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Yorodumi- EMDB-14072: Cryo-EM structure of human full-length alpha4beta3gamma2 GABA(A)R... -
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-Basic information
Entry | Database: EMDB / ID: EMD-14072 | ||||||||||||||||||
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Title | Cryo-EM structure of human full-length alpha4beta3gamma2 GABA(A)R in complex with GABA and nanobody Nb25 | ||||||||||||||||||
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Sample |
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Keywords | pentameric ligand-gated ion channel / neurotransmitter receptor / GABA receptor / MEMBRANE PROTEIN | ||||||||||||||||||
Function / homology | Function and homology information benzodiazepine receptor activity / inhibitory extracellular ligand-gated monoatomic ion channel activity / GABA receptor activation / GABA-gated chloride ion channel activity / cellular response to histamine / inhibitory synapse assembly / GABA-A receptor activity / GABA-A receptor complex / postsynaptic specialization membrane / neurotransmitter receptor activity ...benzodiazepine receptor activity / inhibitory extracellular ligand-gated monoatomic ion channel activity / GABA receptor activation / GABA-gated chloride ion channel activity / cellular response to histamine / inhibitory synapse assembly / GABA-A receptor activity / GABA-A receptor complex / postsynaptic specialization membrane / neurotransmitter receptor activity / synaptic transmission, GABAergic / gamma-aminobutyric acid signaling pathway / chloride transport / chloride channel activity / roof of mouth development / Signaling by ERBB4 / extracellular ligand-gated monoatomic ion channel activity / chloride channel complex / GABA-ergic synapse / regulation of postsynaptic membrane potential / chloride transmembrane transport / dendrite membrane / transmitter-gated monoatomic ion channel activity involved in regulation of postsynaptic membrane potential / cytoplasmic vesicle membrane / postsynapse / postsynaptic membrane / neuron projection / 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: 3.0 Å | ||||||||||||||||||
Authors | Sente A / Desai R | ||||||||||||||||||
Funding support | United Kingdom, 5 items
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Citation | Journal: Nature / Year: 2022 Title: Differential assembly diversifies GABA receptor structures and signalling. Authors: Andrija Sente / Rooma Desai / Katerina Naydenova / Tomas Malinauskas / Youssef Jounaidi / Jonas Miehling / Xiaojuan Zhou / Simonas Masiulis / Steven W Hardwick / Dimitri Y Chirgadze / Keith ...Authors: Andrija Sente / Rooma Desai / Katerina Naydenova / Tomas Malinauskas / Youssef Jounaidi / Jonas Miehling / Xiaojuan Zhou / Simonas Masiulis / Steven W Hardwick / Dimitri Y Chirgadze / Keith W Miller / A Radu Aricescu / Abstract: Type A γ-aminobutyric acid receptors (GABARs) are pentameric ligand-gated chloride channels that mediate fast inhibitory signalling in neural circuits and can be modulated by essential medicines ...Type A γ-aminobutyric acid receptors (GABARs) are pentameric ligand-gated chloride channels that mediate fast inhibitory signalling in neural circuits and can be modulated by essential medicines including general anaesthetics and benzodiazepines. Human GABAR subunits are encoded by 19 paralogous genes that can, in theory, give rise to 495,235 receptor types. However, the principles that govern the formation of pentamers, the permutational landscape of receptors that may emerge from a subunit set and