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- EMDB-46467: Ca2+ bound open-inactivated hSlo1 + beta2N-beta4 channel in deter... -

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Basic information

Entry
Database: EMDB / ID: EMD-46467
TitleCa2+ bound open-inactivated hSlo1 + beta2N-beta4 channel in detergent- Class 2
Map data
Sample
  • Complex: Ca2+ bound BK channel complex of hSlo1 + beta2N-beta4 subunit
    • Complex: Inactivating beta2N-beta4 subunit chimera
      • Protein or peptide: Large-conductance Ca2+-activated K+ channel beta2 subunit,Calcium-activated potassium channel subunit beta-4 chimera
    • Complex: hSlo1 channel
      • Protein or peptide: Isoform 5 of Calcium-activated potassium channel subunit alpha-1
KeywordsPotassium ion channel / calcium and voltage gated ion channel / big potassium channel / human BK / hSlo1 / open-inactivated hSlo1 / ball and chain inactivation / hSlo1 inactivating subunit complex / detergent. / MEMBRANE PROTEIN
Biological speciesHomo sapiens (human)
Methodsingle particle reconstruction / cryo EM / Resolution: 2.96 Å
AuthorsAgarwal S / Nimigean C
Funding support United States, 1 items
OrganizationGrant numberCountry
National Institutes of Health/National Center for Research Resources (NIH/NCRR) United States
CitationJournal: Nat Commun / Year: 2025
Title: Ball-and-chain inactivation of a human large conductance calcium-activated potassium channel.
Authors: Shubhangi Agarwal / Elizabeth D Kim / Sangyun Lee / Alexander Simon / Alessio Accardi / Crina M Nimigean /
Abstract: BK channels are large-conductance calcium (Ca)-activated potassium channels crucial for neuronal excitability, muscle contraction, and neurotransmitter release. The pore-forming (α) subunits co- ...BK channels are large-conductance calcium (Ca)-activated potassium channels crucial for neuronal excitability, muscle contraction, and neurotransmitter release. The pore-forming (α) subunits co-assemble with auxiliary (β and γ) subunits that modulate their function. Previous studies demonstrated that the N-termini of β2-subunits can inactivate BK channels, but with no structural correlate. Here, we investigate BK β2-subunit inactivation using cryo-electron microscopy, electrophysiology and molecular dynamics simulations. We find that the β2 N-terminus occludes the pore only in the Ca-bound open state, via a ball-and-chain mechanism. The first three hydrophobic residues of β2 are crucial for occlusion, while the remainder of the N-terminus remains flexible. Neither the closed channel conformation obtained in the absence of Ca nor an intermediate conformation found in the presence of Ca show density for the N-terminus of the β2 subunit in their pore, likely due to narrower side access portals preventing their entry into the channel pore.
History
DepositionAug 7, 2024-
Header (metadata) releaseMar 5, 2025-
Map releaseMar 5, 2025-
UpdateMar 5, 2025-
Current statusMar 5, 2025Processing site: RCSB / Status: Released

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Structure visualization

Supplemental images

emd_46467.png

Downloads & links

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Map

FileDownload / File: emd_46467.map.gz / Format: CCP4 / Size: 216 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 384 pix.
= 316.8 Å		0.83 Å/pix.
x 384 pix.
= 316.8 Å		0.83 Å/pix.
x 384 pix.
= 316.8 Å

Surface

Projections

Slices (1/3)

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Slices (2/3)

Images are generated by Spider.

Voxel sizeX=Y=Z: 0.825 Å
Density

Histogram

Histogram (log scale)
Contour LevelBy AUTHOR: 0.0143
Minimum - Maximum-0.02222089 - 0.06342679
Average (Standard dev.)0.00018709117 (±0.002763377)
SymmetrySpace group: 1
Details

EMDB XML:

Map geometry
Axis orderXYZ
Origin000
Dimensions384384384
Spacing384384384
CellA=B=C: 316.8 Å
α=β=γ: 90.0 °

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Supplemental data

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Half map: #1

Fileemd_46467_half_map_1.map
Projections & Slices
AxesZYX

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Slices (1/2)
Density Histograms

Histogram

Histogram (log scale)

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Half map: #2

Fileemd_46467_half_map_2.map
Projections & Slices
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Histogram

