[English] 日本語
Yorodumi
- EMDB-0280: CryoEM structure of human full-length alpha1beta3gamma2L GABA(A)R... -

+
Open data


ID or keywords:

Loading...

-
Basic information

Entry
Database: EMDB / ID: EMD-0280
TitleCryoEM structure of human full-length alpha1beta3gamma2L GABA(A)R in complex with bicuculline and megabody Mb38.
Map data
SampleHuman full-length heteromeric alpha1beta3gamma2L GABA(A)R in complex with bicuculline and megabody Mb38.
  • Human full-length heteromeric alpha1beta3gamma2L GABA(A)R
  • megabody Mb38
  • (Gamma-aminobutyric acid receptor subunit ...) x 3
  • Megabody Mb38
  • (ligand) x 5
Function / homology
Function and homology information


benzodiazepine receptor activity / inner ear receptor cell development / inhibitory extracellular ligand-gated ion channel activity / GABA-A receptor complex / GABA-A receptor activity / GABA-gated chloride ion channel 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-A receptor complex / GABA-A receptor activity / GABA-gated chloride ion channel activity / GABA receptor complex / inhibitory synapse assembly / cellular response to histamine / synaptic transmission, GABAergic / gamma-aminobutyric acid signaling pathway / integral component of postsynaptic specialization membrane / innervation / regulation of postsynaptic membrane potential / chloride channel activity / inhibitory postsynaptic potential / chloride channel complex / chloride transmembrane transport / adult behavior / dendrite membrane / cochlea development / GABA-ergic synapse / nervous system process / roof of mouth development / transmitter-gated ion channel activity involved in regulation of postsynaptic membrane potential / ion transmembrane transport / post-embryonic development / regulation of membrane potential / sensory perception of sound / cytoplasmic vesicle membrane / postsynapse / postsynaptic membrane / chemical synaptic transmission / drug binding / negative regulation of neuron apoptotic process / neuron projection / synapse / cell junction / axon / signal transduction / integral component of plasma membrane / identical protein binding / plasma membrane
Gamma-aminobutyric-acid A receptor, alpha 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, gamma subunit ...Gamma-aminobutyric-acid A receptor, alpha 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, gamma subunit / Gamma-aminobutyric-acid A receptor, alpha 1 subunit / Gamma-aminobutyric-acid A receptor, beta subunit
Gamma-aminobutyric acid receptor subunit alpha-1 / Gamma-aminobutyric acid receptor subunit alpha-1 / Gamma-aminobutyric acid receptor subunit gamma-2 / Gamma-aminobutyric acid receptor subunit beta-3
Biological speciesHomo sapiens (human) / Lama glama (llama)
Methodsingle particle reconstruction / cryo EM / Resolution: 3.69 Å
AuthorsMasiulis S / Desai R / Uchanski T / Serna Martin I / Laverty D / Karia D / Malinauskas T / Jasenko Z / Pardon E / Kotecha A / Steyaert J / Miller KW / Aricescu AR
Funding support United Kingdom, Switzerland, United States, 8 items
OrganizationGrant numberCountry
Medical Research Council (United Kingdom)MR/L009609/1 United Kingdom
Medical Research Council (United Kingdom)MC_UP_A025_1013 United Kingdom
Medical Research Council (United Kingdom)MC_UP_1201/15 United Kingdom
Cancer Research UKC20724/A14414 United Kingdom
Biotechnology and Biological Sciences Research CouncilBB/M024709/1 United Kingdom
Swiss National Science Foundation168735 Switzerland
Human Frontier Science ProgramRGP0065/2014 United Kingdom
National Institutes of Health/National Institute of General Medical SciencesGM 58448 United States
CitationJournal: Nature / Year: 2019
Title: GABA receptor signalling mechanisms revealed by structural pharmacology.
Authors: Simonas Masiulis / Rooma Desai / Tomasz Uchański / Itziar Serna Martin / Duncan Laverty / Dimple Karia / Tomas Malinauskas / Jasenko Zivanov / Els Pardon / Abhay Kotecha / Jan Steyaert / ...Authors: Simonas Masiulis / Rooma Desai / Tomasz Uchański / Itziar Serna Martin / Duncan Laverty / Dimple Karia / Tomas Malinauskas / Jasenko Zivanov / Els Pardon / Abhay Kotecha / Jan Steyaert / Keith W Miller / A Radu Aricescu /
Abstract: Type-A γ-aminobutyric (GABA) receptors are ligand-gated chloride channels with a very rich pharmacology. Some of their modulators, including benzodiazepines and general anaesthetics, are among the ...Type-A γ-aminobutyric (GABA) receptors are ligand-gated chloride channels with a very rich pharmacology. Some of their modulators, including benzodiazepines and general anaesthetics, are among the most successful drugs in clinical use and are common substances of abuse. Without reliable structural data, the mechanistic basis for the pharmacological modulation of GABA receptors remains largely unknown. Here we report several high-resolution cryo-electron microscopy structures in which the full-length human α1β3γ2L GABA receptor in lipid nanodiscs is bound to the channel-blocker picrotoxin, the competitive antagonist bicuculline, the agonist GABA (γ-aminobutyric acid), and the classical benzodiazepines alprazolam and diazepam. We describe the binding modes and mechanistic effects of these ligands, the closed and desensitized states of the GABA receptor gating cycle, and the basis for allosteric coupling between the extracellular, agonist-binding region and the transmembrane, pore-forming region. This work provides a structural framework in which to integrate previous physiology and pharmacology research and a rational basis for the development of GABA receptor modulators.
History
DepositionOct 8, 2018-
Header (metadata) releaseNov 7, 2018-
Map releaseJan 2, 2019-
UpdateDec 11, 2019-
Current statusDec 11, 2019Processing site: PDBe / Status: Released

