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- EMDB-43131: Voltage gated potassium ion channel Kv1.2 in complex with DTx -

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Basic information

Entry
Database: EMDB / ID: EMD-43131
TitleVoltage gated potassium ion channel Kv1.2 in complex with DTx
Map dataKv1.2 DTX bound map
Sample
  • Cell: Voltage gated potassium channel Kv1.2 with DTx bound
    • Protein or peptide: Kunitz-type serine protease inhibitor homolog alpha-dendrotoxin
    • Protein or peptide: Potassium voltage-gated channel subfamily A member 2
  • Ligand: POTASSIUM ION
KeywordsIon Channel / Toxin bound / Blocker / MEMBRANE PROTEIN
Function / homology
Function and homology information


optic nerve structural organization / Voltage gated Potassium channels / potassium channel complex / voltage-gated monoatomic ion channel activity involved in regulation of postsynaptic membrane potential / regulation of circadian sleep/wake cycle, non-REM sleep / paranodal junction / potassium ion export across plasma membrane / corpus callosum development / delayed rectifier potassium channel activity / voltage-gated monoatomic ion channel activity involved in regulation of presynaptic membrane potential ...optic nerve structural organization / Voltage gated Potassium channels / potassium channel complex / voltage-gated monoatomic ion channel activity involved in regulation of postsynaptic membrane potential / regulation of circadian sleep/wake cycle, non-REM sleep / paranodal junction / potassium ion export across plasma membrane / corpus callosum development / delayed rectifier potassium channel activity / voltage-gated monoatomic ion channel activity involved in regulation of presynaptic membrane potential / axon initial segment / optic nerve development / juxtaparanode region of axon / outward rectifier potassium channel activity / neuronal cell body membrane / regulation of dopamine secretion / lamellipodium membrane / action potential / voltage-gated potassium channel activity / kinesin binding / potassium channel regulator activity / neuronal action potential / voltage-gated potassium channel complex / axon terminus / potassium ion transmembrane transport / sensory perception of pain / calyx of Held / serine-type endopeptidase inhibitor activity / protein homooligomerization / cerebral cortex development / lamellipodium / toxin activity / presynaptic membrane / perikaryon / postsynaptic membrane / endosome / axon / dendrite / endoplasmic reticulum membrane / glutamatergic synapse / extracellular space / membrane / plasma membrane
Similarity search - Function
Potassium channel, voltage dependent, Kv1.2 / Potassium channel, voltage dependent, Kv1 / Potassium channel, voltage dependent, Kv / : / Potassium channel tetramerisation-type BTB domain / BTB/POZ domain / Voltage-gated potassium channel / Proteinase inhibitor I2, Kunitz, conserved site / Pancreatic trypsin inhibitor (Kunitz) family signature. / BPTI/Kunitz family of serine protease inhibitors. ...Potassium channel, voltage dependent, Kv1.2 / Potassium channel, voltage dependent, Kv1 / Potassium channel, voltage dependent, Kv / : / Potassium channel tetramerisation-type BTB domain / BTB/POZ domain / Voltage-gated potassium channel / Proteinase inhibitor I2, Kunitz, conserved site / Pancreatic trypsin inhibitor (Kunitz) family signature. / BPTI/Kunitz family of serine protease inhibitors. / Pancreatic trypsin inhibitor Kunitz domain / Kunitz/Bovine pancreatic trypsin inhibitor domain / Pancreatic trypsin inhibitor (Kunitz) family profile. / Pancreatic trypsin inhibitor Kunitz domain superfamily / Broad-Complex, Tramtrack and Bric a brac / BTB/POZ domain / Voltage-dependent channel domain superfamily / SKP1/BTB/POZ domain superfamily / Ion transport domain / Ion transport protein
Similarity search - Domain/homology
Kunitz-type serine protease inhibitor homolog alpha-dendrotoxin / Potassium voltage-gated channel subfamily A member 2
Similarity search - Component
Biological speciesKomagataella pastoris (fungus) / Dendroaspis angusticeps (eastern green mamba) / Rattus norvegicus (Norway rat)
Methodsingle particle reconstruction / cryo EM / Resolution: 3.2 Å
AuthorsWu Y / Sigworth FJ
Funding support United States, 1 items
OrganizationGrant numberCountry
National Institutes of Health/National Center for Research Resources (NIH/NCRR)GR108615 United States
CitationJournal: bioRxiv / Year: 2024
Title: Cryo-EM structures of Kv1.2 potassium channels, conducting and non-conducting.
Authors: Yangyu Wu / Yangyang Yan / Youshan Yang / Shumin Bian / Alberto Rivetta / Ken Allen / Fred J Sigworth /
Abstract: We present near-atomic-resolution cryo-EM structures of the mammalian voltage-gated potassium channel Kv1.2 in open, C-type inactivated, toxin-blocked and sodium-bound states at 3.2 Å, 2.5 Å, 3.2 ...We present near-atomic-resolution cryo-EM structures of the mammalian voltage-gated potassium channel Kv1.2 in open, C-type inactivated, toxin-blocked and sodium-bound states at 3.2 Å, 2.5 Å, 3.2 Å, and 2.9Å. These structures, all obtained at nominally zero membrane potential in detergent micelles, reveal distinct ion-occupancy patterns in the selectivity filter. The first two structures are very similar to those reported in the related Shaker channel and the much-studied Kv1.2-2.1 chimeric channel. On the other hand, two new structures show unexpected patterns of ion occupancy. First, the toxin α-Dendrotoxin, like Charybdotoxin, is seen to attach to the negatively-charged channel outer mouth, and a lysine residue penetrates into the selectivity filter, with the terminal amine coordinated by carbonyls, partially disrupting the outermost ion-binding site. In the remainder of the filter two densities of bound ions are observed, rather than three as observed with other toxin-blocked Kv channels. Second, a structure of Kv1.2 in Na solution does not show collapse or destabilization of the selectivity filter, but instead shows an intact selectivity filter with ion density in each binding site. We also attempted to image the C-type inactivated Kv1.2 W366F channel in Na solution, but the protein conformation was seen to be highly variable and only a low-resolution structure could be obtained. These findings present new insights into the stability of the selectivity filter and the mechanism of toxin block of this intensively studied, voltage-gated potassium channel.
History
DepositionDec 13, 2023-
Header (metadata) releaseJul 10, 2024-
Map releaseJul 10, 2024-
UpdateOct 9, 2024-
Current statusOct 9, 2024Processing site: RCSB / Status: Released

