transmembrane transporter activity / glutamine transport / L-serine transmembrane transporter activity / membrane protein complex / high-affinity L-glutamate transmembrane transporter activity / glutamate:sodium symporter activity / L-glutamate import / : / L-glutamate transmembrane transporter activity / L-glutamate transmembrane transport / D-aspartate import across plasma membrane / ligand-gated channel activity / neutral amino acid transport / L-aspartate transmembrane transporter activity / L-aspartate import across plasma membrane / Glutamate Neurotransmitter Release Cycle / neutral L-amino acid transmembrane transporter activity / auditory behavior / L-glutamate import across plasma membrane / symporter activity / transepithelial transport / Amino acid transport across the plasma membrane / amino acid transmembrane transporter activity / intracellular sodium ion homeostasis / cellular response to cocaine / glutamate binding / antiporter activity / neuromuscular process controlling balance / neurotransmitter transport / RHOJ GTPase cycle / protein homotrimerization / RHOQ GTPase cycle / amino acid transport / response to light stimulus / RHOH GTPase cycle / RAC3 GTPase cycle / positive regulation of synaptic transmission / transport across blood-brain barrier / monoatomic ion transport / RAC1 GTPase cycle / potassium ion transmembrane transport / chloride transmembrane transport / basal plasma membrane / erythrocyte differentiation / sensory perception of sound / response to wounding / centriolar satellite / melanosome / signaling receptor activity / virus receptor activity / cytoplasmic vesicle / chemical synaptic transmission / neuron projection / ciliary basal body / response to xenobiotic stimulus / response to antibiotic / neuronal cell body / synapse / perinuclear region of cytoplasm / cell surface / extracellular exosome / metal ion binding / membrane / plasma membrane
Similarity search - Function
Proton glutamate symport protein / Sodium:dicarboxylate symporter / : / 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. / Orthogonal Bundle / Mainly Alpha
Similarity search - Domain/homology
Chem-6Z6 / : / RUBIDIUM ION / Excitatory amino acid transporter 1 / Neutral amino acid transporter B(0)
Similarity search - Component
Biological speciesHomo sapiens (human)
MethodX-RAY DIFFRACTION / SYNCHROTRON / MOLECULAR REPLACEMENT / molecular replacement / Resolution: 3.91 Å
AuthorsCanul-Tec, J.C. / Legrand, P. / Reyes, N.
Funding support France, 1items
OrganizationGrant numberCountry
European Research Council (ERC)309657 France
CitationJournal: EMBO J / Year: 2022
Title: The ion-coupling mechanism of human excitatory amino acid transporters.
Authors: Juan C Canul-Tec / Anand Kumar / Jonathan Dhenin / Reda Assal / Pierre Legrand / Martial Rey / Julia Chamot-Rooke / Nicolas Reyes /
Abstract: Excitatory amino acid transporters (EAATs) maintain glutamate gradients in the brain essential for neurotransmission and to prevent neuronal death. They use ionic gradients as energy source and co- ...Excitatory amino acid transporters (EAATs) maintain glutamate gradients in the brain essential for neurotransmission and to prevent neuronal death. They use ionic gradients as energy source and co-transport transmitter into the cytoplasm with Na and H , while counter-transporting K to re-initiate the transport cycle. However, the molecular mechanisms underlying ion-coupled transport remain incompletely understood. Here, we present 3D X-ray crystallographic and cryo-EM structures, as well as thermodynamic analysis of human EAAT1 in different ion bound conformations, including elusive counter-transport ion bound states. Binding energies of Na and H , and unexpectedly Ca , are coupled to neurotransmitter binding. Ca competes for a conserved Na site, suggesting a regulatory role for Ca in glutamate transport at the synapse, while H binds to a conserved glutamate residue stabilizing substrate occlusion. The counter-transported ion binding site overlaps with that of glutamate, revealing the K -based mechanism to exclude the transmitter during the transport cycle and to prevent its neurotoxic release on the extracellular side.
History
DepositionNov 8, 2020Deposition site: PDBE / Processing site: PDBE
Revision 1.0Oct 13, 2021Provider: repository / Type: Initial release
Revision 1.1Dec 1, 2021Group: Database references / Category: citation / citation_author
Item: _citation.country / _citation.journal_abbrev ..._citation.country / _citation.journal_abbrev / _citation.journal_id_ASTM / _citation.journal_id_CSD / _citation.journal_id_ISSN / _citation.page_first / _citation.page_last / _citation.pdbx_database_id_DOI / _citation.pdbx_database_id_PubMed / _citation.title / _citation.year
Revision 1.2Jan 12, 2022Group: Database references / Category: citation / citation_author
Item: _citation.journal_volume / _citation.page_first ..._citation.journal_volume / _citation.page_first / _citation.page_last / _citation.year / _citation_author.identifier_ORCID
Revision 1.3Jan 19, 2022Group: Database references / Category: citation / citation_author
Item: _citation.page_first / _citation.page_last / _citation_author.identifier_ORCID
Revision 1.4Jan 31, 2024Group: Data collection / Refinement description
Category: chem_comp_atom / chem_comp_bond / pdbx_initial_refinement_model

