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- PDB-7m6m: Full length alpha1 Glycine receptor in presence of 32uM Tetrahydr... -

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Basic information

Entry
Database: PDB / ID: 7m6m
TitleFull length alpha1 Glycine receptor in presence of 32uM Tetrahydrocannabinol
ComponentsGlycine receptor subunit alphaZ1
KeywordsMEMBRANE PROTEIN / Ions Ligands Receptors / Glycine receptor Recombinant Proteins
Function / homology
Function and homology information


Neurotransmitter receptors and postsynaptic signal transmission / extracellularly glycine-gated ion channel activity / extracellularly glycine-gated chloride channel activity / synaptic transmission, glycinergic / cellular response to ethanol / cellular response to zinc ion / regulation of neuron differentiation / glycine binding / chloride channel complex / ligand-gated monoatomic ion channel activity ...Neurotransmitter receptors and postsynaptic signal transmission / extracellularly glycine-gated ion channel activity / extracellularly glycine-gated chloride channel activity / synaptic transmission, glycinergic / cellular response to ethanol / cellular response to zinc ion / regulation of neuron differentiation / glycine binding / chloride channel complex / ligand-gated monoatomic ion channel activity / neuropeptide signaling pathway / response to amino acid / monoatomic ion transport / chloride transmembrane transport / transmitter-gated monoatomic ion channel activity involved in regulation of postsynaptic membrane potential / central nervous system development / cellular response to amino acid stimulus / transmembrane signaling receptor activity / perikaryon / postsynaptic membrane / neuron projection / dendrite / synapse / zinc ion binding / membrane / plasma membrane
Similarity search - Function
Glycine receptor alpha1 / Glycine receptor alpha / Gamma-aminobutyric acid A receptor/Glycine receptor alpha / Neurotransmitter-gated ion-channel, conserved site / Neurotransmitter-gated ion-channels signature. / Neurotransmitter-gated ion-channel transmembrane domain / Neurotransmitter-gated ion-channel transmembrane region / Neurotransmitter-gated ion-channel transmembrane domain superfamily / Neuronal acetylcholine receptor / Neurotransmitter-gated ion-channel ...Glycine receptor alpha1 / Glycine receptor alpha / Gamma-aminobutyric acid A receptor/Glycine receptor alpha / Neurotransmitter-gated ion-channel, conserved site / Neurotransmitter-gated ion-channels signature. / Neurotransmitter-gated ion-channel transmembrane domain / Neurotransmitter-gated ion-channel transmembrane region / Neurotransmitter-gated ion-channel transmembrane domain superfamily / Neuronal acetylcholine receptor / Neurotransmitter-gated ion-channel / Neurotransmitter-gated ion-channel ligand-binding domain / Neurotransmitter-gated ion-channel ligand-binding domain superfamily / Neurotransmitter-gated ion-channel ligand binding domain
Similarity search - Domain/homology
Chem-TCI / Glycine receptor subunit alphaZ1
Similarity search - Component
Biological speciesDanio rerio (zebrafish)
MethodELECTRON MICROSCOPY / single particle reconstruction / cryo EM / Resolution: 3.09 Å
AuthorsKumar, A. / Chakrapani, S.
Funding support United States, 2items
OrganizationGrant numberCountry
American Heart Association20POST35210394 United States
National Institutes of Health/National Institute of General Medical Sciences (NIH/NIGMS)R35GM134896 United States
Citation
Journal: Nat Commun / Year: 2022
Title: Structural basis for cannabinoid-induced potentiation of alpha1-glycine receptors in lipid nanodiscs.