Histogram (log scale)

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Sample components

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Entire : Ca2+ bound BK channel complex of hSlo1 + beta2N-beta4 subunit

EntireName: Ca2+ bound BK channel complex of hSlo1 + beta2N-beta4 subunit
Components
  • Complex: Ca2+ bound BK channel complex of hSlo1 + beta2N-beta4 subunit
    • Complex: Inactivating beta2N-beta4 subunit chimera
      • Protein or peptide: Large-conductance Ca2+-activated K+ channel beta2 subunit,Calcium-activated potassium channel subunit beta-4 chimera
    • Complex: hSlo1 channel
      • Protein or peptide: Isoform 5 of Calcium-activated potassium channel subunit alpha-1

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Supramolecule #1: Ca2+ bound BK channel complex of hSlo1 + beta2N-beta4 subunit

SupramoleculeName: Ca2+ bound BK channel complex of hSlo1 + beta2N-beta4 subunit
type: complex / ID: 1 / Parent: 0 / Macromolecule list: all
Source (natural)Organism: Homo sapiens (human)

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Supramolecule #2: Inactivating beta2N-beta4 subunit chimera

SupramoleculeName: Inactivating beta2N-beta4 subunit chimera / type: complex / ID: 2 / Parent: 1 / Macromolecule list: #2
Source (natural)Organism: Homo sapiens (human)

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Supramolecule #3: hSlo1 channel

SupramoleculeName: hSlo1 channel / type: complex / ID: 3 / Parent: 1 / Macromolecule list: #1
Source (natural)Organism: Homo sapiens (human)

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Macromolecule #1: Isoform 5 of Calcium-activated potassium channel subunit alpha-1

MacromoleculeName: Isoform 5 of Calcium-activated potassium channel subunit alpha-1
type: protein_or_peptide / ID: 1 / Enantiomer: LEVO
Source (natural)Organism: Homo sapiens (human)
Recombinant expressionOrganism: Homo sapiens (human)
SequenceString: MDALIIPVTM EVPCDSRGQR MWWAFLASSM VTFFGGLFII LLWRTLKYLW TVCCHCGGKT KEAQKINNGS SQADGTLKPV DEKEEAVAA EVGWMTSVKD WAGVMISAQT LTGRVLVVLV FALSIGALVI YFIDSSNPIE SCQNFYKDFT LQIDMAFNVF F LLYFGLRF ...String:
MDALIIPVTM EVPCDSRGQR MWWAFLASSM VTFFGGLFII LLWRTLKYLW TVCCHCGGKT KEAQKINNGS SQADGTLKPV DEKEEAVAA EVGWMTSVKD WAGVMISAQT LTGRVLVVLV FALSIGALVI YFIDSSNPIE SCQNFYKDFT LQIDMAFNVF F LLYFGLRF IAANDKLWFW LEVNSVVDFF TVPPVFVSVY LNRSWLGLRF LRALRLIQFS EILQFLNILK TSNSIKLVNL LS IFISTWL TAAGFIHLVE NSGDPWENFQ NNQALTYWEC VYLLMVTMST VGYGDVYAKT TLGRLFMVFF ILGGLAMFAS YVP EIIELI GNRKKYGGSY SAVSGRKHIV VCGHITLESV SNFLKDFLHK DRDDVNVEIV FLHNISPNLE LEALFKRHFT QVEF YQGSV LNPHDLARVK IESADACLIL ANKYCADPDA EDASNIMRVI SIKNYHPKIR IITQMLQYHN KAHLLNIPSW NWKEG DDAI CLAELKLGFI AQSCLAQGLS TMLANLFSMR SFIKIEEDTW QKYYLEGVSN EMYTEYLSSA FVGLSFPTVC ELCFVK LKL LMIAIEYKSA NRESRILINP GNHLKIQEGT LGFFIASDAK EVKRAFFYCK ACHDDITDPK RIKKCGCKRL EDEQPST LS PKKKQRNGGM RNSPNTSPKL MRHDPLLIPG NDQIDNMDSN VKKYDSTGMF HWCAPKEIEK VILTRSEAAM TVLSGHVV V CIFGDVSSAL IGLRNLVMPL RASNFHYHEL KHIVFVGSIE YLKREWETLH NFPKVSILPG TPLSRADLRA VNINLCDMC VILSANQNNI DDTSLQDKEC ILASLNIKSM QFDDSIGVLQ ANSQGFTPPG MDRSSPDNSP VHGMLRQPSI TTGVNIPIIT ELVNDTNVQ FLDQDDDDDP DTELYLTQPF ACGTAFAVSV LDSLMSATYF NDNILTLIRT LVTGGATPEL EALIAEENAL R GGYSTPQT LANRDRCRVA QLALLDGPFA DLGDGGCYGD LFCKALKTYN MLCFGIYRLR DAHLSTPSQC TKRYVITNPP YE FELVPTD LIFCLMQFD