-
Structure visualization

Structure viewerMolecule:
MolmilJmol/JSmol

Downloads & links

-
Assembly

Deposited unit
A: Excitatory amino acid transporter 1,Neutral amino acid transporter B(0),Excitatory amino acid transporter 1
hetero molecules


Theoretical massNumber of molelcules
Total (without water)57,2435
Polymers56,4601
Non-polymers7834
Water00
1
A: Excitatory amino acid transporter 1,Neutral amino acid transporter B(0),Excitatory amino acid transporter 1
hetero molecules

A: Excitatory amino acid transporter 1,Neutral amino acid transporter B(0),Excitatory amino acid transporter 1
hetero molecules

A: Excitatory amino acid transporter 1,Neutral amino acid transporter B(0),Excitatory amino acid transporter 1
hetero molecules


Theoretical massNumber of molelcules
Total (without water)171,72915
Polymers169,3813
Non-polymers2,34812
Water0
TypeNameSymmetry operationNumber
identity operation1_555x,y,z1
crystal symmetry operation2_565-y,x-y+1,z1
crystal symmetry operation3_455-x+y-1,-x,z1
Buried area10360 Å2
ΔGint-144 kcal/mol
Surface area48960 Å2
MethodPISA
Unit cell
Length a, b, c (Å)124.375, 124.375, 91.438
Angle α, β, γ (deg.)90.000, 90.000, 120.000
Int Tables number173
Space group name H-MP63

-
Components

#1: Protein Excitatory amino acid transporter 1,Neutral amino acid transporter B(0),Excitatory amino acid transporter 1 / Sodium-dependent glutamate/aspartate transporter 1 / GLAST-1 / Solute carrier family 1 member 3 / ...Sodium-dependent glutamate/aspartate transporter 1 / GLAST-1 / Solute carrier family 1 member 3 / ATB(0) / Baboon M7 virus receptor / RD114/simian type D retrovirus receptor / Sodium-dependent neutral amino acid transporter type 2 / Solute carrier family 1 member 5 / Sodium-dependent glutamate/aspartate transporter 1 / GLAST-1 / Solute carrier family 1 member 3


Mass: 56460.324 Da / Num. of mol.: 1
Source method: isolated from a genetically manipulated source
Details: Rubidium and Barium ions and allosteric inhibitor UCPH101,Rubidium and Barium ions and allosteric inhibitor UCPH101,Rubidium and Barium ions and allosteric inhibitor UCPH101
Source: (gene. exp.) Homo sapiens (human)
Gene: SLC1A3, EAAT1, GLAST, GLAST1, SLC1A5, ASCT2, M7V1, RDR, RDRC
Plasmid: pcDNA3.1 / Cell (production host): embrionic / Cell line (production host): HEK-293F / Production host: Homo sapiens (human) / Tissue (production host): embrionic kidney / References: UniProt: P43003, UniProt: Q15758
#2: Chemical ChemComp-6Z6 / 2-Amino-5,6,7,8-tetrahydro-4-(4-methoxyphenyl)-7-(naphthalen-1-yl)-5-oxo-4H-chromene-3-carbonitrile


Mass: 422.475 Da / Num. of mol.: 1 / Source method: obtained synthetically / Formula: C27H22N2O3 / Feature type: SUBJECT OF INVESTIGATION
#3: Chemical ChemComp-BA / BARIUM ION


Mass: 137.327 Da / Num. of mol.: 2 / Source method: obtained synthetically / Formula: Ba
#4: Chemical ChemComp-RB / RUBIDIUM ION


Mass: 85.468 Da / Num. of mol.: 1 / Source method: obtained synthetically / Formula: Rb
Has ligand of interestY

-
Experimental details

-
Experiment

ExperimentMethod: X-RAY DIFFRACTION / Number of used crystals: 1

-
Sample preparation

CrystalDensity Matthews: 3.62 Å3/Da / Density % sol: 65.98 %
Crystal growTemperature: 277 K / Method: vapor diffusion, hanging drop / pH: 8.2
Details: 28% PEG400, 100 mM Tris pH8.2, 50 mM Calcium chloride, 50 mM Barium chloride
PH range: 8-8.4