the effect that this has on GABAergic signalling remain largely unknown. Here we use cryogenic electron microscopy to determine the structures of extrasynaptic GABARs assembled from α4, β3 and δ subunits, and their counterparts incorporating γ2 instead of δ subunits. In each case, we identified two receptor subtypes with distinct stoichiometries and arrangements, all four differing from those previously observed for synaptic, α1-containing receptors. This, in turn, affects receptor responses to physiological and synthetic modulators by creating or eliminating ligand-binding sites at subunit interfaces. We provide structural and functional evidence that selected GABAR arrangements can act as coincidence detectors, simultaneously responding to two neurotransmitters: GABA and histamine. Using assembly simulations and single-cell RNA sequencing data, we calculated the upper bounds for receptor diversity in recombinant systems and in vivo. We propose that differential assembly is a pervasive mechanism for regulating the physiology and pharmacology of GABARs. | ||||||||||||||||||
History |
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-Structure visualization
Supplemental images |
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-Downloads & links
-EMDB archive
Map data | emd_14072.map.gz | 71.6 MB | EMDB map data format | |
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Header (meta data) | emd-14072-v30.xml emd-14072.xml | 18.4 KB 18.4 KB | Display Display | EMDB header |
Images | emd_14072.png | 59.8 KB | ||
Filedesc metadata | emd-14072.cif.gz | 7.2 KB | ||
Archive directory | http://ftp.pdbj.org/pub/emdb/structures/EMD-14072 ftp://ftp.pdbj.org/pub/emdb/structures/EMD-14072 | HTTPS FTP |
-Related structure data
Related structure data | 7qnaMC 7qn5C 7qn6C 7qn7C 7qn8C 7qn9C 7qnbC 7qncC 7qndC 7qneC M: atomic model generated by this map C: citing same article (ref.) |
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Similar structure data | Similarity search - Function & homologyF&H Search |
EM raw data | EMPIAR-10909 (Title: Cryo-EM micrographs of GABA(A)Rs purified from cells expressing human full-length alpha4, beta3 and gamma2 subunits, in presence of GABA and nanobody Nb25 Data size: 3.8 TB Data #1: Unaligned multi-frame micrographs of GABA(A) receptors purified from cells expressing alpha4, beta3 and gamma2 subunits [micrographs - multiframe]) |
-Links
EMDB pages | EMDB (EBI/PDBe) / EMDataResource |
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Related items in Molecule of the Month |
-Map
File | Download / File: emd_14072.map.gz / Format: CCP4 / Size: 93 MB / Type: IMAGE STORED AS FLOATING POINT NUMBER (4 BYTES) | ||||||||||||||||||||||||||||||||||||
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Projections & slices | Image control
Images are generated by Spider. | ||||||||||||||||||||||||||||||||||||
Voxel size | X=Y=Z: 0.896 Å | ||||||||||||||||||||||||||||||||||||
Density |
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Symmetry | Space group: 1 | ||||||||||||||||||||||||||||||||||||
Details | EMDB XML:
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-Supplemental data
-Sample components
-Entire : Human full-length alpha4beta3gamma2 GABA(A)R in complex with GABA...
Entire | Name: Human full-length alpha4beta3gamma2 GABA(A)R in complex with GABA and Nanobody Nb25 |
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Components |
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-Supramolecule #1: Human full-length alpha4beta3gamma2 GABA(A)R in complex with GABA...
