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- EMDB-48217: Structure of the human TWIK-2 potassium channel in complex with p... -

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Basic information

Entry
Database: EMDB / ID: EMD-48217
TitleStructure of the human TWIK-2 potassium channel in complex with pimozide
Map dataTWIK2 bound to pimozide
Sample
  • Complex: human TWIK-2 potassium channel in complex with pimozide
    • Protein or peptide: Potassium channel subfamily K member 6
  • Ligand: POTASSIUM ION
  • Ligand: 3-[1-[4,4-bis(4-fluorophenyl)butyl]piperidin-4-yl]-1~{H}-benzimidazol-2-one
KeywordsTwo-pore potassium channel K2P6 TWIK2 K2P channel pimozide drug NLRP3 inflammasome / MEMBRANE PROTEIN
Function / homology
Function and homology information


regulation of lysosome size / Tandem of pore domain in a weak inwardly rectifying K+ channels (TWIK) / Phase 4 - resting membrane potential / potassium ion leak channel activity / regulation of resting membrane potential / inward rectifier potassium channel activity / negative regulation of systemic arterial blood pressure / outward rectifier potassium channel activity / positive regulation of NLRP3 inflammasome complex assembly / potassium channel activity ...regulation of lysosome size / Tandem of pore domain in a weak inwardly rectifying K+ channels (TWIK) / Phase 4 - resting membrane potential / potassium ion leak channel activity / regulation of resting membrane potential / inward rectifier potassium channel activity / negative regulation of systemic arterial blood pressure / outward rectifier potassium channel activity / positive regulation of NLRP3 inflammasome complex assembly / potassium channel activity / voltage-gated potassium channel complex / potassium ion transmembrane transport / potassium ion transport / late endosome membrane / lysosomal membrane / metal ion binding / plasma membrane
Similarity search - Function
Two pore domain potassium channel, TWIK-2 / 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
Potassium channel subfamily K member 6
Similarity search - Component
Biological speciesHomo sapiens (human)
Methodsingle particle reconstruction / cryo EM / Resolution: 3.17 Å
AuthorsKhanra NK / Long SB
Funding support United States, 1 items
OrganizationGrant numberCountry
National Institutes of Health/National Institute of General Medical Sciences (NIH/NIGMS)R35GM131921 United States
CitationJournal: Proc Natl Acad Sci U S A / Year: 2025
Title: Structure of the human TWIK-2 potassium channel and its inhibition by pimozide.
Authors: Nandish K Khanra / Chongyuan Wang / Bryce D Delgado / Stephen B Long /
Abstract: The potassium channel TWIK-2 is crucial for ATP-induced activation of the NLRP3 inflammasome in macrophages. The channel is a member of the two-pore domain potassium (K2P) channel superfamily and an ...The potassium channel TWIK-2 is crucial for ATP-induced activation of the NLRP3 inflammasome in macrophages. The channel is a member of the two-pore domain potassium (K2P) channel superfamily and an emerging therapeutic target to mitigate severe inflammatory injury involving NLRP3 activation. We report the cryo-EM structure of human TWIK-2. In comparison to other K2P channels, the structure reveals an unusual "up" conformation of Tyr111 in the selectivity filter and a resulting SF1-P1 pocket behind the filter. Density for acyl chains is present in fenestrations within the transmembrane region that connects the central cavity of the pore to the lipid membrane. Despite its importance as a drug target, limited pharmacological tools are available for TWIK-2. A previous study suggested that the FDA-approved small molecule pimozide might inhibit TWIK-2. Using a reconstituted system, we show that pimozide directly inhibits the channel and we determine a cryo-EM structure of a complex with the drug. Pimozide displaces the acyl chains within the fenestrations and binds below the selectivity filter where it would impede ion permeation. The drug may access its binding site by lateral diffusion in the membrane, suggesting that other hydrophobic small molecules could have utility for inhibiting TWIK-2. The work defines the structure of TWIK-2 and provides a structural foundation for development of more specific inhibitors with potential utility as anti-inflammatory drugs.
History
DepositionDec 6, 2024-
Header (metadata) releaseApr 23, 2025-
Map releaseApr 23, 2025-
UpdateNov 5, 2025-
Current statusNov 5, 2025Processing site: RCSB / Status: Released