-
Data collection

DiffractionMean temperature: 100 K / Serial crystal experiment: N
Diffraction sourceSource: SYNCHROTRON / Site: SOLEIL / Beamline: PROXIMA 1 / Wavelength: 0.979 Å
DetectorType: DECTRIS PILATUS 6M / Detector: PIXEL / Date: Sep 2, 2016
RadiationMonochromator: channel cut monochromator / Protocol: SINGLE WAVELENGTH / Monochromatic (M) / Laue (L): M / Scattering type: x-ray
Radiation wavelengthWavelength: 0.979 Å / Relative weight: 1
ReflectionResolution: 3.91→46.41 Å / Num. obs: 7403 / % possible obs: 99.8 % / Redundancy: 16.7 % / Biso Wilson estimate: 142.47 Å2 / CC1/2: 1 / Rmerge(I) obs: 0.126 / Rpim(I) all: 0.032 / Rrim(I) all: 0.13 / Net I/σ(I): 9 / Num. measured all: 123824
Reflection shell

Diffraction-ID: 1

Resolution (Å)Redundancy (%)Rmerge(I) obsNum. measured allNum. unique obsCC1/2Rpim(I) allRrim(I) allNet I/σ(I) obs% possible all
3.91-4.0114.69.76780355490.1482.60110.1280.399.1
17.47-46.41130.03311488810.010.03458.495.1

-
Phasing

PhasingMethod: molecular replacement
Phasing MRModel details: Phaser MODE: MR_AUTO
Highest resolutionLowest resolution
Rotation3.91 Å46.41 Å
Translation3.91 Å46.41 Å

-
Processing

Software
NameVersionClassification
XDSJan 31, 2020 Built=20200417data reduction
Aimless0.5.32data scaling
PHASER2.7.17phasing
BUSTER2.10.3refinement
PDB_EXTRACT3.27data extraction
RefinementMethod to determine structure: MOLECULAR REPLACEMENT
Starting model: 5LM4

5lm4


Resolution: 3.91→23 Å / Cor.coef. Fo:Fc: 0.85 / Cor.coef. Fo:Fc free: 0.835 / Cross valid method: THROUGHOUT / σ(F): 0 / SU Rfree Blow DPI: 1.016
RfactorNum. reflection% reflectionSelection details
Rfree0.291 228 4.92 %RANDOM
Rwork0.246 ---
obs0.248 4632 63 %-
Displacement parametersBiso max: 270.4 Å2 / Biso mean: 149.42 Å2 / Biso min: 17.78 Å2
Baniso -1Baniso -2Baniso -3
1--4.5403 Å20 Å20 Å2
2---4.5403 Å20 Å2
3---9.0806 Å2
Refine analyzeLuzzati coordinate error obs: 0.68 Å
Refinement stepCycle: final / Resolution: 3.91→23 Å
ProteinNucleic acidLigandSolventTotal
Num. atoms2933 0 35 0 2968
Biso mean--152.24 --
Num. residues----386
Refine LS restraints
Refine-IDTypeNumberRestraint functionWeightDev ideal
X-RAY DIFFRACTIONt_dihedral_angle_d1028SINUSOIDAL2
X-RAY DIFFRACTIONt_trig_c_planes44HARMONIC2
X-RAY DIFFRACTIONt_gen_planes464HARMONIC5
X-RAY DIFFRACTIONt_it3014HARMONIC20
X-RAY DIFFRACTIONt_nbd3SEMIHARMONIC5
X-RAY DIFFRACTIONt_improper_torsion
X-RAY DIFFRACTIONt_pseud_angle
X-RAY DIFFRACTIONt_chiral_improper_torsion425SEMIHARMONIC5
X-RAY DIFFRACTIONt_sum_occupancies
X-RAY DIFFRACTIONt_utility_distance
X-RAY DIFFRACTIONt_utility_angle
X-RAY DIFFRACTIONt_utility_torsion
X-RAY DIFFRACTIONt_ideal_dist_contact3668SEMIHARMONIC4
X-RAY DIFFRACTIONt_bond_d3014HARMONIC20.009
X-RAY DIFFRACTIONt_angle_deg4098HARMONIC21
X-RAY DIFFRACTIONt_omega_torsion1.69
X-RAY DIFFRACTIONt_other_torsion21.14
LS refinement shellResolution: 3.91→4.37 Å / Rfactor Rfree error: 0 / Total num. of bins used: 5
RfactorNum. reflection% reflection
Rfree0.2955 28 4.66 %
Rwork0.2104 573 -
all0.2137 601 -
obs--28.8 %
Refinement TLS params.

