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- EMDB-21028: Cryo-EM structure of Ca2+-free hsSlo1-beta4 channel complex -

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Basic information

Entry
Database: EMDB / ID: EMD-21028
TitleCryo-EM structure of Ca2+-free hsSlo1-beta4 channel complex
Map dataB factor-sharpened Ca2 -free hsSlo1-beta4 channel complex
Sample
  • Complex: hsSlo1 alpha subunit-beta4 subunit complex
    • Protein or peptide: Calcium-activated potassium channel subunit alpha-1
    • Protein or peptide: Calcium-activated potassium channel subunit beta-4
  • Ligand: (1R)-2-{[(S)-{[(2S)-2,3-dihydroxypropyl]oxy}(hydroxy)phosphoryl]oxy}-1-[(hexadecanoyloxy)methyl]ethyl (9Z)-octadec-9-enoate
  • Ligand: CHOLESTEROL
  • Ligand: 2-acetamido-2-deoxy-beta-D-glucopyranose
KeywordsHigh conductance Ca2+-activated K+ channel / Slo1 channel / BK channel / MaxiK channel / Slo1 beta subunit / beta4 subunit / TRANSPORT PROTEIN
Function / homology
Function and homology information


Acetylcholine inhibits contraction of outer hair cells / micturition / Ca2+ activated K+ channels / large conductance calcium-activated potassium channel activity / response to carbon monoxide / calcium-activated potassium channel activity / negative regulation of cell volume / regulation of neurotransmitter secretion / voltage-gated monoatomic ion channel activity involved in regulation of presynaptic membrane potential / smooth muscle contraction involved in micturition ...Acetylcholine inhibits contraction of outer hair cells / micturition / Ca2+ activated K+ channels / large conductance calcium-activated potassium channel activity / response to carbon monoxide / calcium-activated potassium channel activity / negative regulation of cell volume / regulation of neurotransmitter secretion / voltage-gated monoatomic ion channel activity involved in regulation of presynaptic membrane potential / smooth muscle contraction involved in micturition / intracellular potassium ion homeostasis / Sensory processing of sound by inner hair cells of the cochlea / response to osmotic stress / cGMP effects / action potential / voltage-gated potassium channel activity / detection of calcium ion / regulation of vasoconstriction / potassium channel regulator activity / neuronal action potential / voltage-gated potassium channel complex / potassium ion transmembrane transport / regulation of membrane potential / potassium ion transport / caveola / response to calcium ion / vasodilation / actin binding / chemical synaptic transmission / postsynaptic membrane / response to hypoxia / positive regulation of apoptotic process / apical plasma membrane / synapse / identical protein binding / membrane / metal ion binding / plasma membrane
Similarity search - Function
Potassium channel, calcium-activated, BK, beta subunit / Calcium-activated potassium channel, beta subunit / : / Ca2+-activated K+ channel Slowpoke, TrkA_C like domain / : / Calcium-activated potassium channel BK, alpha subunit / : / Calcium-activated BK potassium channel alpha subunit / Calcium-activated potassium channel slowpoke-like RCK domain / RCK N-terminal domain profile. ...Potassium channel, calcium-activated, BK, beta subunit / Calcium-activated potassium channel, beta subunit / : / Ca2+-activated K+ channel Slowpoke, TrkA_C like domain / : / Calcium-activated potassium channel BK, alpha subunit / : / Calcium-activated BK potassium channel alpha subunit / Calcium-activated potassium channel slowpoke-like RCK domain / RCK N-terminal domain profile. / Ion transport domain / Ion transport protein / NAD(P)-binding domain superfamily
Similarity search - Domain/homology
Calcium-activated potassium channel subunit alpha-1 / Calcium-activated potassium channel subunit beta-4
Similarity search - Component
Biological speciesHomo sapiens (human)
Methodsingle particle reconstruction / cryo EM / Resolution: 3.5 Å
AuthorsTao X / MacKinnon R
Funding support United States, 1 items
OrganizationGrant numberCountry
National Institutes of Health/National Institute of General Medical Sciences (NIH/NIGMS)GM43949 United States
CitationJournal: Elife / Year: 2019
Title: Molecular structures of the human Slo1 K channel in complex with β4.
Authors: Xiao Tao / Roderick MacKinnon /
Abstract: Slo1 is a Ca- and voltage-activated K channel that underlies skeletal and smooth muscle contraction, audition, hormone secretion and neurotransmitter release. In mammals, Slo1 is regulated by ...Slo1 is a Ca- and voltage-activated K channel that underlies skeletal and smooth muscle contraction, audition, hormone secretion and neurotransmitter release. In mammals, Slo1 is regulated by auxiliary proteins that confer tissue-specific gating and pharmacological properties. This study presents cryo-EM structures of Slo1 in complex with the auxiliary protein, β4. Four β4, each containing two transmembrane helices, encircle Slo1, contacting it through helical interactions inside the membrane. On the extracellular side, β4 forms a tetrameric crown over the pore. Structures with high and low Ca concentrations show that identical gating conformations occur in the absence and presence of β4, implying that β4 serves to modulate the relative stabilities of 'pre-existing' conformations rather than creating new ones. The effects of β4 on scorpion toxin inhibition kinetics are explained by the crown, which constrains access but does not prevent binding.
History
DepositionNov 25, 2019-
Header (metadata) releaseDec 25, 2019-
Map releaseDec 25, 2019-
UpdateOct 30, 2024-
Current statusOct 30, 2024Processing site: RCSB / Status: Released