-
Structure visualization

Movie
  • Surface view with section colored by density value
  • Surface level: 0.07
  • Imaged by UCSF Chimera
  • Download
  • Surface view colored by cylindrical radius
  • Surface level: 0.07
  • Imaged by UCSF Chimera
  • Download
  • Surface view with fitted model
  • Atomic models: : PDB-6huk
  • Surface level: 0.07
  • Imaged by UCSF Chimera
  • Download
Movie viewer
Structure viewerEM map:
SurfViewMolmilJmol/JSmol
Supplemental images

Downloads & links

-
Map

FileDownload / File: emd_0280.map.gz / Format: CCP4 / Size: 42.9 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 224 pix.
= 239.68 Å
1.07 Å/pix.
x 224 pix.
= 239.68 Å
1.07 Å/pix.
x 224 pix.
= 239.68 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: 0.07 / Movie #1: 0.07
Minimum - Maximum-0.32086563 - 0.4944583
Average (Standard dev.)0.0012150024 (±0.015643317)
SymmetrySpace group: 1
Details

EMDB XML:

Map geometry
Axis orderXYZ
Origin000
Dimensions224224224
Spacing224224224
CellA=B=C: 239.68001 Å
α=β=γ: 90.0 °

CCP4 map header:

modeImage stored as Reals
Å/pix. X/Y/Z1.071.071.07
M x/y/z224224224
origin x/y/z0.0000.0000.000
length x/y/z239.680239.680239.680
α/β/γ90.00090.00090.000
MAP C/R/S123
start NC/NR/NS000
NC/NR/NS224224224
D min/max/mean-0.3210.4940.001

-
Supplemental data

-
Additional map: Un-sharpened combined map

Fileemd_0280_additional.map
AnnotationUn-sharpened combined map
Projections & Slices
AxesZYX

Projections

Slices (1/2)
Density Histograms

-
Half map: Half-map 1

Fileemd_0280_half_map_1.map
AnnotationHalf-map 1
Projections & Slices
AxesZYX

Projections

Slices (1/2)
Density Histograms

-
Half map: Half-map 2

Fileemd_0280_half_map_2.map
AnnotationHalf-map 2
Projections & Slices
AxesZYX

Projections

Slices (1/2)
Density Histograms

-
Sample components

+
Entire Human full-length heteromeric alpha1beta3gamma2L GABA(A)R in comp...

EntireName: Human full-length heteromeric alpha1beta3gamma2L GABA(A)R in complex with bicuculline and megabody Mb38.
Number of components: 12

+
Component #1: protein, Human full-length heteromeric alpha1beta3gamma2L GABA(A)...

ProteinName: Human full-length heteromeric alpha1beta3gamma2L GABA(A)R in complex with bicuculline and megabody Mb38.
Recombinant expression: No
MassTheoretical: 330 kDa

+
Component #2: protein, Human full-length heteromeric alpha1beta3gamma2L GABA(A)R

ProteinName: Human full-length heteromeric alpha1beta3gamma2L GABA(A)R
Recombinant expression: No
SourceSpecies: Homo sapiens (human)
Source (engineered)Expression System: Homo sapiens (human) / Cell of expression system: HEK293S
Source (natural)Location in cell: Plasma membrane / Organ or tissue: Brain

+
Component #3: protein, megabody Mb38

ProteinName: megabody Mb38 / Recombinant expression: No
SourceSpecies: Lama glama (llama)
Source (engineered)Expression System: Esherichia coli