Method: refined / Refine-ID: X-RAY DIFFRACTION

IDL11 (°2)L12 (°2)L13 (°2)L22 (°2)L23 (°2)L33 (°2)S11 (Å °)S12 (Å °)S13 (Å °)S21 (Å °)S22 (Å °)S23 (Å °)S31 (Å °)S32 (Å °)S33 (Å °)T11 (Å2)T12 (Å2)T13 (Å2)T22 (Å2)T23 (Å2)T33 (Å2)Origin x (Å)Origin y (Å)Origin z (Å)
1-0.7440.4295-0.01650.744-0.67510-0.2578-0.1237-0.13440.0197-0.01150.27480.1491-0.14140.2692-0.08730.04490.00440.0933-0.1268-0.2341-45.086637.47110.321
2-0.3293-0.12290.76521.41370.34010.0454-0.12410.15030.24790.1249-0.36020.31510.13760.0490.4842-0.57850.09280.01150.3057-0.0297-0.3046-33.140226.24390.513
Refinement TLS group
IDRefine-IDRefine TLS-IDSelection detailsAuth asym-IDAuth seq-ID
1X-RAY DIFFRACTION1{A|42 - 111 A|170 - 283}A42 - 111
2X-RAY DIFFRACTION1{A|42 - 111 A|170 - 283}A170 - 283
3X-RAY DIFFRACTION2{A|112 - 147 A|290 - 486}A112 - 147
4X-RAY DIFFRACTION2{A|112 - 147 A|290 - 486}A290 - 486

+
About Yorodumi

-
News

-
Feb 9, 2022. New format data for meta-information of EMDB entries

New format data for meta-information of EMDB entries

  • Version 3 of the EMDB header file is now the official format.
  • The previous official version 1.9 will be removed from the archive.

Related info.:EMDB header

External links:wwPDB to switch to version 3 of the EMDB data model

-
Aug 12, 2020. Covid-19 info

Covid-19 info

URL: https://pdbj.org/emnavi/covid19.php

New page: Covid-19 featured information page in EM Navigator.

Related info.:Covid-19 info / Mar 5, 2020. Novel coronavirus structure data

+
Mar 5, 2020. Novel coronavirus structure data

Novel coronavirus structure data

Related info.:Yorodumi Speices / Aug 12, 2020. Covid-19 info

External links:COVID-19 featured content - PDBj / Molecule of the Month (242)：Coronavirus Proteases

+
Jan 31, 2019. EMDB accession codes are about to change! (news from PDBe EMDB page)

EMDB accession codes are about to change! (news from PDBe EMDB page)

  • The allocation of 4 digits for EMDB accession codes will soon come to an end. Whilst these codes will remain in use, new EMDB accession codes will include an additional digit and will expand incrementally as the available range of codes is exhausted. The current 4-digit format prefixed with “EMD-” (i.e. EMD-XXXX) will advance to a 5-digit format (i.e. EMD-XXXXX), and so on. It is currently estimated that the 4-digit codes will be depleted around Spring 2019, at which point the 5-digit format will come into force.
  • The EM Navigator/Yorodumi systems omit the EMD- prefix.

Related info.:Q: What is EMD? / ID/Accession-code notation in Yorodumi/EM Navigator

External links:EMDB Accession Codes are Changing Soon! / Contact to PDBj

+
Jul 12, 2017. Major update of PDB

Major update of PDB

  • wwPDB released updated PDB data conforming to the new PDBx/mmCIF dictionary.
  • This is a major update changing the version number from 4 to 5, and with Remediation, in which all the entries are updated.
  • In this update, many items about electron microscopy experimental information are reorganized (e.g. em_software).
  • Now, EM Navigator and Yorodumi are based on the updated data.

External links:wwPDB Remediation / Enriched Model Files Conforming to OneDep Data Standards Now Available in the PDB FTP Archive

-
Yorodumi

Thousand views of thousand structures

  • Yorodumi is a browser for structure data from EMDB, PDB, SASBDB, etc.
  • This page is also the successor to EM Navigator detail page, and also detail information page/front-end page for Omokage search.
  • The word "yorodu" (or yorozu) is an old Japanese word meaning "ten thousand". "mi" (miru) is to see.

Related info.:EMDB / PDB / SASBDB / Comparison of 3 databanks / Yorodumi Search / Aug 31, 2016. New EM Navigator & Yorodumi / Yorodumi Papers / Jmol/JSmol / Function and homology information / Changes in new EM Navigator and Yorodumi

Read more