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- EMDB-7817: Human GABA-A receptor alpha1-beta2-gamma2 subtype in complex with... -

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Basic information

Entry
Database: EMDB / ID: EMD-7817
TitleHuman GABA-A receptor alpha1-beta2-gamma2 subtype in complex with GABA and flumazenil, conformation A
Map data
SampleHuman GABA-A receptor alpha1-beta2-gamma2 subtype in complex with GABA and flumazenil, conformation A
  • (Gamma-aminobutyric acid receptor subunit ...) x 3
  • Kappa Fab Light Chain
  • IgG2b Fab Heavy Chain
  • (ligand) x 6
Function / homology
Function and homology information


benzodiazepine receptor activity / inner ear receptor cell development / inhibitory extracellular ligand-gated ion channel activity / GABA-gated chloride ion channel activity / GABA-A receptor complex / GABA-A receptor activity / GABA receptor complex / inhibitory synapse assembly / cellular response to histamine / synaptic transmission, GABAergic ...benzodiazepine receptor activity / inner ear receptor cell development / inhibitory extracellular ligand-gated ion channel activity / GABA-gated chloride ion channel activity / GABA-A receptor complex / GABA-A receptor activity / GABA receptor complex / inhibitory synapse assembly / cellular response to histamine / synaptic transmission, GABAergic / gamma-aminobutyric acid signaling pathway / regulation of postsynaptic membrane potential / integral component of postsynaptic specialization membrane / innervation / chloride channel activity / chloride channel complex / chloride transmembrane transport / adult behavior / dendrite membrane / cochlea development / GABA-ergic synapse / nervous system process / transmitter-gated ion channel activity involved in regulation of postsynaptic membrane potential / ion transmembrane transport / regulation of membrane potential / post-embryonic development / sensory perception of sound / cytoplasmic vesicle membrane / postsynapse / chemical synaptic transmission / postsynaptic membrane / drug binding / negative regulation of neuron apoptotic process / neuron projection / synapse / cell junction / axon / signal transduction / integral component of plasma membrane / extracellular exosome / plasma membrane / cytosol
Gamma-aminobutyric-acid A receptor, alpha 1 subunit / Neurotransmitter-gated ion-channel ligand-binding domain superfamily / Neurotransmitter-gated ion-channel transmembrane domain superfamily / Neurotransmitter-gated ion-channel, conserved site / Neurotransmitter-gated ion-channel ligand-binding domain / Neurotransmitter-gated ion-channel / Neurotransmitter-gated ion-channel transmembrane domain / Gamma-aminobutyric acid A receptor/Glycine receptor alpha / Gamma-aminobutyric-acid A receptor, gamma 2 subunit / Gamma-aminobutyric-acid A receptor, alpha subunit ...Gamma-aminobutyric-acid A receptor, alpha 1 subunit / Neurotransmitter-gated ion-channel ligand-binding domain superfamily / Neurotransmitter-gated ion-channel transmembrane domain superfamily / Neurotransmitter-gated ion-channel, conserved site / Neurotransmitter-gated ion-channel ligand-binding domain / Neurotransmitter-gated ion-channel / Neurotransmitter-gated ion-channel transmembrane domain / Gamma-aminobutyric acid A receptor/Glycine receptor alpha / Gamma-aminobutyric-acid A receptor, gamma 2 subunit / Gamma-aminobutyric-acid A receptor, alpha subunit / Gamma-aminobutyric-acid A receptor, gamma subunit / Gamma-aminobutyric-acid A receptor, beta subunit
Gamma-aminobutyric acid receptor subunit alpha-1 / Gamma-aminobutyric acid receptor subunit gamma-2 / Gamma-aminobutyric acid receptor subunit beta-2
Biological speciesHomo sapiens (human) / house mouse (house mouse)
Methodsingle particle reconstruction / cryo EM / Resolution: 3.92 Å
AuthorsZhu S / Noviello CM / Teng J / Walsh Jr RM / Kim JJ / Hibbs RE
Funding support United States, 1 items
OrganizationGrant numberCountry
Welch FoundationI-1812 United States
CitationJournal: Nature / Year: 2018
Title: Structure of a human synaptic GABA receptor.
Authors: Shaotong Zhu / Colleen M Noviello / Jinfeng Teng / Richard M Walsh / Jeong Joo Kim / Ryan E Hibbs /
Abstract: Fast inhibitory neurotransmission in the brain is principally mediated by the neurotransmitter GABA (γ-aminobutyric acid) and its synaptic target, the type A GABA receptor (GABA receptor). ...Fast inhibitory neurotransmission in the brain is principally mediated by the neurotransmitter GABA (γ-aminobutyric acid) and its synaptic target, the type A GABA receptor (GABA receptor). Dysfunction of this receptor results in neurological disorders and mental illnesses including epilepsy, anxiety and insomnia. The GABA receptor is also a prolific target for therapeutic, illicit and recreational drugs, including benzodiazepines, barbiturates, anaesthetics and ethanol. Here we present high-resolution cryo-electron microscopy structures of the human α1β2γ2 GABA receptor, the predominant isoform in the adult brain, in complex with GABA and the benzodiazepine site antagonist flumazenil, the first-line clinical treatment for benzodiazepine overdose. The receptor architecture reveals unique heteromeric interactions for this important class of inhibitory neurotransmitter receptor. This work provides a template for understanding receptor modulation by GABA and benzodiazepines, and will assist rational approaches to therapeutic targeting of this receptor for neurological disorders and mental illness.
Validation ReportPDB-ID: 6d6u