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Structure visualization

Supplemental images

emd_43131.png

Downloads & links

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Map

FileDownload / File: emd_43131.map.gz / Format: CCP4 / Size: 64 MB / Type: IMAGE STORED AS FLOATING POINT NUMBER (4 BYTES)
AnnotationKv1.2 DTX bound map
Projections & slices

Image control

Size
Brightness
Contrast
Others
AxesZ (Sec.)Y (Row.)X (Col.)
1.07 Å/pix.
x 256 pix.
= 273.408 Å		1.07 Å/pix.
x 256 pix.
= 273.408 Å		1.07 Å/pix.
x 256 pix.
= 273.408 Å

Surface

Projections

Slices (1/3)

Slices (1/2)

Slices (2/3)

Images are generated by Spider.

Voxel sizeX=Y=Z: 1.068 Å
Density

Histogram

Histogram (log scale)
Contour LevelBy AUTHOR: 0.843
Minimum - Maximum-5.9528065 - 7.4049854
Average (Standard dev.)0.0038585216 (±0.1958854)
SymmetrySpace group: 1
Details

EMDB XML:

Map geometry
Axis orderXYZ
Origin000
Dimensions256256256
Spacing256256256
CellA=B=C: 273.408 Å
α=β=γ: 90.0 °

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Supplemental data

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Half map: Kv1.2 DTX bound half map

Fileemd_43131_half_map_1.map
AnnotationKv1.2 DTX bound half map
Projections & Slices
AxesZYX

Projections

Slices (1/2)
Density Histograms

Histogram

Histogram (log scale)

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Half map: Kv1.2 DTX bound half map

Fileemd_43131_half_map_2.map
AnnotationKv1.2 DTX bound half map
Projections & Slices
AxesZYX

Projections

Slices (1/2)
Density Histograms

Histogram

Histogram (log scale)

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Sample components

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Entire : Voltage gated potassium channel Kv1.2 with DTx bound

EntireName: Voltage gated potassium channel Kv1.2 with DTx bound
Components
  • Cell: Voltage gated potassium channel Kv1.2 with DTx bound
    • Protein or peptide: Kunitz-type serine protease inhibitor homolog alpha-dendrotoxin
    • Protein or peptide: Potassium voltage-gated channel subfamily A member 2
  • Ligand: POTASSIUM ION