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Macromolecule #2: Large-conductance Ca2+-activated K+ channel beta2 subunit,Calcium...

MacromoleculeName: Large-conductance Ca2+-activated K+ channel beta2 subunit,Calcium-activated potassium channel subunit beta-4 chimera
type: protein_or_peptide / ID: 2 / Enantiomer: LEVO
Source (natural)Organism: Homo sapiens (human)
Recombinant expressionOrganism: Homo sapiens (human)
SequenceString: FIWTSGRTSS SYRHDEKRNI YQKIRDHDLL DKRKTVTALK AGEDKSIRLG LFLIISGVVS LFIFGFCWLS PALQDLQATE ANCTVLSVQQ IGEVFECTFT CGADCRGTSQ YPCVQVYVNN SESNSRALLH SDEHQLLTNP KCSYIPPCK RENQKNLESV MNWQQYWKDE ...String:
FIWTSGRTSS SYRHDEKRNI YQKIRDHDLL DKRKTVTALK AGEDKSIRLG LFLIISGVVS LFIFGFCWLS PALQDLQATE ANCTVLSVQQ IGEVFECTFT CGADCRGTSQ YPCVQVYVNN SESNSRALLH SDEHQLLTNP KCSYIPPCK RENQKNLESV MNWQQYWKDE IGSQPFTCYF NQHQRPDDVL LHRTHDEIVL LHCFLWPLVT FVVGVLIVV LTICAKSLAV KAEAMKKRKF S

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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

Concentration9 mg/mL
BufferpH: 8
Details: 20 mM Tris-HCl pH 8.0, 450 mM KCl, 5 mM EDTA, 15 mM MgCl2, 0.02% GDN and 0.05 mg/ml POPE:POPC:POPA 5:5:1 (w:w:w).
GridModel: UltrAuFoil R1.2/1.3 / Material: GOLD / Mesh: 300 / Pretreatment - Type: GLOW DISCHARGE / Pretreatment - Time: 80 sec.
VitrificationCryogen name: ETHANE / Chamber humidity: 100 % / Chamber temperature: 288.15 K / Instrument: FEI VITROBOT MARK IV

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Electron microscopy

MicroscopeFEI TITAN KRIOS
Image recordingFilm or detector model: GATAN K3 BIOQUANTUM (6k x 4k) / Average electron dose: 48.45 e/Å2
Electron beamAcceleration voltage: 300 kV / Electron source: FIELD EMISSION GUN
Electron opticsIllumination mode: FLOOD BEAM / Imaging mode: BRIGHT FIELD / Nominal defocus max: 3.0 µm / Nominal defocus min: 0.5 µm
Experimental equipment
Model: Titan Krios / Image courtesy: FEI Company

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Image processing

Startup modelType of model: PDB ENTRY
PDB model - PDB ID:

6v22

Final reconstructionApplied symmetry - Point group: C1 (asymmetric) / Resolution.type: BY AUTHOR / Resolution: 2.96 Å / Resolution method: FSC 0.143 CUT-OFF / Software - Name: RELION (ver. 4.0.0) / Number images used: 32657
Initial angle assignmentType: ANGULAR RECONSTITUTION / Software - Name: RELION (ver. 4.0.0)
Final angle assignmentType: ANGULAR RECONSTITUTION / Software - Name: RELION (ver. 4.0.0)
Final 3D classificationSoftware - Name: RELION (ver. 4.0.0)
FSC plot (resolution estimation)

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Atomic model buiding 1

Initial modelPDB ID:

6v22


Chain - Source name: PDB / Chain - Initial model type: experimental model
RefinementSpace: REAL / Protocol: RIGID BODY FIT

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