Supramolecule | Name: Human full-length alpha4beta3gamma2 GABA(A)R in complex with GABA and Nanobody Nb25 type: complex / ID: 1 / Parent: 0 / Macromolecule list: #1-#4 |
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Source (natural) | Organism: Homo sapiens (human) |
-Macromolecule #1: Gamma-aminobutyric acid receptor subunit alpha-4
Macromolecule | Name: Gamma-aminobutyric acid receptor subunit alpha-4 / type: protein_or_peptide / ID: 1 / Number of copies: 1 / Enantiomer: LEVO |
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Source (natural) | Organism: Homo sapiens (human) |
Molecular weight | Theoretical: 61.696047 KDa |
Recombinant expression | Organism: Homo sapiens (human) |
Sequence | String: MVSAKKVPAI ALSAGVSFAL LRFLCLAVCL NESPGQNQKE EKLCTENFTR ILDSLLDGYD NRLRPGFGGP VTEVKTDIYV TSFGPVSDV EMEYTMDVFF RQTWIDKRLK YDGPIEILRL NNMMVTKVWT PDTFFRNGKK SVSHNMTAPN KLFRIMRNGT I LYTMRLTI ...String: MVSAKKVPAI ALSAGVSFAL LRFLCLAVCL NESPGQNQKE EKLCTENFTR ILDSLLDGYD NRLRPGFGGP VTEVKTDIYV TSFGPVSDV EMEYTMDVFF RQTWIDKRLK YDGPIEILRL NNMMVTKVWT PDTFFRNGKK SVSHNMTAPN KLFRIMRNGT I LYTMRLTI SAECPMRLVD FPMDGHACPL KFGSYAYPKS EMIYTWTKGP EKSVEVPKES SSLVQYDLIG QTVSSETIKS IT GEYIVMT VYFHLRRKMG YFMIQTYIPC IMTVILSQVS FWINKESVPA RTVFGITTVL TMTTLSISAR HSLPKVSYAT AMD WFIAVC FAFVFSALIE FAAVNYFTNI QMEKAKRKTS KPPQEVPAAP VQREKHPEAP LQNTNANLNM RKRTNALVHS ESDV GNRTE VGNHSSKSST VVQESSKGTP RSYLASSPNP FSRANAAETI SAARALPSAS PTSIRTGYMP RKASVGSAST RHVFG SRLQ RIKTTVNTIG ATGKLSATPP PSAPPPSGSG TSKIDKYARI LFPVTFGAFN MVYWVVYLSK DTMEKSESLM UniProtKB: Gamma-aminobutyric acid receptor subunit alpha-4 |
-Macromolecule #2: Gamma-aminobutyric acid receptor subunit beta-3
Macromolecule | Name: Gamma-aminobutyric acid receptor subunit beta-3 / type: protein_or_peptide / ID: 2 / Number of copies: 3 / Enantiomer: LEVO |
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Source (natural) | Organism: Homo sapiens (human) |
Molecular weight | Theoretical: 54.180348 KDa |
Recombinant expression | Organism: Homo sapiens (human) |
Sequence | String: MWGLAGGRLF GIFSAPVLVA VVCCAQSVND PGNMSFVKET VDKLLKGYDI RLRPDFGGPP VCVGMNIDIA SIDMVSEVNM DYTLTMYFQ QYWRDKRLAY SGIPLNLTLD NRVADQLWVP DTYFLNDKKS FVHGVTVKNR MIRLHPDGTV LYGLRITTTA A CMMDLRRY ...String: MWGLAGGRLF GIFSAPVLVA VVCCAQSVND 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 UniProtKB: Gamma-aminobutyric acid receptor subunit beta-3 |
-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: 55.666809 KDa |
Recombinant expression | Organism: Homo sapiens (human) |
Sequence | String: MGILPSPGMP ALLSLVSLLS VLLMGCVAET GQKSDDDYED YTSNKTWVLT PKVPEGDVTV ILNNLLEGYD NKLRPDIGVK PTLIHTDMY VNSIGPVNAI NMEYTIDIFF AQTWYDRRLK FNSTIKVLRL NSNMVGKIWI PDTFFRNSKK ADAHWITTPN R MLRIWNDG ...String: MGILPSPGMP ALLSLVSLLS VLLMGCVAET GQKSDDDYED YTSNKTWVLT PKVPEGDVTV ILNNLLEGYD NKLRPDIGVK PTLIHTDMY VNSIGPVNAI NMEYTIDIFF AQTWYDRRLK FNSTIKVLRL NSNMVGKIWI PDTFFRNSKK ADAHWITTPN R MLRIWNDG RVLYTLRLTI DAECQLQLHN FPMDEHSCPL EFSSYGYPRE EIVYQWKRSS VEVGDTRSWR LYQFSFVGLR NT TEVVKTT SGDYVVMSVY FDLSRRMGYF TIQTYIPCTL IVVLSWVSFW INKDAVPART SLGITTVLTM TTLSTIARKS LPK VSYVTA MDLFVSVCFI FVFSALVEYG TLHYFVSNRK PSKDKDKKKK NPLLRMFSFK APTIDIRPRS ATIQMNNATH LQER DEEYG YECLDGKDCA SFFCCFEDCR TGAWRHGRIH IRIAKMDSYA RIFFPTAFCL FNLVYWVSYL YLGTGGSGGS GGSTE TSQV APA UniProtKB: Gamma-aminobutyric acid receptor subunit gamma-2 |
