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- EMDB-50741: Structure of the human two pore domain potassium ion channel THIK... -

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Basic information

Entry
Database: EMDB / ID: EMD-50741
TitleStructure of the human two pore domain potassium ion channel THIK-1 (K2P13.1) in a closed conformation
Map data
Sample
  • Complex: K2P13.1 homodimer
    • Protein or peptide: Potassium channel subfamily K member 13
  • Ligand: LINOLEIC ACID
  • Ligand: POTASSIUM ION
KeywordsPotassium channel / MEMBRANE PROTEIN
Function / homology
Function and homology information


regulation of excitatory synapse pruning / Tandem pore domain halothane-inhibited K+ channel (THIK) / regulation of NLRP3 inflammasome complex assembly / Phase 4 - resting membrane potential / potassium ion leak channel activity / regulation of resting membrane potential / outward rectifier potassium channel activity / monoatomic ion channel complex / potassium channel activity / potassium ion transmembrane transport ...regulation of excitatory synapse pruning / Tandem pore domain halothane-inhibited K+ channel (THIK) / regulation of NLRP3 inflammasome complex assembly / Phase 4 - resting membrane potential / potassium ion leak channel activity / regulation of resting membrane potential / outward rectifier potassium channel activity / monoatomic ion channel complex / potassium channel activity / potassium ion transmembrane transport / protein heterodimerization activity / metal ion binding / identical protein binding / plasma membrane
Similarity search - Function
Two pore domain potassium channel, THIK / Two pore domain potassium channel / Potassium channel domain / Ion channel
Similarity search - Domain/homology
Potassium channel subfamily K member 13
Similarity search - Component
Biological speciesHomo sapiens (human)
Methodsingle particle reconstruction / cryo EM / Resolution: 3.16 Å
AuthorsRodstrom KEJ / Tucker SJ
Funding support United Kingdom, 5 items
OrganizationGrant numberCountry
Biotechnology and Biological Sciences Research Council (BBSRC)BB/T002018/1 United Kingdom
Biotechnology and Biological Sciences Research Council (BBSRC)BB/S008608/1 United Kingdom
Medical Research Council (MRC, United Kingdom)MR/W017741/1 United Kingdom
Wellcome TrustWT084655MA United Kingdom
Wellcome Trust102161/B/13/Z United Kingdom
CitationJournal: Nat Struct Mol Biol / Year: 2025
Title: Cryo-EM structure of the human THIK-1 K2P K channel reveals a lower Y gate regulated by lipids and anesthetics.
Authors: Karin E J Rödström / Bisher Eymsh / Peter Proks / Mehtab S Hayre / Sönke Cordeiro / Edward Mendez-Otalvaro / Christian Madry / Anna Rowland / Wojciech Kopec / Simon Newstead / Thomas ...Authors: Karin E J Rödström / Bisher Eymsh / Peter Proks / Mehtab S Hayre / Sönke Cordeiro / Edward Mendez-Otalvaro / Christian Madry / Anna Rowland / Wojciech Kopec / Simon Newstead / Thomas Baukrowitz / Marcus Schewe / Stephen J Tucker /
Abstract: THIK-1 (KCNK13) is a halothane-inhibited and anionic-lipid-activated two-pore domain (K2P) K channel implicated in microglial activation and neuroinflammation, and a current target for the treatment ...THIK-1 (KCNK13) is a halothane-inhibited and anionic-lipid-activated two-pore domain (K2P) K channel implicated in microglial activation and neuroinflammation, and a current target for the treatment of neurodegenerative disorders, for example Alzheimer's disease and amyothropic lateral sclerosis (ALS). However, compared to other K2P channels, little is known about the structural and functional properties of THIK-1. Here we present a 3.16-Å-resolution cryo-EM structure of human THIK-1 that reveals several distinct features, in particular, a tyrosine in M4 that contributes to a lower 'Y gate' that opens upon activation by physiologically relevant G-protein-coupled receptor and lipid signaling pathways. We demonstrate that linoleic acid bound within a modulatory pocket adjacent to the filter influences channel activity, and that halothane inhibition involves a binding site within the inner cavity, both resulting in conformational changes to the Y gate. Finally, the extracellular cap domain contains positively charged residues that line the ion exit pathway and contribute to the distinct biophysical properties of this channel. Overall, our results provide structural insights into THIK-1 function and identify distinct regulatory sites that expand its potential as a drug target for the modulation of microglial function.
History
DepositionJun 24, 2024-
Header (metadata) releaseMar 5, 2025-
Map releaseMar 5, 2025-
UpdateMar 5, 2025-
Current statusMar 5, 2025Processing site: PDBe / Status: Released