SummaryFull reportAbout validation report
History
DepositionApr 22, 2018-
Header (metadata) releaseJun 6, 2018-
Map releaseJun 27, 2018-
UpdateDec 18, 2019-
Current statusDec 18, 2019Processing site: RCSB / Status: Released

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Structure visualization

Movie
  • Surface view with section colored by density value
  • Surface level: 0.024
  • Imaged by UCSF Chimera
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  • Surface view colored by cylindrical radius
  • Surface level: 0.024
  • Imaged by UCSF Chimera
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  • Surface view with fitted model
  • Atomic models: : PDB-6d6u
  • Surface level: 0.024
  • Imaged by UCSF Chimera
  • Download
  • Simplified surface model + fitted atomic model
  • Atomic models: PDB-6d6u
  • Imaged by Jmol
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Movie viewer
Structure viewerEM map:
SurfViewMolmilJmol/JSmol
Supplemental images

Downloads & links

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Map

FileDownload / File: emd_7817.map.gz / Format: CCP4 / Size: 67 MB / Type: IMAGE STORED AS FLOATING POINT NUMBER (4 BYTES)
Projections & slices

Image control

Size
Brightness
Contrast
Others
AxesZ (Sec.)Y (Row.)X (Col.)
1.07 Å/pix.
x 260 pix.
= 278.2 Å
1.07 Å/pix.
x 260 pix.
= 278.2 Å
1.07 Å/pix.
x 260 pix.
= 278.2 Å

Surface

Projections

Slices (1/3)

Slices (1/2)

Slices (2/3)

Images are generated by Spider.

Voxel sizeX=Y=Z: 1.07 Å
Density
Contour LevelBy AUTHOR: 0.024 / Movie #1: 0.024
Minimum - Maximum-0.1750069 - 0.25294805
Average (Standard dev.)0.00041850033 (±0.006033654)
SymmetrySpace group: 1
Details

EMDB XML:

Map geometry
Axis orderXYZ
Origin000
Dimensions260260260
Spacing260260260
CellA=B=C: 278.2 Å
α=β=γ: 90.0 °

CCP4 map header:

modeImage stored as Reals
Å/pix. X/Y/Z1.071.071.07
M x/y/z260260260
origin x/y/z0.0000.0000.000
length x/y/z278.200278.200278.200
α/β/γ90.00090.00090.000
start NX/NY/NZ000
NX/NY/NZ384384384
MAP C/R/S123
start NC/NR/NS000
NC/NR/NS260260260
D min/max/mean-0.1750.2530.000

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Supplemental data

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Sample components

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Entire Human GABA-A receptor alpha1-beta2-gamma2 subtype in complex with...

EntireName: Human GABA-A receptor alpha1-beta2-gamma2 subtype in complex with GABA and flumazenil, conformation A
Details: Fab fragments generated by proteolytic cleavage of IgG antibody
Number of components: 12

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Component #1: protein, Human GABA-A receptor alpha1-beta2-gamma2 subtype in com...

ProteinName: Human GABA-A receptor alpha1-beta2-gamma2 subtype in complex with GABA and flumazenil, conformation A
Details: Fab fragments generated by proteolytic cleavage of IgG antibody
Recombinant expression: No
MassTheoretical: 348 kDa
SourceSpecies: Homo sapiens (human)

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Component #2: protein, Gamma-aminobutyric acid receptor subunit beta-2,Gamma-am...

ProteinName: Gamma-aminobutyric acid receptor subunit beta-2,Gamma-aminobutyric acid receptor subunit beta-2
Number of Copies: 2 / Recombinant expression: No
MassTheoretical: 39.521691 kDa
SourceSpecies: Homo sapiens (human)
Source (engineered)Expression System: Homo sapiens (human)

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Component #3: protein, Gamma-aminobutyric acid receptor subunit alpha-1,Gamma-a...

