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- PDB-7sk0: TWIK1 in MSP1D1 lipid nanodisc at pH 7.4 -

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Basic information

Entry
Database: PDB / ID: 7sk0
TitleTWIK1 in MSP1D1 lipid nanodisc at pH 7.4
ComponentsPotassium channel subfamily K member 1
KeywordsTRANSPORT PROTEIN / K+ ion Channel / pH
Function / homology
Function and homology information


Tandem of pore domain in a weak inwardly rectifying K+ channels (TWIK) / Phase 4 - resting membrane potential / inward rectifier potassium channel complex / potassium channel complex / stabilization of membrane potential / regulation of resting membrane potential / potassium ion leak channel activity / inward rectifier potassium channel activity / sodium channel activity / anchoring junction ...Tandem of pore domain in a weak inwardly rectifying K+ channels (TWIK) / Phase 4 - resting membrane potential / inward rectifier potassium channel complex / potassium channel complex / stabilization of membrane potential / regulation of resting membrane potential / potassium ion leak channel activity / inward rectifier potassium channel activity / sodium channel activity / anchoring junction / plasma membrane => GO:0005886 / potassium channel activity / sodium ion transmembrane transport / potassium ion transmembrane transport / response to nicotine / synaptic membrane / brush border membrane / recycling endosome / perikaryon / membrane => GO:0016020 / endosome / apical plasma membrane / dendrite / identical protein binding / cytoplasm
Similarity search - Function
Two pore domain potassium channel, TWIK-1 / Two pore domain potassium channel, TWIK family / Two pore domain potassium channel, TASK family / Two pore domain potassium channel / Potassium channel domain / Ion channel
Similarity search - Domain/homology
: / N-OCTANE / Potassium channel subfamily K member 1
Similarity search - Component
Biological speciesRattus norvegicus (Norway rat)
MethodELECTRON MICROSCOPY / single particle reconstruction / cryo EM / Resolution: 3.33 Å
AuthorsTurney, T.S. / Brohawn, S.G.
Funding support United States, 2items
OrganizationGrant numberCountry
Other private United States
National Institutes of Health/National Institute of General Medical Sciences (NIH/NIGMS)GM123496 United States
Citation
Journal: Nat Commun / Year: 2022
Title: Structural Basis for pH-gating of the K channel TWIK1 at the selectivity filter.
Authors: Toby S Turney / Vivian Li / Stephen G Brohawn /
Abstract: TWIK1 (K2P1.1, KCNK1) is a widely expressed pH-gated two-pore domain K channel (K2P) that contributes to cardiac rhythm generation and insulin release from pancreatic beta cells. TWIK1 displays ...TWIK1 (K2P1.1, KCNK1) is a widely expressed pH-gated two-pore domain K channel (K2P) that contributes to cardiac rhythm generation and insulin release from pancreatic beta cells. TWIK1 displays unique properties among K2Ps including low basal activity and inhibition by extracellular protons through incompletely understood mechanisms. Here, we present cryo-EM structures of TWIK1 in lipid nanodiscs at high and low pH that reveal a previously undescribed gating mechanism at the K selectivity filter. At high pH, TWIK1 adopts an open conformation. At low pH, protonation of an extracellular histidine results in a cascade of conformational changes that close the channel by sealing the top of the selectivity filter, displacing the helical cap to block extracellular ion access pathways, and opening gaps for lipid block of the intracellular cavity. These data provide a mechanistic understanding for extracellular pH-gating of TWIK1 and illustrate how diverse mechanisms have evolved to gate the selectivity filter of K channels.
#1: Journal: Biorxiv / Year: 2021
Title: Structural Basis for pH-Gating of the K + Channel TWIK1 at the Selectivity Filter
Authors: Turney, T.S. / Li, V. / Brohawn, S.G.
History
DepositionOct 19, 2021Deposition site: RCSB / Processing site: RCSB
Revision 1.0Nov 24, 2021Provider: repository / Type: Initial release
Revision 1.1Jun 22, 2022Group: Database references / Category: citation / citation_author