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Structure visualization

Movie
  • Surface view with section colored by density value
  • Surface level: 6.7
  • Imaged by UCSF Chimera
  • Download
  • Surface view colored by cylindrical radius
  • Surface level: 6.7
  • Imaged by UCSF Chimera
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  • Surface view with fitted model
  • Atomic models: PDB-6v35
  • Surface level: 6.7
  • Imaged by UCSF Chimera
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Movie viewer
Structure viewerEM map:
SurfViewMolmilJmol/JSmol
Supplemental images

emd_21028.png

Downloads & links

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Map

FileDownload / File: emd_21028.map.gz / Format: CCP4 / Size: 64 MB / Type: IMAGE STORED AS FLOATING POINT NUMBER (4 BYTES)
AnnotationB factor-sharpened Ca2 -free hsSlo1-beta4 channel complex
Projections & slices

Image control

Size
Brightness
Contrast
Others
AxesZ (Sec.)Y (Row.)X (Col.)
1.3 Å/pix.
x 256 pix.
= 332.8 Å		1.3 Å/pix.
x 256 pix.
= 332.8 Å		1.3 Å/pix.
x 256 pix.
= 332.8 Å

Surface

Projections

Slices (1/3)

Slices (1/2)

Slices (2/3)

Images are generated by Spider.

Voxel sizeX=Y=Z: 1.3 Å
Density

Histogram

Histogram (log scale)
Contour LevelBy AUTHOR: 6.7 / Movie #1: 6.7
Minimum - Maximum-9.618023000000001 - 21.471024
Average (Standard dev.)-0.34174907 (±1.078085)
SymmetrySpace group: 1
Details

EMDB XML:

Map geometry
Axis orderXYZ
Origin000
Dimensions256256256
Spacing256256256
CellA=B=C: 332.8 Å
α=β=γ: 90.0 °

CCP4 map header:

modeImage stored as Reals
Å/pix. X/Y/Z1.31.31.3
M x/y/z256256256
origin x/y/z0.0000.0000.000
length x/y/z332.800332.800332.800
α/β/γ90.00090.00090.000
start NX/NY/NZ000
NX/NY/NZ162182277
MAP C/R/S123
start NC/NR/NS000
NC/NR/NS256256256
D min/max/mean-9.61821.471-0.342

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Supplemental data

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Sample components

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Entire : hsSlo1 alpha subunit-beta4 subunit complex

EntireName: hsSlo1 alpha subunit-beta4 subunit complex
Components
  • Complex: hsSlo1 alpha subunit-beta4 subunit complex
    • Protein or peptide: Calcium-activated potassium channel subunit alpha-1
    • Protein or peptide: Calcium-activated potassium channel subunit beta-4
  • Ligand: (1R)-2-{[(S)-{[(2S)-2,3-dihydroxypropyl]oxy}(hydroxy)phosphoryl]oxy}-1-[(hexadecanoyloxy)methyl]ethyl (9Z)-octadec-9-enoate
  • Ligand: CHOLESTEROL
  • Ligand: 2-acetamido-2-deoxy-beta-D-glucopyranose

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Supramolecule #1: hsSlo1 alpha subunit-beta4 subunit complex

SupramoleculeName: hsSlo1 alpha subunit-beta4 subunit complex / type: complex / ID: 1 / Parent: 0 / Macromolecule list: #1-#2
Details: human high conductance Ca2+-activated K+ channel Slo1 (BK) alpha subunit and beta4 subunit
Source (natural)Organism: Homo sapiens (human)
Molecular weightTheoretical: 580 KDa

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Macromolecule #1: Calcium-activated potassium channel subunit alpha-1

MacromoleculeName: Calcium-activated potassium channel subunit alpha-1 / type: protein_or_peptide / ID: 1 / Number of copies: 4 / Enantiomer: LEVO
Source (natural)Organism: Homo sapiens (human)
Molecular weightTheoretical: 119.988062 KDa
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 LIFCLMQFDS NSLEVLFQ

UniProtKB: Calcium-activated potassium channel subunit alpha-1

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Macromolecule #2: Calcium-activated potassium channel subunit beta-4