ProteinName: Gamma-aminobutyric acid receptor subunit alpha-1,Gamma-aminobutyric acid receptor subunit alpha-1
Number of Copies: 2 / Recombinant expression: No
MassTheoretical: 41.061211 kDa
SourceSpecies: Homo sapiens (human)
Source (engineered)Expression System: Homo sapiens (human)

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Component #4: protein, Gamma-aminobutyric acid receptor subunit gamma-2,Gamma-a...

ProteinName: Gamma-aminobutyric acid receptor subunit gamma-2,Gamma-aminobutyric acid receptor subunit gamma-2
Number of Copies: 1 / Recombinant expression: No
MassTheoretical: 45.35466 kDa
SourceSpecies: Homo sapiens (human)
Source (engineered)Expression System: Homo sapiens (human)

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Component #5: protein, Kappa Fab Light Chain

ProteinName: Kappa Fab Light Chain / Number of Copies: 2 / Recombinant expression: No
MassTheoretical: 23.505943 kDa
SourceSpecies: house mouse (house mouse) / Strain: BALB/c

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Component #6: protein, IgG2b Fab Heavy Chain

ProteinName: IgG2b Fab Heavy Chain / Number of Copies: 2 / Recombinant expression: No
MassTheoretical: 49.811043 kDa
SourceSpecies: house mouse (house mouse) / Strain: BALB/c

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Component #7: ligand, N-ACETYL-D-GLUCOSAMINE

LigandName: N-ACETYL-D-GLUCOSAMINEN-Acetylglucosamine / Number of Copies: 10 / Recombinant expression: No
MassTheoretical: 0.221208 kDa

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Component #8: ligand, BETA-D-MANNOSE

LigandName: BETA-D-MANNOSE / Number of Copies: 4 / Recombinant expression: No
MassTheoretical: 0.180156 kDa

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Component #9: ligand, GAMMA-AMINO-BUTANOIC ACID

LigandName: GAMMA-AMINO-BUTANOIC ACID / Number of Copies: 2 / Recombinant expression: No
MassTheoretical: 0.10312 kDa

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Component #10: ligand, CHOLESTEROL HEMISUCCINATE

LigandName: CHOLESTEROL HEMISUCCINATE / Number of Copies: 12 / Recombinant expression: No
MassTheoretical: 0.486726 kDa

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Component #11: ligand, ALPHA-D-MANNOSE

LigandName: ALPHA-D-MANNOSE / Number of Copies: 7 / Recombinant expression: No
MassTheoretical: 0.180156 kDa

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Component #12: ligand, ethyl 8-fluoro-5-methyl-6-oxo-5,6-dihydro-4H-imidazo[1,5-...

LigandName: ethyl 8-fluoro-5-methyl-6-oxo-5,6-dihydro-4H-imidazo[1,5-a][1,4]benzodiazepine-3-carboxylate
Number of Copies: 1 / Recombinant expression: No
MassTheoretical: 0.303288 kDa

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Experimental details

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Sample preparation

SpecimenSpecimen state: Particle / Method: cryo EM
Sample solutionSpecimen conc.: 6 mg/mL / pH: 7.4
Support filmSample was glow discharged at 30 mA for 80 seconds using a PELCO easiGLow
VitrificationInstrument: FEI VITROBOT MARK IV / Cryogen name: ETHANE / Temperature: 277 K / Humidity: 100 % / Details: 3 seconds blot time.

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Electron microscopy imaging

Experimental equipment
Model: Titan Krios / Image courtesy: FEI Company
ImagingMicroscope: FEI TITAN KRIOS
Electron gunElectron source: FIELD EMISSION GUN / Accelerating voltage: 300 kV / Electron dose: 47 e/Å2 / Illumination mode: FLOOD BEAM
LensMagnification: 46730.0 X (calibrated) / Cs: 2.7 mm / Imaging mode: BRIGHT FIELD / Defocus: 500.0 - 4500.0 nm / Energy filter: GIF Quantum LS / Energy window: -10-10 eV
Specimen HolderModel: FEI TITAN KRIOS AUTOGRID HOLDER / Temperature: (80.0 - 90.0 K)
CameraDetector: GATAN K2 SUMMIT (4k x 4k)

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Image acquisition

Image acquisitionNumber of digital images: 5594

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Image processing

ProcessingMethod: single particle reconstruction / Applied symmetry: C1 (asymmetric) / Number of projections: 292662
3D reconstructionAlgorithm: BACK PROJECTION / Software: RELION / Resolution: 3.92 Å / Resolution method: FSC 0.143 CUT-OFF
FSC plot (resolution estimation)

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Atomic model buiding

Modeling #1Refinement space: REAL
Output model

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