Authors: Arvind Kumar / Kayla Kindig / Shanlin Rao / Afroditi-Maria Zaki / Sandip Basak / Mark S P Sansom / Philip C Biggin / Sudha Chakrapani /
Abstract: Nociception and motor coordination are critically governed by glycine receptor (GlyR) function at inhibitory synapses. Consequentially, GlyRs are attractive targets in the management of chronic pain ...Nociception and motor coordination are critically governed by glycine receptor (GlyR) function at inhibitory synapses. Consequentially, GlyRs are attractive targets in the management of chronic pain and in the treatment of several neurological disorders. High-resolution mechanistic details of GlyR function and its modulation are just emerging. While it has been known that cannabinoids such as Δ-tetrahydrocannabinol (THC), the principal psychoactive constituent in marijuana, potentiate GlyR in the therapeutically relevant concentration range, the molecular mechanism underlying this effect is still not understood. Here, we present Cryo-EM structures of full-length GlyR reconstituted into lipid nanodisc in complex with THC under varying concentrations of glycine. The GlyR-THC complexes are captured in multiple conformational states that reveal the basis for THC-mediated potentiation, manifested as different extents of opening at the level of the channel pore. Taken together, these structural findings, combined with molecular dynamics simulations and functional analysis, provide insights into the potential THC binding site and the allosteric coupling to the channel pore.
#1: Journal: Nat Commun / Year: 2020
Title: Mechanisms of activation and desensitization of full-length glycine receptor in lipid nanodiscs.
Authors: Arvind Kumar / Sandip Basak / Shanlin Rao / Yvonne Gicheru / Megan L Mayer / Mark S P Sansom / Sudha Chakrapani /
Abstract: Glycinergic synapses play a central role in motor control and pain processing in the central nervous system. Glycine receptors (GlyRs) are key players in mediating fast inhibitory neurotransmission ...Glycinergic synapses play a central role in motor control and pain processing in the central nervous system. Glycine receptors (GlyRs) are key players in mediating fast inhibitory neurotransmission at these synapses. While previous high-resolution structures have provided insights into the molecular architecture of GlyR, several mechanistic questions pertaining to channel function are still unanswered. Here, we present Cryo-EM structures of the full-length GlyR protein complex reconstituted into lipid nanodiscs that are captured in the unliganded (closed), glycine-bound (open and desensitized), and allosteric modulator-bound conformations. A comparison of these states reveals global conformational changes underlying GlyR channel gating and modulation. The functional state assignments were validated by molecular dynamics simulations, and the observed permeation events are in agreement with the anion selectivity and conductance of GlyR. These studies provide the structural basis for gating, ion selectivity, and single-channel conductance properties of GlyR in a lipid environment.
History
DepositionMar 26, 2021Deposition site: RCSB / Processing site: RCSB
Revision 1.0Aug 3, 2022Provider: repository / Type: Initial release
Revision 1.1Aug 10, 2022Group: Structure summary / Category: struct / Item: _struct.title
Revision 1.2Sep 7, 2022Group: Database references / Category: citation / citation_author
Item: _citation.journal_abbrev / _citation.journal_id_CSD ..._citation.journal_abbrev / _citation.journal_id_CSD / _citation.journal_id_ISSN / _citation.journal_volume / _citation.page_first / _citation.page_last / _citation.pdbx_database_id_DOI / _citation.pdbx_database_id_PubMed / _citation.title / _citation.year