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Structure visualization

Supplemental images

emd_48217.png

Downloads & links

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Map

FileDownload / File: emd_48217.map.gz / Format: CCP4 / Size: 216 MB / Type: IMAGE STORED AS FLOATING POINT NUMBER (4 BYTES)
AnnotationTWIK2 bound to pimozide
Projections & slices

Image control

Size
Brightness
Contrast
Others
AxesZ (Sec.)Y (Row.)X (Col.)
0.73 Å/pix.
x 384 pix.
= 278.4 Å		0.73 Å/pix.
x 384 pix.
= 278.4 Å		0.73 Å/pix.
x 384 pix.
= 278.4 Å

Surface

Projections

Slices (1/3)

Slices (1/2)

Slices (2/3)

Images are generated by Spider.

Voxel sizeX=Y=Z: 0.725 Å
Density

Histogram

Histogram (log scale)
Contour LevelBy AUTHOR: 0.45
Minimum - Maximum-2.758652 - 4.0185432
Average (Standard dev.)-0.0009879048 (±0.049192887)
SymmetrySpace group: 1
Details

EMDB XML:

Map geometry
Axis orderXYZ
Origin000
Dimensions384384384
Spacing384384384
CellA=B=C: 278.40002 Å
α=β=γ: 90.0 °

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Supplemental data

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Half map: TWIK2 bound to pimozide

Fileemd_48217_half_map_1.map
AnnotationTWIK2 bound to pimozide
Projections & Slices
AxesZYX

Projections

Slices (1/2)
Density Histograms

Histogram

Histogram (log scale)

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Half map: TWIK2 bound to pimozide

Fileemd_48217_half_map_2.map
AnnotationTWIK2 bound to pimozide
Projections & Slices
AxesZYX

Projections

Slices (1/2)
Density Histograms

Histogram

Histogram (log scale)

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Sample components

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Entire : human TWIK-2 potassium channel in complex with pimozide

EntireName: human TWIK-2 potassium channel in complex with pimozide
Components
  • Complex: human TWIK-2 potassium channel in complex with pimozide
    • Protein or peptide: Potassium channel subfamily K member 6
  • Ligand: POTASSIUM ION
  • Ligand: 3-[1-[4,4-bis(4-fluorophenyl)butyl]piperidin-4-yl]-1~{H}-benzimidazol-2-one

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Supramolecule #1: human TWIK-2 potassium channel in complex with pimozide

SupramoleculeName: human TWIK-2 potassium channel in complex with pimozide
type: complex / ID: 1 / Parent: 0 / Macromolecule list: #1
Details: human TWIK-2 potassium channel in complex with pimozide
Source (natural)Organism: Homo sapiens (human)
Molecular weightTheoretical: 33.74728 KDa

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Macromolecule #1: Potassium channel subfamily K member 6

MacromoleculeName: Potassium channel subfamily K member 6 / type: protein_or_peptide / ID: 1 / Number of copies: 2 / Enantiomer: LEVO
Source (natural)Organism: Homo sapiens (human)
Molecular weightTheoretical: 33.775113 KDa
Recombinant expressionOrganism: Homo sapiens (human)
SequenceString: MRRGALLAGA LAAYAAYLVL GALLVARLEG PHEARLRAEL ETLRAQLLQR SPCVAAPALD AFVERVLAAG RLGRVVLANA SGSANASDP AWDFASALFF ASTLITTVGY GYTTPLTDAG KAFSIAFALL GVPTTMLLLT ASAQRLSLLL THVPLSWLSM R WGWDPRRA ...String:
MRRGALLAGA LAAYAAYLVL GALLVARLEG PHEARLRAEL ETLRAQLLQR SPCVAAPALD AFVERVLAAG RLGRVVLANA SGSANASDP AWDFASALFF ASTLITTVGY GYTTPLTDAG KAFSIAFALL GVPTTMLLLT ASAQRLSLLL THVPLSWLSM R WGWDPRRA ACWHLVALLG VVVTVCFLVP AVIFAHLEEA WSFLDAFYFC FISLSTIGLG DYVPGEAPGQ PYRALYKVLV TV YLFLGLV AMVLVLQTFR HVSDLHGLTE LILLPPPCPA SFNADEDDRV DILGPQPESH QQLSASSHTD YASIPR

