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Yorodumi- EMDB-4411: Cryo-EM structure of the human synaptic alpha1-beta3-gamma2 GABAA... -
+Open data
-Basic information
Entry | Database: EMDB / ID: EMD-4411 | ||||||||||||||||||||||||
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Title | Cryo-EM structure of the human synaptic alpha1-beta3-gamma2 GABAA receptor in complex with Megabody38 in a lipid nanodisc | ||||||||||||||||||||||||
Map data | Cryo-EM map of a GABAA receptor in lipid nanodisc | ||||||||||||||||||||||||
Sample |
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Function / homology | Function and homology information benzodiazepine receptor activity / GABA receptor complex / cellular response to histamine / GABA receptor activation / GABA-A receptor activity / GABA-gated chloride ion channel activity / GABA-A receptor complex / inhibitory synapse assembly / Signaling by ERBB4 / synaptic transmission, GABAergic ...benzodiazepine receptor activity / GABA receptor complex / cellular response to histamine / GABA receptor activation / GABA-A receptor activity / GABA-gated chloride ion channel activity / GABA-A receptor complex / inhibitory synapse assembly / Signaling by ERBB4 / synaptic transmission, GABAergic / gamma-aminobutyric acid signaling pathway / postsynaptic specialization membrane / neurotransmitter receptor activity / adult behavior / chloride channel activity / roof of mouth development / chloride channel complex / transmembrane transporter complex / GABA-ergic synapse / regulation of postsynaptic membrane potential / chloride transmembrane transport / dendrite membrane / post-embryonic development / transmitter-gated monoatomic ion channel activity involved in regulation of postsynaptic membrane potential / cytoplasmic vesicle membrane / postsynapse / neuron projection / axon / synapse / signal transduction / identical protein binding / plasma membrane Similarity search - Function | ||||||||||||||||||||||||
Biological species | Homo sapiens (human) / Lama glama (llama) | ||||||||||||||||||||||||
Method | single particle reconstruction / cryo EM / Resolution: 3.2 Å | ||||||||||||||||||||||||
Authors | Laverty D / Desai R / Uchanski T / Masiulis S / Wojciech JS / Malinauskas T / Zivanov J / Pardon E / Steyaert J / Miller KW / Aricescu AR | ||||||||||||||||||||||||
Funding support | United Kingdom, Switzerland, United States, 7 items
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Citation | Journal: Nature / Year: 2019 Title: Cryo-EM structure of the human α1β3γ2 GABA receptor in a lipid bilayer. Authors: Duncan Laverty / Rooma Desai / Tomasz Uchański / Simonas Masiulis / Wojciech J Stec / Tomas Malinauskas / Jasenko Zivanov / Els Pardon / Jan Steyaert / Keith W Miller / A Radu Aricescu / Abstract: Type A γ-aminobutyric acid (GABA) receptors are pentameric ligand-gated ion channels and the main drivers of fast inhibitory neurotransmission in the vertebrate nervous system. Their dysfunction is ...Type A γ-aminobutyric acid (GABA) receptors are pentameric ligand-gated ion channels and the main drivers of fast inhibitory neurotransmission in the vertebrate nervous system. Their dysfunction is implicated in a range of neurological disorders, including depression, epilepsy and schizophrenia. Among the numerous assemblies that are theoretically possible, the most prevalent in the brain are the α1β2/3γ2 GABA receptors. The β3 subunit has an important role in maintaining inhibitory tone, and the expression of this subunit alone is sufficient to rescue inhibitory synaptic transmission in β1-β3 triple knockout neurons. So far, efforts to generate accurate structural models for heteromeric GABA receptors have been hampered by the use of engineered receptors and the presence of detergents. Notably, some recent cryo-electron microscopy reconstructions have reported 'collapsed' conformations; however, these disagree with the structure of the prototypical pentameric ligand-gated ion channel the Torpedo nicotinic acetylcholine