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Structure visualization

Supplemental images

emd_50741.png

Downloads & links

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Map

FileDownload / File: emd_50741.map.gz / Format: CCP4 / Size: 64 MB / Type: IMAGE STORED AS FLOATING POINT NUMBER (4 BYTES)
Projections & slices

Image control

Size
Brightness
Contrast
Others
AxesZ (Sec.)Y (Row.)X (Col.)
0.83 Å/pix.
x 256 pix.
= 212.992 Å		0.83 Å/pix.
x 256 pix.
= 212.992 Å		0.83 Å/pix.
x 256 pix.
= 212.992 Å

Surface

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Slices (2/3)

Images are generated by Spider.

Voxel sizeX=Y=Z: 0.832 Å
Density

Histogram

Histogram (log scale)
Contour LevelBy AUTHOR: 0.25
Minimum - Maximum-1.6273953 - 2.1717148
Average (Standard dev.)0.0006606604 (±0.043420766)
SymmetrySpace group: 1
Details

EMDB XML:

Map geometry
Axis orderXYZ
Origin000
Dimensions256256256
Spacing256256256
CellA=B=C: 212.992 Å
α=β=γ: 90.0 °

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Supplemental data

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Mask #1

Fileemd_50741_msk_1.map
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AxesZYX

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Histogram

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Mask #2

Fileemd_50741_msk_2.map
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Half map: Half map, A

Fileemd_50741_half_map_1.map
AnnotationHalf map, A
Projections & Slices
AxesZYX

Projections

Slices (1/2)
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Histogram

Histogram (log scale)

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Half map: Half map, B

Fileemd_50741_half_map_2.map
AnnotationHalf map, B
Projections & Slices
AxesZYX

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Histogram

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Sample components

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Entire : K2P13.1 homodimer

EntireName: K2P13.1 homodimer
Components
  • Complex: K2P13.1 homodimer
    • Protein or peptide: Potassium channel subfamily K member 13
  • Ligand: LINOLEIC ACID
  • Ligand: POTASSIUM ION

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Supramolecule #1: K2P13.1 homodimer

SupramoleculeName: K2P13.1 homodimer / type: complex / ID: 1 / Parent: 0 / Macromolecule list: #1
Details: Protein generated by removal of the 10xHis and FLAG purification tags with HRV 3C protease cleavage
Source (natural)Organism: Homo sapiens (human)
Molecular weightTheoretical: 670 KDa

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Macromolecule #1: Potassium channel subfamily K member 13

MacromoleculeName: Potassium channel subfamily K member 13 / type: protein_or_peptide / ID: 1 / Details: K2P13.1 residues G9-G297 / Number of copies: 2 / Enantiomer: LEVO
Source (natural)Organism: Homo sapiens (human)
Molecular weightTheoretical: 33.63798 KDa
Recombinant expressionOrganism: Spodoptera frugiperda (fall armyworm)
SequenceString: MGPGHLNEDN ARFLLLAALI VLYLLGGAAV FSALELAHER QAKQRWEERL ANFSRGHNLS RDELRGFLRH YEEATRAGIR VDNVRPRWD FTGAFYFVGT VVSTIGFGMT TPATVGGKIF LIFYGLVGCS STILFFNLFL ERLITIIAYI MKSCHQRQLR R RGALPQES ...String:
MGPGHLNEDN ARFLLLAALI VLYLLGGAAV FSALELAHER QAKQRWEERL ANFSRGHNLS RDELRGFLRH YEEATRAGIR VDNVRPRWD FTGAFYFVGT VVSTIGFGMT TPATVGGKIF LIFYGLVGCS STILFFNLFL ERLITIIAYI MKSCHQRQLR R RGALPQES LKDAGQCEVD SLAGWKPSVY YVMLILCTAS ILISCCASAM YTPIEGWSYF DSLYFCFVAF STIGFGDLVS SQ NAHYESQ GLYRFANFVF ILMGVCCIYS LFNVISILIK QSLNWILRKM DSGAELEVLF Q