UniProtKB: Potassium channel subfamily K member 6

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Macromolecule #2: POTASSIUM ION

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

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Macromolecule #3: 3-[1-[4,4-bis(4-fluorophenyl)butyl]piperidin-4-yl]-1~{H}-benzimid...

MacromoleculeName: 3-[1-[4,4-bis(4-fluorophenyl)butyl]piperidin-4-yl]-1~{H}-benzimidazol-2-one
type: ligand / ID: 3 / Number of copies: 1 / Formula: 1II
Molecular weightTheoretical: 461.546 Da
Chemical component information

ChemComp-1II:
3-[1-[4,4-bis(4-fluorophenyl)butyl]piperidin-4-yl]-1~{H}-benzimidazol-2-one

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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

BufferpH: 7.5
Component:
ConcentrationFormulaName
20.0 mMHEPES2-[4-(2-hydroxyethyl)piperazin-1-yl]ethanesulfonic acid
150.0 mMKClPotassium Chloride
0.01 %LMNG2,2-didecylpropane-1,3-bis-beta-D-maltopyranoside
0.001 %CHSCholesteryl Hemisuccinate
0.0033 %GDNglyco-diosgenin
0.5 mMPimozide1-[1-[4,4-Bis(4-fluorophenyl)butyl]-4-piperidinyl]-1,3-dihydro-2H-benzimidazol-2-one

Details: 20 mM HEPES, 150 mM KCl, pH 7.5, 0.01 % LMNG, 0.001 % CHS, 0.0033 % GDN, 0.5 mM pimozide
GridModel: Quantifoil R1.2/1.3 / Material: GOLD / Mesh: 400 / Support film - Material: CARBON / Support film - topology: HOLEY
VitrificationCryogen name: ETHANE / Chamber humidity: 90 % / Chamber temperature: 277.15 K / Instrument: FEI VITROBOT MARK IV
Details: 4.0 uL sample, blot time = 3.0 sec, blot force = 0.
DetailsMonodisperse recombinantly purified TWIK2 bound to pimozide

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Electron microscopy

MicroscopeTFS KRIOS
Specialist opticsEnergy filter - Slit width: 20 eV
Image recordingFilm or detector model: TFS FALCON 4i (4k x 4k) / Average electron dose: 51.38 e/Å2
Electron beamAcceleration voltage: 300 kV / Electron source: FIELD EMISSION GUN
Electron opticsIllumination mode: FLOOD BEAM / Imaging mode: BRIGHT FIELD / Cs: 2.7 mm / Nominal defocus max: 2.5 µm / Nominal defocus min: 1.0 µm / Nominal magnification: 165000
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

Particle selectionNumber selected: 6263529
CTF correctionSoftware - Name: RELION (ver. 3.1) / Software - details: CTFFIND 4.1 / Type: PHASE FLIPPING AND AMPLITUDE CORRECTION
Startup modelType of model: OTHER
Final reconstructionApplied symmetry - Point group: C2 (2 fold cyclic) / Resolution.type: BY AUTHOR / Resolution: 3.17 Å / Resolution method: FSC 0.143 CUT-OFF / Number images used: 156956
Initial angle assignmentType: MAXIMUM LIKELIHOOD
Final angle assignmentType: MAXIMUM LIKELIHOOD
FSC plot (resolution estimation)

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Atomic model buiding 1

RefinementOverall B value: 124.2
Output model

PDB-9mel:
Structure of the human TWIK-2 potassium channel in complex with pimozide

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