receptor, the large body of structural work on homologous homopentameric receptor variants and the logic of an ion-channel architecture. Here we present a high-resolution cryo-electron microscopy structure of the full-length human α1β3γ2L-a major synaptic GABA receptor isoform-that is functionally reconstituted in lipid nanodiscs. The receptor is bound to a positive allosteric modulator 'megabody' and is in a desensitized conformation. Each GABA receptor pentamer contains two phosphatidylinositol-4,5-bisphosphate molecules, the head groups of which occupy positively charged pockets in the intracellular juxtamembrane regions of α1 subunits. Beyond this level, the intracellular M3-M4 loops are largely disordered, possibly because interacting post-synaptic proteins are not present. This structure illustrates the molecular principles of heteromeric GABA receptor organization and provides a reference framework for future mechanistic investigations of GABAergic signalling and pharmacology. | ||||||||||||||||||||||||
History |
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-Structure visualization
Movie |
Movie viewer |
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Structure viewer | EM map: SurfViewMolmilJmol/JSmol |
Supplemental images |
-Downloads & links
-EMDB archive
Map data | emd_4411.map.gz | 5.4 MB | EMDB map data format | |
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Header (meta data) | emd-4411-v30.xml emd-4411.xml | 26.6 KB 26.6 KB | Display Display | EMDB header |
Images | emd_4411.png | 69.7 KB | ||
Masks | emd_4411_msk_1.map | 64 MB | Mask map | |
Others | emd_4411_half_map_1.map.gz emd_4411_half_map_2.map.gz | 50.1 MB 50 MB | ||
Archive directory | http://ftp.pdbj.org/pub/emdb/structures/EMD-4411 ftp://ftp.pdbj.org/pub/emdb/structures/EMD-4411 | HTTPS FTP |
-Validation report
Summary document | emd_4411_validation.pdf.gz | 497.6 KB | Display | EMDB validaton report |
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Full document | emd_4411_full_validation.pdf.gz | 497.2 KB | Display | |
Data in XML | emd_4411_validation.xml.gz | 12.2 KB | Display | |
Data in CIF | emd_4411_validation.cif.gz | 14.3 KB | Display | |
Arichive directory | https://ftp.pdbj.org/pub/emdb/validation_reports/EMD-4411 ftp://ftp.pdbj.org/pub/emdb/validation_reports/EMD-4411 | HTTPS FTP |
-Related structure data
Related structure data | 6i53MC M: atomic model generated by this map C: citing same article (ref.) |
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Similar structure data |
-Links
EMDB pages | EMDB (EBI/PDBe) / EMDataResource |
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Related items in Molecule of the Month |
-Map
File | Download / File: emd_4411.map.gz / Format: CCP4 / Size: 64 MB / Type: IMAGE STORED AS FLOATING POINT NUMBER (4 BYTES) | ||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||
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Annotation | Cryo-EM map of a GABAA receptor in lipid nanodisc | ||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||
Voxel size | X=Y=Z: 1.07 Å | ||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||
Density |
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Symmetry | Space group: 1 | ||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||
Details | EMDB XML:
CCP4 map header:
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-Supplemental data
-Mask #1
File | emd_4411_msk_1.map | ||||||||||||
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Projections & Slices |
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Density Histograms |
-Half map: #2
File | emd_4411_half_map_1.map | ||||||||||||
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Projections & Slices |
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Density Histograms |
-Half map: #1
File | emd_4411_half_map_2.map | ||||||||||||
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Projections & Slices |
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Density Histograms |
