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- PDB-6vm3: Full length Glycine receptor reconstituted in lipid nanodisc in G... -

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Basic information

Entry
Database: PDB / ID: 6vm3
TitleFull length Glycine receptor reconstituted in lipid nanodisc in Gly/IVM-conformation (State-3)
ComponentsGlycine receptor subunit alphaZ1
KeywordsMEMBRANE PROTEIN / Ions Ligands Receptors / Glycine receptor Recombinant Proteins Glycine
Function / homology
Function and homology information


transmitter-gated monoatomic ion channel activity / Neurotransmitter receptors and postsynaptic signal transmission / extracellularly glycine-gated ion channel activity / extracellularly glycine-gated chloride channel activity / cellular response to ethanol / cellular response to zinc ion / regulation of neuron differentiation / neurotransmitter receptor activity / glycine binding / chloride channel complex ...transmitter-gated monoatomic ion channel activity / Neurotransmitter receptors and postsynaptic signal transmission / extracellularly glycine-gated ion channel activity / extracellularly glycine-gated chloride channel activity / cellular response to ethanol / cellular response to zinc ion / regulation of neuron differentiation / neurotransmitter receptor activity / glycine binding / chloride channel complex / ligand-gated monoatomic ion channel activity / transmembrane transporter complex / response to amino acid / monoatomic ion transport / chloride transmembrane transport / 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
GLYCINE / Chem-IVM / Glycine receptor subunit alphaZ1
Similarity search - Component
Biological speciesDanio rerio (zebrafish)
MethodELECTRON MICROSCOPY / single particle reconstruction / cryo EM / Resolution: 3.07 Å
AuthorsKumar, A. / Basak, S. / Chakrapani, S.
Funding support United States, 2items
OrganizationGrant numberCountry
National Institutes of Health/National Institute of General Medical Sciences (NIH/NIGMS)5R01GM108921-05 United States
National Institutes of Health/National Institute of General Medical Sciences (NIH/NIGMS)1R01GM131216-01 United States
Citation
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.
#1: Journal: Nature / Year: 2015
Title: Glycine receptor mechanism elucidated by electron cryo-microscopy.
Authors: Juan Du / Wei Lü / Shenping Wu / Yifan Cheng / Eric Gouaux /
Abstract: The strychnine-sensitive glycine receptor (GlyR) mediates inhibitory synaptic transmission in the spinal cord and brainstem and is linked to neurological disorders, including autism and hyperekplexia. ...The strychnine-sensitive glycine receptor (GlyR) mediates inhibitory synaptic transmission in the spinal cord and brainstem and is linked to neurological disorders, including autism and hyperekplexia. Understanding of molecular mechanisms and pharmacology of glycine receptors has been hindered by a lack of high-resolution structures. Here we report electron cryo-microscopy structures of the zebrafish α1 GlyR with strychnine, glycine, or glycine and ivermectin (glycine/ivermectin). Strychnine arrests the receptor in an antagonist-bound closed ion channel state, glycine stabilizes the receptor in an agonist-bound open channel state, and the glycine/ivermectin complex adopts a potentially desensitized or partially open state. Relative to the glycine-bound state, strychnine expands the agonist-binding pocket via outward movement of the C loop, promotes rearrangement of the extracellular and transmembrane domain 'wrist' interface, and leads to rotation of the transmembrane domain towards the pore axis, occluding the ion conduction pathway. These structures illuminate the GlyR mechanism and define a rubric to interpret structures of Cys-loop receptors.
History
DepositionJan 27, 2020Deposition site: RCSB / Processing site: RCSB
Revision 1.0Jul 29, 2020Provider: repository / Type: Initial release
Revision 1.1Aug 12, 2020Group: Database references / Structure summary / Category: chem_comp / citation / citation_author
Item: _chem_comp.pdbx_synonyms / _citation.journal_volume ..._chem_comp.pdbx_synonyms / _citation.journal_volume / _citation.page_first / _citation.page_last / _citation.pdbx_database_id_PubMed / _citation.title / _citation_author.identifier_ORCID / _citation_author.name
Revision 1.2Mar 6, 2024Group: Data collection / Database references / Category: chem_comp_atom / chem_comp_bond / database_2
Item: _database_2.pdbx_DOI / _database_2.pdbx_database_accession

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Structure visualization

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Assembly

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


Theoretical massNumber of molelcules
Total (without water)260,98120
Polymers254,1095
Non-polymers6,87315
Water00
1


  • Idetical with deposited unit
  • defined by author
  • Evidence: gel filtration
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-GlcpN]{[(2+1)][<C1O1>]{}}}LINUCSPDB-CARE
#3: Chemical
ChemComp-GLY / GLYCINE


Type: peptide linking / Mass: 75.067 Da / Num. of mol.: 5 / Source method: obtained synthetically / Formula: C2H5NO2
#4: Chemical
ChemComp-IVM / (2aE,4E,5'S,6S,6'R,7S,8E,11R,13R,15S,17aR,20R,20aR,20bS)-6'-[(2S)-butan-2-yl]-20,20b-dihydroxy-5',6,8,19-tetramethyl-17 -oxo-3',4',5',6,6',10,11,14,15,17,17a,20,20a,20b-tetradecahydro-2H,7H-spiro[11,15-methanofuro[4,3,2-pq][2,6]benzodioxacy clooctadecine-13,2'-pyran]-7-yl 2,6-dideoxy-4-O-(2,6-dideoxy-3-O-methyl-alpha-L-arabino-hexopyranosyl)-3-O-methyl-alpha-L-arabino-hexopyranoside / 22,23-DIHYDROAVERMECTIN B1A / IVERMECTIN


Mass: 875.093 Da / Num. of mol.: 5 / Source method: obtained synthetically / Formula: C48H74O14 / Feature type: SUBJECT OF INVESTIGATION / Comment: antiparasitic*YM
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 ChlorideNaCl1
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 FIELD / 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 (6k x 4k) / Num. of grids imaged: 8 / Num. of real images: 2389
EM imaging opticsEnergyfilter name: GIF Bioquantum / Energyfilter slit width: 20 eV
Image scansMovie frames/image: 40

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Processing

SoftwareName: PHENIX / Version: 1.17.1_3660: / 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: 3.07 Å / Resolution method: FSC 0.143 CUT-OFF / Num. of particles: 27516 / Num. of class averages: 1 / Symmetry type: POINT
Refine LS restraints
Refine-IDTypeDev idealNumber
ELECTRON MICROSCOPYf_bond_d0.01115565
ELECTRON MICROSCOPYf_angle_d1.48321125
ELECTRON MICROSCOPYf_dihedral_angle_d26.9452070
ELECTRON MICROSCOPYf_chiral_restr0.0792440
ELECTRON MICROSCOPYf_plane_restr0.0072575

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