MacromoleculeName: Calcium-activated potassium channel subunit beta-4 / type: protein_or_peptide / ID: 2 / Number of copies: 4 / Enantiomer: LEVO
Source (natural)Organism: Homo sapiens (human)
Molecular weightTheoretical: 24.991756 KDa
Recombinant expressionOrganism: Homo sapiens (human)
SequenceString: MAKLRVAYEY TEAEDKSIRL GLFLIISGVV SLFIFGFCWL SPALQDLQAT EANCTVLSVQ QIGEVFECTF TCGADCRGTS QYPCVQVYV NNSESNSRAL LHSDEHQLLT NPKCSYIPPC KRENQKNLES VMNWQQYWKD EIGSQPFTCY FNQHQRPDDV L LHRTHDEI ...String:
MAKLRVAYEY TEAEDKSIRL GLFLIISGVV SLFIFGFCWL SPALQDLQAT EANCTVLSVQ QIGEVFECTF TCGADCRGTS QYPCVQVYV NNSESNSRAL LHSDEHQLLT NPKCSYIPPC KRENQKNLES VMNWQQYWKD EIGSQPFTCY FNQHQRPDDV L LHRTHDEI VLLHCFLWPL VTFVVGVLIV VLTICAKSLA VKAEAMKKRK FSSNSLEVLF Q

UniProtKB: Calcium-activated potassium channel subunit beta-4

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Macromolecule #3: (1R)-2-{[(S)-{[(2S)-2,3-dihydroxypropyl]oxy}(hydroxy)phosphoryl]o...

MacromoleculeName: (1R)-2-{[(S)-{[(2S)-2,3-dihydroxypropyl]oxy}(hydroxy)phosphoryl]oxy}-1-[(hexadecanoyloxy)methyl]ethyl (9Z)-octadec-9-enoate
type: ligand / ID: 3 / Number of copies: 40 / Formula: PGW
Molecular weightTheoretical: 749.007 Da

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Macromolecule #4: CHOLESTEROL

MacromoleculeName: CHOLESTEROL / type: ligand / ID: 4 / Number of copies: 8 / Formula: CLR
Molecular weightTheoretical: 386.654 Da
Chemical component information

ChemComp-CLR:
CHOLESTEROL

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Macromolecule #5: 2-acetamido-2-deoxy-beta-D-glucopyranose

MacromoleculeName: 2-acetamido-2-deoxy-beta-D-glucopyranose / type: ligand / ID: 5 / Number of copies: 8 / Formula: NAG
Molecular weightTheoretical: 221.208 Da
Chemical component information

ChemComp-NAG:
2-acetamido-2-deoxy-beta-D-glucopyranose

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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

Concentration8.5 mg/mL
BufferpH: 8
Component:
ConcentrationFormulaName
450.0 mMKClpotassium chloride
20.0 mMC4H11NO3Tris
0.5 mMC56H92O29Digitonin
5.0 mMC14H24N2O10EGTA
15.0 mMMgCl2magnesium chloride
GridModel: Quantifoil R1.2/1.3 / Material: GOLD / Mesh: 400 / Support film - Material: CARBON / Support film - topology: HOLEY / Pretreatment - Type: GLOW DISCHARGE / Pretreatment - Time: 12 sec. / Pretreatment - Atmosphere: AIR
VitrificationCryogen name: ETHANE / Chamber humidity: 100 % / Chamber temperature: 295 K / Instrument: FEI VITROBOT MARK IV / Details: Blot for 4 seconds before plunging..

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Electron microscopy

MicroscopeFEI TITAN KRIOS
Image recordingFilm or detector model: GATAN K2 SUMMIT (4k x 4k) / Detector mode: SUPER-RESOLUTION / Digitization - Frames/image: 1-50 / Number grids imaged: 1 / Number real images: 3405 / Average exposure time: 15.0 sec. / Average electron dose: 89.0 e/Å2
Electron beamAcceleration voltage: 300 kV / Electron source: FIELD EMISSION GUN
Electron opticsC2 aperture diameter: 70.0 µm / Illumination mode: FLOOD BEAM / Imaging mode: BRIGHT FIELD / Cs: 2.7 mm / Nominal defocus max: 2.0 µm / Nominal defocus min: 0.7000000000000001 µm
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
Final reconstructionNumber classes used: 1 / Applied symmetry - Point group: C4 (4 fold cyclic) / Resolution.type: BY AUTHOR / Resolution: 3.5 Å / Resolution method: FSC 0.143 CUT-OFF / Software - Name: FREALIGN (ver. 9.11) / Number images used: 42842
Initial angle assignmentType: PROJECTION MATCHING / Software - Name: cryoSPARC (ver. 2.9.0)
Final angle assignmentType: PROJECTION MATCHING / Software - Name: FREALIGN (ver. 9.11)
Final 3D classificationNumber classes: 3 / Software - Name: RELION (ver. 3.0.8)

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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: OTHER
Output model

PDB-6v35:
Cryo-EM structure of Ca2+-free hsSlo1-beta4 channel complex

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