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Assembly

Deposited unit
A: Potassium channel subfamily K member 1
B: Potassium channel subfamily K member 1
hetero molecules


Theoretical massNumber of molelcules
Total (without water)76,9569
Polymers76,5322
Non-polymers4247
Water0
1


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

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Components

#1: Protein Potassium channel subfamily K member 1 / Inward rectifying potassium channel protein TWIK-1 / rTWIK / Potassium channel K2P1


Mass: 38265.984 Da / Num. of mol.: 2
Source method: isolated from a genetically manipulated source
Source: (gene. exp.) Rattus norvegicus (Norway rat) / Gene: Kcnk1 / Production host: Komagataella pastoris (fungus) / References: UniProt: Q9Z2T2
#2: Chemical ChemComp-OCT / N-OCTANE / Octane


Mass: 114.229 Da / Num. of mol.: 2 / Source method: obtained synthetically / Formula: C8H18
#3: Chemical
ChemComp-K / POTASSIUM ION


Mass: 39.098 Da / Num. of mol.: 5 / Source method: obtained synthetically / Formula: K / 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: TWIK1 in MSP1D1 Lipid Nanodisc at pH 7.4 / Type: COMPLEX / Entity ID: #1 / Source: RECOMBINANT
Source (natural)Organism: Rattus norvegicus (Norway rat)
Source (recombinant)Organism: Komagataella pastoris (fungus)
Buffer solutionpH: 7.4
Buffer component
IDConc.NameFormulaBuffer-ID
120 mMHEPESC8H18N2O4S1
2150 mMPotassium ChlorideKCl1
31 mMEDTAEthylenediaminetetraacetic acidC10H16N2O81
SpecimenEmbedding applied: NO / Shadowing applied: NO / Staining applied: NO / Vitrification applied: YES
Specimen supportGrid type: Quantifoil R1.2/1.3
VitrificationInstrument: FEI VITROBOT MARK III / Cryogen name: ETHANE / Humidity: 100 % / Chamber temperature: 277 K

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Electron microscopy imaging

Experimental equipment
Model: Talos Arctica / Image courtesy: FEI Company
MicroscopyModel: FEI TALOS ARCTICA
Electron gunElectron source: FIELD EMISSION GUN / Accelerating voltage: 200 kV / Illumination mode: FLOOD BEAM
Electron lensMode: BRIGHT FIELDBright-field microscopy
Image recordingElectron dose: 49.715 e/Å2 / Film or detector model: GATAN K3 (6k x 4k)

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Processing

SoftwareName: PHENIX / Version: 1.18.2_3874: / Classification: refinement
EM software
IDNameVersionCategory
1RELION3.1particle selection
4CTFFIND4CTF correction
7Cootmodel fitting
9cryoSPARC2initial Euler assignment
10RELION3.1final Euler assignment
11RELION3.1classification
12RELION3.13D reconstruction
13PHENIXmodel refinement
CTF correctionType: PHASE FLIPPING AND AMPLITUDE CORRECTION
Particle selectionNum. of particles selected: 2872868
SymmetryPoint symmetry: C2 (2 fold cyclic)
3D reconstructionResolution: 3.33 Å / Resolution method: FSC 0.143 CUT-OFF / Num. of particles: 92438 / Symmetry type: POINT
Atomic model buildingB value: 147.12 / Space: REAL / Target criteria: Correlation coefficient
Atomic model buildingPDB-ID: 3UKM
Refine LS restraints
Refine-IDTypeDev idealNumber
ELECTRON MICROSCOPYf_bond_d0.0094264
ELECTRON MICROSCOPYf_angle_d1.0455790
ELECTRON MICROSCOPYf_dihedral_angle_d11.388558
ELECTRON MICROSCOPYf_chiral_restr0.058658
ELECTRON MICROSCOPYf_plane_restr0.008712

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