-Macromolecule #4: Nanobody Nb25
Macromolecule | Name: Nanobody Nb25 / 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: 13.341741 KDa |
Recombinant expression | Organism: Escherichia coli (E. coli) |
Sequence | String: QVQLVESGGG LVQGSLRLSC AASGHTFNYP IMGWFRQAPG KEREFVGAIS WSGGSTSYAD SVKDRFTISR DNAKNTVYLE MNNLKPEDT AVYYCAAKGR YSGGLYYPTN YDYWGQGTQV TV |
-Macromolecule #7: 2-acetamido-2-deoxy-beta-D-glucopyranose
Macromolecule | Name: 2-acetamido-2-deoxy-beta-D-glucopyranose / type: ligand / ID: 7 / Number of copies: 2 / Formula: NAG |
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Molecular weight | Theoretical: 221.208 Da |
Chemical component information | ChemComp-NAG: |
-Macromolecule #8: GAMMA-AMINO-BUTANOIC ACID
Macromolecule | Name: GAMMA-AMINO-BUTANOIC ACID / type: ligand / ID: 8 / Number of copies: 1 / Formula: ABU |
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Molecular weight | Theoretical: 103.12 Da |
Chemical component information | ChemComp-ABU: |
-Experimental details
-Structure determination
Method | cryo EM |
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Processing | single particle reconstruction |
Aggregation state | particle |
-Sample preparation
Buffer | pH: 7.6 Component:
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Grid | Model: UltrAuFoil R1.2/1.3 / Material: GOLD / Pretreatment - Type: GLOW DISCHARGE / Pretreatment - Time: 30 sec. / Pretreatment - Atmosphere: AIR / Details: 30 mA (EasyGlow) | |||||||||
Vitrification | Cryogen name: ETHANE / Chamber humidity: 95 % / Chamber temperature: 287 K |
-Electron microscopy
Microscope | FEI TITAN KRIOS |
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Electron beam | Acceleration voltage: 300 kV / Electron source: FIELD EMISSION GUN |
Electron optics | C2 aperture diameter: 50.0 µm / Illumination mode: FLOOD BEAM / Imaging mode: BRIGHT FIELDBright-field microscopy / Cs: 2.7 mm / Nominal defocus max: 2.1 µm / Nominal defocus min: 0.7000000000000001 µm / Nominal magnification: 130000 |
Specialist optics | Energy filter - Name: GIF Bioquantum / Energy filter - Slit width: 20 eV |
Sample stage | Cooling holder cryogen: NITROGEN |
Image recording | Film or detector model: GATAN K3 (6k x 4k) / Average electron dose: 47.46 e/Å2 |
Experimental equipment | Model: Titan Krios / Image courtesy: FEI Company |
-Image processing
Startup model | Type of model: OTHER / Details: AB INITIO MODEL |
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Initial angle assignment | Type: MAXIMUM LIKELIHOOD / Software - Name: RELION (ver. 3.1) |
Final 3D classification | Software - Name: RELION (ver. 3.1) |
Final angle assignment | Type: MAXIMUM LIKELIHOOD / Software - Name: RELION (ver. 3.1) |
Final reconstruction | Resolution.type: BY AUTHOR / Resolution: 3.0 Å / Resolution method: FSC 0.143 CUT-OFF / Number images used: 55567 |
-Atomic model buiding 1
Refinement | Protocol: AB INITIO MODEL |
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Output model | PDB-7qna: |