-Sample components
-Entire : Human alpha1-beta3-gamma2 GABA-A receptor in complex with Megabod...
Entire | Name: Human alpha1-beta3-gamma2 GABA-A receptor in complex with Megabody38 and in lipid nanodiscs |
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Components |
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-Supramolecule #1: Human alpha1-beta3-gamma2 GABA-A receptor in complex with Megabod...
Supramolecule | Name: Human alpha1-beta3-gamma2 GABA-A receptor in complex with Megabody38 and in lipid nanodiscs type: complex / ID: 1 / Parent: 0 / Macromolecule list: #1-#4 |
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Molecular weight | Theoretical: 400 KDa |
-Supramolecule #2: Human alpha1-beta3-gamma2 GABA-A receptor
Supramolecule | Name: Human alpha1-beta3-gamma2 GABA-A receptor / type: complex / ID: 2 / Parent: 1 / Macromolecule list: #1-#3 |
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Source (natural) | Organism: Homo sapiens (human) |
Recombinant expression | Organism: Homo sapiens (human) / Recombinant strain: Embryonic kidney cells |
-Supramolecule #3: Megabody38
Supramolecule | Name: Megabody38 / type: complex / ID: 3 / Parent: 1 / Macromolecule list: #4 |
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Source (natural) | Organism: Lama glama (llama) |
Recombinant expression | Organism: Escherichia coli (E. coli) |
-Macromolecule #1: Gamma-aminobutyric acid receptor subunit beta-3
Macromolecule | Name: Gamma-aminobutyric acid receptor subunit beta-3 / type: protein_or_peptide / ID: 1 / Number of copies: 2 / Enantiomer: LEVO |
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Source (natural) | Organism: Homo sapiens (human) |
Molecular weight | Theoretical: 54.444578 KDa |
Recombinant expression | Organism: Homo sapiens (human) |
Sequence | String: MCSGLLELLL PIWLSWTLGT RGSEPRSVND PGNMSFVKET VDKLLKGYDI RLRPDFGGPP VCVGMNIDIA SIDMVSEVNM DYTLTMYFQ QYWRDKRLAY SGIPLNLTLD NRVADQLWVP DTYFLNDKKS FVHGVTVKNR MIRLHPDGTV LYGLRITTTA A CMMDLRRY ...String: MCSGLLELLL PIWLSWTLGT RGSEPRSVND PGNMSFVKET VDKLLKGYDI RLRPDFGGPP VCVGMNIDIA SIDMVSEVNM DYTLTMYFQ QYWRDKRLAY SGIPLNLTLD NRVADQLWVP DTYFLNDKKS FVHGVTVKNR MIRLHPDGTV LYGLRITTTA A CMMDLRRY PLDEQNCTLE IESYGYTTDD IEFYWRGGDK AVTGVERIEL PQFSIVEHRL VSRNVVFATG AYPRLSLSFR LK RNIGYFI LQTYMPSILI TILSWVSFWI NYDASAARVA LGITTVLTMT TINTHLRETL PKIPYVKAID MYLMGCFVFV FLA LLEYAF VNYIFFGRGP QRQKKLAEKT AKAKNDRSKS ESNRVDAHGN ILLTSLEVHN EMNEVSGGIG DTRNSAISFD NSGI QYRKQ SMPREGHGRF LGDRSLPHKK THLRRRSSQL KIKIPDLTDV NAIDRWSRIV FPFTFSLFNL VYWLYYVN |
-Macromolecule #2: Gamma-aminobutyric acid receptor subunit alpha-1
Macromolecule | Name: Gamma-aminobutyric acid receptor subunit alpha-1 / type: protein_or_peptide / ID: 2 / Number of copies: 2 / Enantiomer: LEVO |
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Source (natural) | Organism: Homo sapiens (human) |
Molecular weight | Theoretical: 52.916602 KDa |
Recombinant expression | Organism: Homo sapiens (human) |
Sequence | String: MKKSPGLSDY LWAWTLFLST LTGRSYGDYK DDDDKQPSLQ DELKDNTTVF TRILDRLLDG YDNRLRPGLG ERVTEVKTDI FVTSFGPVS DHDMEYTIDV FFRQSWKDER LKFKGPMTVL RLNNLMASKI WTPDTFFHNG KKSVAHNMTM PNKLLRITED G TLLYTMRL ...String: MKKSPGLSDY LWAWTLFLST LTGRSYGDYK DDDDKQPSLQ DELKDNTTVF TRILDRLLDG YDNRLRPGLG ERVTEVKTDI FVTSFGPVS DHDMEYTIDV FFRQSWKDER LKFKGPMTVL RLNNLMASKI WTPDTFFHNG KKSVAHNMTM PNKLLRITED G TLLYTMRL TVRAECPMHL EDFPMDAHAC PLKFGSYAYT RAEVVYEWTR EPARSVVVAE DGSRLNQYDL LGQTVDSGIV QS STGEYVV MTTHFHLKRK IGYFVIQTYL PCIMTVILSQ VSFWLNRESV PARTVFGVTT VLTMTTLSIS ARNSLPKVAY ATA MDWFIA VCYAFVFSAL IEFATVNYFT KRGYAWDGKS VVPEKPKKVK DPLIKKNNTY APTATSYTPN LARGDPGLAT IAKS ATIEP KEVKPETKPP EPKKTFNSVS KIDRLSRIAF PLLFGIFNLV YWATYLNREP QLKAPTPHQ |
-Macromolecule #3: Gamma-aminobutyric acid receptor subunit gamma-2
Macromolecule | Name: Gamma-aminobutyric acid receptor subunit gamma-2 / type: protein_or_peptide / ID: 3 / Number of copies: 1 / Enantiomer: LEVO |
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Source (natural) | Organism: Homo sapiens (human) |