UniProtKB: Potassium channel subfamily K member 13

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Macromolecule #2: LINOLEIC ACID

MacromoleculeName: LINOLEIC ACID / type: ligand / ID: 2 / Number of copies: 2 / Formula: EIC
Molecular weightTheoretical: 280.445 Da
Chemical component information

ChemComp-EIC:
LINOLEIC ACID

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Macromolecule #3: POTASSIUM ION

MacromoleculeName: POTASSIUM ION / type: ligand / ID: 3 / Number of copies: 2 / Formula: K
Molecular weightTheoretical: 39.098 Da

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Experimental details

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Structure determination

Methodcryo EM
Processingsingle particle reconstruction
Aggregation stateparticle

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Sample preparation

Concentration4 mg/mL
BufferpH: 7.5
Component:
ConcentrationFormulaName
20.0 mMC8H18N2O4S4-(2-Hydroxyethyl)piperazine-1-ethanesulfonic acid
200.0 mMKClpotassium chloride
0.12 % w/vC27H52O12octyl glucose neopentyl glycol
0.012 % w/vC31H50O4C4H11NO3cholesteryl hemisuccinate tris salt

Details: 20 mM HEPES pH 7.5, 200 mM KCl, 0.12% w/v OGNG, 0.012% w/v CHS, pH was adjusted using potassium hydroxide
GridModel: Au-flat 1.2/1.3 / Material: GOLD / Mesh: 300 / Support film - Material: GOLD / Support film - topology: HOLEY / Pretreatment - Type: GLOW DISCHARGE / Pretreatment - Time: 120 sec.
VitrificationCryogen name: ETHANE / Chamber humidity: 100 % / Chamber temperature: 277 K / Instrument: FEI VITROBOT MARK IV / Details: Grid blotted for 4 seconds.
DetailsMonodisperse sample.

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Electron microscopy

MicroscopeFEI TITAN KRIOS
Specialist opticsEnergy filter - Name: GIF Bioquantum / Energy filter - Slit width: 20 eV
Image recordingFilm or detector model: GATAN K3 BIOQUANTUM (6k x 4k) / Number grids imaged: 1 / Number real images: 21171 / Average electron dose: 42.54 e/Å2
Electron beamAcceleration voltage: 300 kV / Electron source: FIELD EMISSION GUN
Electron opticsIllumination mode: FLOOD BEAM / Imaging mode: BRIGHT FIELD / Nominal defocus max: 2.3000000000000003 µm / Nominal defocus min: 1.0 µm / Nominal magnification: 105000
Sample stageSpecimen holder model: FEI TITAN KRIOS AUTOGRID HOLDER / Cooling holder cryogen: NITROGEN
Experimental equipment
Model: Titan Krios / Image courtesy: FEI Company

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Image processing

Startup modelType of model: NONE / Details: Ab-initio
Final reconstructionApplied symmetry - Point group: C2 (2 fold cyclic) / Resolution.type: BY AUTHOR / Resolution: 3.16 Å / Resolution method: FSC 0.143 CUT-OFF / Software - Name: cryoSPARC (ver. 4.4.0) / Number images used: 302189
Initial angle assignmentType: MAXIMUM LIKELIHOOD / Software - Name: cryoSPARC (ver. 4.4.0)
Final angle assignmentType: MAXIMUM LIKELIHOOD / Software - Name: cryoSPARC (ver. 4.4.0)
FSC plot (resolution estimation)

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Atomic model buiding 1

DetailsAn initial model was built manually in COOT
RefinementSpace: REAL / Protocol: FLEXIBLE FIT
Output model

PDB-9ft7:
Structure of the human two pore domain potassium ion channel THIK-1 (K2P13.1) in a closed conformation

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