+
Component #4: 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
Details: Potential signal peptide: MKKSPGLSDY LWAWTLFLST LTGRSYG FLAG tag: DYKDDDDK,Potential signal peptide: MKKSPGLSDY LWAWTLFLST LTGRSYG FLAG tag: DYKDDDDK
Number of Copies: 2 / Recombinant expression: No
MassTheoretical: 52.916602 kDa
SourceSpecies: Homo sapiens (human)
Source (engineered)Expression System: Homo sapiens (human)

+
Component #5: protein, Gamma-aminobutyric acid receptor subunit beta-3

ProteinName: Gamma-aminobutyric acid receptor subunit beta-3 / Number of Copies: 2 / Recombinant expression: No
MassTheoretical: 54.444578 kDa
SourceSpecies: Homo sapiens (human)
Source (engineered)Expression System: Homo sapiens (human)

+
Component #6: protein, Gamma-aminobutyric acid receptor subunit gamma-2

ProteinName: Gamma-aminobutyric acid receptor subunit gamma-2 / Details: Linker sequence: GGSGGSGGSGK 1D4 tag: TETSQVAPA / Number of Copies: 1 / Recombinant expression: No
MassTheoretical: 56.922055 kDa
SourceSpecies: Homo sapiens (human)
Source (engineered)Expression System: Homo sapiens (human)

+
Component #7: protein, Megabody Mb38

ProteinName: Megabody Mb38 / Details: 6His tag: HHHHHH EPEA tag: EPEA / Number of Copies: 1 / Recombinant expression: No
MassTheoretical: 57.784301 kDa
SourceSpecies: Lama glama (llama)
Source (engineered)Expression System: Escherichia coli (E. coli)

+
Component #8: ligand, bicuculline methochloride

LigandName: bicuculline methochloride / Number of Copies: 2 / Recombinant expression: No
MassTheoretical: 0.367352 kDa

+
Component #9: ligand, [(2R)-2-octanoyloxy-3-[oxidanyl-[(1R,2R,3S,4R,5R,6S)-2,3,...

LigandName: [(2R)-2-octanoyloxy-3-[oxidanyl-[(1R,2R,3S,4R,5R,6S)-2,3,6-tris(oxidanyl)-4,5-diphosphonooxy-cyclohexyl]oxy-phosphoryl]oxy-propyl] octanoate
Number of Copies: 2 / Recombinant expression: No
MassTheoretical: 0.746566 kDa

+
Component #10: ligand, N-ACETYL-D-GLUCOSAMINE

LigandName: N-ACETYL-D-GLUCOSAMINEN-Acetylglucosamine / Number of Copies: 14 / Recombinant expression: No
MassTheoretical: 0.221208 kDa

+
Component #11: ligand, BETA-D-MANNOSE

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

+
Component #12: ligand, ALPHA-D-MANNOSE

LigandName: ALPHA-D-MANNOSE / Number of Copies: 12 / Recombinant expression: No
MassTheoretical: 0.180156 kDa

-
Experimental details

-
Sample preparation

SpecimenSpecimen state: Particle / Method: cryo EM
Sample solutionSpecimen conc.: 0.1 mg/mL / pH: 7.6
VitrificationInstrument: FEI VITROBOT MARK IV / Cryogen name: ETHANE

-
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: 30 e/Å2 / Illumination mode: FLOOD BEAM
LensMagnification: 75000.0 X (nominal) / Cs: 2.7 mm / Imaging mode: BRIGHT FIELD / Defocus: 500.0 - 700.0 nm
Specimen HolderModel: FEI TITAN KRIOS AUTOGRID HOLDER
CameraDetector: FEI FALCON III (4k x 4k)

-
Image processing

ProcessingMethod: single particle reconstruction / Applied symmetry: C1 (asymmetric) / Number of projections: 30536
3D reconstructionSoftware: RELION / Resolution: 3.69 Å / Resolution method: FSC 0.143 CUT-OFF

-
Atomic model buiding

Modeling #1Refinement protocol: flexible

+
About Yorodumi

-
News

-
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. (see PDBe EMDB page)
  • 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 at PDBe / 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. See below links for details.
  • 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

+
Jun 16, 2017. Omokage search with filter

Omokage search with filter

  • Result of Omokage search can be filtered by keywords and the database types

Related info.:Omokage search

+
Sep 15, 2016. EM Navigator & Yorodumi renewed

EM Navigator & Yorodumi renewed

  • New versions of EM Navigator and Yorodumi started

Related info.:Changes in new EM Navigator and Yorodumi

+
Aug 31, 2016. New EM Navigator & Yorodumi

New EM Navigator & Yorodumi

  • In 15th Sep 2016, the development versions of EM Navigator and Yorodumi will replace the official versions.
  • Current version will continue as 'legacy version' for some time.

Related info.:Changes in new EM Navigator and Yorodumi / EM Navigator / Yorodumi

Read more

-
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.

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