Molecular weight | Theoretical: 56.922055 KDa |
Recombinant expression | Organism: Homo sapiens (human) |
Sequence | String: MSSPNIWSTG SSVYSTPVFS QKMTVWILLL LSLYPGFTSQ KSDDDYEDYA SNKTWVLTPK VPEGDVTVIL NNLLEGYDNK LRPDIGVKP TLIHTDMYVN SIGPVNAINM EYTIDIFFAQ TWYDRRLKFN STIKVLRLNS NMVGKIWIPD TFFRNSKKAD A HWITTPNR ...String: MSSPNIWSTG SSVYSTPVFS QKMTVWILLL LSLYPGFTSQ KSDDDYEDYA SNKTWVLTPK VPEGDVTVIL NNLLEGYDNK LRPDIGVKP TLIHTDMYVN SIGPVNAINM EYTIDIFFAQ TWYDRRLKFN STIKVLRLNS NMVGKIWIPD TFFRNSKKAD A HWITTPNR MLRIWNDGRV LYTLRLTIDA ECQLQLHNFP MDEHSCPLEF SSYGYPREEI VYQWKRSSVE VGDTRSWRLY QF SFVGLRN TTEVVKTTSG DYVVMSVYFD LSRRMGYFTI QTYIPCTLIV VLSWVSFWIN KDAVPARTSL GITTVLTMTT LST IARKSL PKVSYVTAMD LFVSVCFIFV FSALVEYGTL HYFVSNRKPS KDKDKKKKNP LLRMFSFKAP TIDIRPRSAT IQMN NATHL QERDEEYGYE CLDGKDCASF FCCFEDCRTG AWRHGRIHIR IAKMDSYARI FFPTAFCLFN LVYWVSYLYL GGSGG SGGS GKTETSQVAP A |
-Macromolecule #4: Megabody38
Macromolecule | Name: Megabody38 / type: protein_or_peptide / ID: 4 / Number of copies: 1 / Enantiomer: LEVO |
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Source (natural) | Organism: Lama glama (llama) |
Molecular weight | Theoretical: 13.462909 KDa |
Recombinant expression | Organism: Escherichia coli (E. coli) |
Sequence | String: QVQLQESGGG LVQAGGSLRV SCAASGRTFT AYIMAWFRQA PGKEREFLAA MDQGRIQYYG DSVRGRFTIS RDYAKNSVDL QLDGLRPED TAVYYCAAGA GFWGLRTASS YHYWGQGTQV TVSS |
-Macromolecule #9: (2S)-3-(hexadecanoyloxy)-2-[(9Z)-octadec-9-enoyloxy]propyl 2-(tri...
Macromolecule | Name: (2S)-3-(hexadecanoyloxy)-2-[(9Z)-octadec-9-enoyloxy]propyl 2-(trimethylammonio)ethyl phosphate type: ligand / ID: 9 / Number of copies: 3 / Formula: POV |
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Molecular weight | Theoretical: 760.076 Da |
Chemical component information | ChemComp-POV: |
-Macromolecule #10: [(2R)-2-octanoyloxy-3-[oxidanyl-[(1R,2R,3S,4R,5R,6S)-2,3,6-tris(o...
Macromolecule | Name: [(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 type: ligand / ID: 10 / Number of copies: 2 / Formula: PIO |
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Molecular weight | Theoretical: 746.566 Da |
Chemical component information | ChemComp-PIO: |
-Experimental details
-Structure determination
Method | cryo EM |
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Processing | single particle reconstruction |
Aggregation state | particle |
-Sample preparation
Concentration | 0.5 mg/mL | |||||||||
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Buffer | pH: 7.2 Component:
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Grid | Model: Quantifoil, UltrAuFoil / Material: GOLD / Pretreatment - Type: GLOW DISCHARGE | |||||||||
Vitrification | Cryogen name: ETHANE / Chamber humidity: 100 % / Chamber temperature: 287.15 K / Instrument: FEI VITROBOT MARK IV | |||||||||
Details | The purified receptor-nanodisc complex was mixed with Mb38 prior to grid preparation. |
-Electron microscopy
Microscope | FEI TITAN KRIOS |
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Specialist optics | Phase plate: VOLTA PHASE PLATE |
Image recording | Film or detector model: FEI FALCON III (4k x 4k) / Detector mode: COUNTING / Number grids imaged: 1 / Number real images: 784 / Average exposure time: 60.0 sec. / Average electron dose: 30.84 e/Å2 |
Electron beam | Acceleration voltage: 300 kV / Electron source: FIELD EMISSION GUN |
Electron optics | C2 aperture diameter: 50.0 µm / Calibrated defocus max: 0.7000000000000001 µm / Calibrated defocus min: 0.5 µm / Calibrated magnification: 75000 / Illumination mode: FLOOD BEAM / Imaging mode: BRIGHT FIELD / Cs: 2.7 mm / Nominal defocus max: 0.7000000000000001 µm / Nominal defocus min: 0.5 µm / Nominal magnification: 75000 |
Sample stage | Specimen holder model: FEI TITAN KRIOS AUTOGRID HOLDER |
Experimental equipment | Model: Titan Krios / Image courtesy: FEI Company |
+Image processing
-Atomic model buiding 1
Refinement | Space: REAL / Protocol: AB INITIO MODEL |
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Output model | PDB-6i53: |