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Supramolecule #1: Voltage gated potassium channel Kv1.2 with DTx bound

SupramoleculeName: Voltage gated potassium channel Kv1.2 with DTx bound / type: cell / ID: 1 / Parent: 0 / Macromolecule list: #1-#2
Source (natural)Organism: Komagataella pastoris (fungus) / Strain: SMD1168

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Macromolecule #1: Kunitz-type serine protease inhibitor homolog alpha-dendrotoxin

MacromoleculeName: Kunitz-type serine protease inhibitor homolog alpha-dendrotoxin
type: protein_or_peptide / ID: 1 / Number of copies: 1 / Enantiomer: LEVO
Source (natural)Organism: Dendroaspis angusticeps (eastern green mamba)
Molecular weightTheoretical: 7.015129 KDa
SequenceString:
GPRRKLCILH RNPGRCYDKI PAFYYNQKKK QCERFDWSGC GGNSNRFKTI EECRRTCIG

UniProtKB: Kunitz-type serine protease inhibitor homolog alpha-dendrotoxin

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Macromolecule #2: Potassium voltage-gated channel subfamily A member 2

MacromoleculeName: Potassium voltage-gated channel subfamily A member 2 / type: protein_or_peptide / ID: 2 / Number of copies: 4 / Enantiomer: LEVO
Source (natural)Organism: Rattus norvegicus (Norway rat)
Molecular weightTheoretical: 60.59157 KDa
Recombinant expressionOrganism: Komagataella pastoris (fungus)
SequenceString: MSAWSHPQFE KGGGSGGGSG GSAWSHPQFE KLVPRGSMTV ATGDPVDEAA AHPGHPQDTY DPEADHECCE RVVINISGLR FETQLKTLA QFPETLLGDP KKRMRYFDPL RNEYFFDRNR PSFDAILYYY QSGGRLRRPV NVPLDIFSEE IRFYELGEEA M EMFREDEG ...String:
MSAWSHPQFE KGGGSGGGSG GSAWSHPQFE KLVPRGSMTV ATGDPVDEAA AHPGHPQDTY DPEADHECCE RVVINISGLR FETQLKTLA QFPETLLGDP KKRMRYFDPL RNEYFFDRNR PSFDAILYYY QSGGRLRRPV NVPLDIFSEE IRFYELGEEA M EMFREDEG YIKEEERPLP ENEFQRQVWL LFEYPESSGP ARIIAIVSVM VILISIVSFC LETLPIFRDE NEDMHGSGVT FH TYSQSTI GYQQSTSFTD PFFIVETLCI IWFSFEFLVR FFACPSKAGF FTNIMNIIDI VAIIPYFITL GTELAEKPED AQQ GQQAMS LAILRVIRLV RVFRIFKLSR HSKGLQILGQ TLKASMRELG LLIFFLFIGV ILFSSAVYFA EADERDSQFP SIPD AFWWA VVSMTTVGYG DMVPTTIGGK IVGSLCAIAG VLTIALPVPV IVSNFNYFYH RETEGEEQAQ YLQVTSCPKI PSSPD LKKS RSASTISKSD YMEIQEGVNN SNEDFREENL KTANCTLANT NYVNITKMLT DV

UniProtKB: Potassium voltage-gated channel subfamily A member 2

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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

Concentration0.1 mg/mL
BufferpH: 7.4
VitrificationCryogen name: ETHANE

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Electron microscopy

MicroscopeFEI TITAN KRIOS
Image recordingFilm or detector model: GATAN K3 (6k x 4k) / Average electron dose: 50.0 e/Å2
Electron beamAcceleration voltage: 300 kV / Electron source: FIELD EMISSION GUN
Electron opticsIllumination mode: FLOOD BEAM / Imaging mode: BRIGHT FIELD / Nominal defocus max: 2.0 µm / Nominal defocus min: 1.0 µm
Experimental equipment
Model: Titan Krios / Image courtesy: FEI Company

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Image processing

Startup modelType of model: NONE
Final reconstructionResolution.type: BY AUTHOR / Resolution: 3.2 Å / Resolution method: FSC 0.143 CUT-OFF / Number images used: 312202
Initial angle assignmentType: ANGULAR RECONSTITUTION
Final angle assignmentType: ANGULAR RECONSTITUTION
FSC plot (resolution estimation)

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