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Structure visualization

Structure viewerMolecule:
MolmilJmol/JSmol

Downloads & links

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Assembly

Deposited unit
A: Glycine receptor subunit alphaZ1
B: Glycine receptor subunit alphaZ1
C: Glycine receptor subunit alphaZ1
E: Glycine receptor subunit alphaZ1
D: Glycine receptor subunit alphaZ1
hetero molecules


Theoretical massNumber of molelcules
Total (without water)257,80315
Polymers254,1095
Non-polymers3,69410
Water0
1


  • Idetical with deposited unit
  • defined by author
  • Evidence: microscopy
TypeNameSymmetry operationNumber
identity operation1_5551

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Components

#1: Protein
Glycine receptor subunit alphaZ1 /


Mass: 50821.711 Da / Num. of mol.: 5
Source method: isolated from a genetically manipulated source
Source: (gene. exp.) Danio rerio (zebrafish) / Gene: glra1 / Production host: Spodoptera frugiperda (fall armyworm) / References: UniProt: O93430
#2: Polysaccharide
2-acetamido-2-deoxy-beta-D-glucopyranose-(1-4)-2-acetamido-2-deoxy-beta-D-glucopyranose


Type: oligosaccharide / Mass: 424.401 Da / Num. of mol.: 5
Source method: isolated from a genetically manipulated source
DescriptorTypeProgram
DGlcpNAcb1-4DGlcpNAcb1-ROHGlycam Condensed SequenceGMML 1.0
WURCS=2.0/1,2,1/[a2122h-1b_1-5_2*NCC/3=O]/1-1/a4-b1WURCSPDB2Glycan 1.1.0
[][D-1-deoxy-GlcpNAc]{[(4+1)][b-D-GlcpNAc]{}}LINUCSPDB-CARE
#3: Chemical
ChemComp-TCI / (6aR,10aR)-6,6,9-trimethyl-3-pentyl-6a,7,8,10a-tetrahydro-6H-benzo[c]chromen-1-ol / Tetrahydrocannabinol / Tetrahydrocannabinol


Mass: 314.462 Da / Num. of mol.: 5 / Source method: obtained synthetically / Formula: C21H30O2 / Feature type: SUBJECT OF INVESTIGATION
Has ligand of interestY

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Experimental details

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Experiment

ExperimentMethod: ELECTRON MICROSCOPY
EM experimentAggregation state: PARTICLE / 3D reconstruction method: single particle reconstruction

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Sample preparation

ComponentName: Glycine receptor subunit alpha Z1 / Type: COMPLEX / Entity ID: #1 / Source: RECOMBINANT
Molecular weightValue: 250 kDa/nm / Experimental value: NO
Source (natural)Organism: Danio rerio (zebrafish)
Source (recombinant)Organism: Spodoptera frugiperda (fall armyworm)
Buffer solutionpH: 8
Buffer component
IDConc.NameFormulaBuffer-ID
150 mMTris4H11NO31
2200 mMSodium ChlorideNaClSodium chloride1
SpecimenConc.: 0.1 mg/ml / Embedding applied: NO / Shadowing applied: NO / Staining applied: NO / Vitrification applied: YES
Details: Full length Zebrafish GlyR alpha1 homopentamer reconsituted in Nanodisc
VitrificationInstrument: FEI VITROBOT MARK IV / Cryogen name: ETHANE / Humidity: 100 % / Chamber temperature: 4 K
Details: 3.5 ul of 0.1 mg/ml protein solution was applied on a grid in the Vitrobot MkIV chamber set to 100% RH at 4 degC for 30s and then blotted for 2 s and plunged

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Electron microscopy imaging

Experimental equipment
Model: Titan Krios / Image courtesy: FEI Company
MicroscopyModel: FEI TITAN KRIOS
Electron gunElectron source: FIELD EMISSION GUN / Accelerating voltage: 300 kV / Illumination mode: FLOOD BEAM
Electron lensMode: BRIGHT FIELDBright-field microscopy / Nominal magnification: 130000 X / Nominal defocus max: 2500 nm / Nominal defocus min: 1000 nm / Cs: 2.7 mm / C2 aperture diameter: 100 µm / Alignment procedure: COMA FREE
Specimen holderCryogen: NITROGEN / Specimen holder model: FEI TITAN KRIOS AUTOGRID HOLDER
Image recordingAverage exposure time: 9 sec. / Electron dose: 40 e/Å2 / Detector mode: SUPER-RESOLUTION / Film or detector model: GATAN K3 BIOQUANTUM (6k x 4k) / Num. of grids imaged: 8 / Num. of real images: 9700
EM imaging opticsEnergyfilter name: GIF Bioquantum / Energyfilter slit width: 20 eV
Image scansMovie frames/image: 40

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Processing

SoftwareName: PHENIX / Version: 1.19_4092: / Classification: refinement
EM software
IDNameVersionCategory
2LatitudeSimage acquisition
13RELION3.13D reconstruction
CTF correctionType: PHASE FLIPPING AND AMPLITUDE CORRECTION
SymmetryPoint symmetry: C5 (5 fold cyclic)
3D reconstructionResolution: 3.09 Å / Resolution method: FSC 0.143 CUT-OFF / Num. of particles: 25539 / Symmetry type: POINT
Refine LS restraints
Refine-IDTypeDev idealNumber
ELECTRON MICROSCOPYf_bond_d0.00814545
ELECTRON MICROSCOPYf_angle_d1.32719745
ELECTRON MICROSCOPYf_dihedral_angle_d11.6745325
ELECTRON MICROSCOPYf_chiral_restr0.082240
ELECTRON MICROSCOPYf_plane_restr0.0082435

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