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- PDB-9o48: Cryo-EM structure of the human SK2-4 chimera/calmodulin channel c... -

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Basic information

Entry
Database: PDB / ID: 9o48
TitleCryo-EM structure of the human SK2-4 chimera/calmodulin channel complex in the Ca2+ bound state
Components
  • Calmodulin-1
  • Intermediate conductance calcium-activated potassium channel protein 4,Small conductance calcium-activated potassium channel protein 2 chimera
KeywordsTRANSPORT PROTEIN / Ion channel / Calcium / Potassium / Calmodulin
Function / homology
Function and homology information


intermediate conductance calcium-activated potassium channel activity / saliva secretion / small conductance calcium-activated potassium channel activity / Acetylcholine inhibits contraction of outer hair cells / membrane repolarization during atrial cardiac muscle cell action potential / stabilization of membrane potential / Ca2+ activated K+ channels / macropinocytosis / calcium-activated potassium channel activity / regulation of calcium ion import across plasma membrane ...intermediate conductance calcium-activated potassium channel activity / saliva secretion / small conductance calcium-activated potassium channel activity / Acetylcholine inhibits contraction of outer hair cells / membrane repolarization during atrial cardiac muscle cell action potential / stabilization of membrane potential / Ca2+ activated K+ channels / macropinocytosis / calcium-activated potassium channel activity / regulation of calcium ion import across plasma membrane / positive regulation of potassium ion transmembrane transport / inward rectifier potassium channel activity / cell volume homeostasis / regulation of potassium ion transmembrane transport / CaM pathway / Cam-PDE 1 activation / Sodium/Calcium exchangers / Calmodulin induced events / Reduction of cytosolic Ca++ levels / Activation of Ca-permeable Kainate Receptor / CREB1 phosphorylation through the activation of CaMKII/CaMKK/CaMKIV cascasde / Loss of phosphorylation of MECP2 at T308 / phospholipid translocation / CREB1 phosphorylation through the activation of Adenylate Cyclase / CaMK IV-mediated phosphorylation of CREB / PKA activation / negative regulation of high voltage-gated calcium channel activity / Glycogen breakdown (glycogenolysis) / positive regulation of ryanodine-sensitive calcium-release channel activity / CLEC7A (Dectin-1) induces NFAT activation / Activation of RAC1 downstream of NMDARs / negative regulation of calcium ion export across plasma membrane / organelle localization by membrane tethering / mitochondrion-endoplasmic reticulum membrane tethering / autophagosome membrane docking / presynaptic endocytosis / regulation of cardiac muscle cell action potential / negative regulation of ryanodine-sensitive calcium-release channel activity / Synthesis of IP3 and IP4 in the cytosol / regulation of cell communication by electrical coupling involved in cardiac conduction / positive regulation of T cell receptor signaling pathway / Phase 0 - rapid depolarisation / Negative regulation of NMDA receptor-mediated neuronal transmission / Unblocking of NMDA receptors, glutamate binding and activation / alpha-actinin binding / RHO GTPases activate PAKs / calcineurin-mediated signaling / Ion transport by P-type ATPases / Uptake and function of anthrax toxins / regulation of ryanodine-sensitive calcium-release channel activity / Long-term potentiation / Calcineurin activates NFAT / Regulation of MECP2 expression and activity / protein phosphatase activator activity / DARPP-32 events / Smooth Muscle Contraction / immune system process / catalytic complex / potassium channel activity / detection of calcium ion / regulation of cardiac muscle contraction / RHO GTPases activate IQGAPs / regulation of cardiac muscle contraction by regulation of the release of sequestered calcium ion / presynaptic cytosol / calcium channel inhibitor activity / cellular response to interferon-beta / Protein methylation / Activation of AMPK downstream of NMDARs / Ion homeostasis / regulation of release of sequestered calcium ion into cytosol by sarcoplasmic reticulum / eNOS activation / regulation of calcium-mediated signaling / Tetrahydrobiopterin (BH4) synthesis, recycling, salvage and regulation / titin binding / voltage-gated potassium channel complex / potassium ion transmembrane transport / sperm midpiece / substantia nigra development / calcium channel complex / calyx of Held / FCERI mediated Ca+2 mobilization / Ras activation upon Ca2+ influx through NMDA receptor / FCGR3A-mediated IL10 synthesis / adenylate cyclase activator activity / regulation of heart rate / Antigen activates B Cell Receptor (BCR) leading to generation of second messengers / protein serine/threonine kinase activator activity / VEGFR2 mediated cell proliferation / sarcomere / regulation of cytokinesis / VEGFR2 mediated vascular permeability / positive regulation of protein secretion / Translocation of SLC2A4 (GLUT4) to the plasma membrane / positive regulation of receptor signaling pathway via JAK-STAT / spindle microtubule / establishment of localization in cell / RAF activation / Transcriptional activation of mitochondrial biogenesis / defense response / potassium ion transport
Similarity search - Function
Calmodulin-binding domain / Potassium channel, calcium-activated, SK / SK, calmodulin-binding domain superfamily / Calmodulin binding domain / Calcium-activated SK potassium channel / Calmodulin binding domain / Potassium channel domain / Ion channel / : / EF-hand domain pair ...Calmodulin-binding domain / Potassium channel, calcium-activated, SK / SK, calmodulin-binding domain superfamily / Calmodulin binding domain / Calcium-activated SK potassium channel / Calmodulin binding domain / Potassium channel domain / Ion channel / : / EF-hand domain pair / EF-hand, calcium binding motif / EF-Hand 1, calcium-binding site / EF-hand calcium-binding domain. / EF-hand calcium-binding domain profile. / EF-hand domain / EF-hand domain pair
Similarity search - Domain/homology
: / Intermediate conductance calcium-activated potassium channel protein 4 / Calmodulin-1 / Small conductance calcium-activated potassium channel protein 2
Similarity search - Component
Biological speciesHomo sapiens (human)
MethodELECTRON MICROSCOPY / single particle reconstruction / cryo EM / Resolution: 3.1 Å
AuthorsCassell, S.J. / Khoshouei, M. / Wilhelm, W.A. / Whicher, J.R.
Funding support1items
OrganizationGrant numberCountry
Not funded
CitationJournal: To Be Published
Title: Mechanism of SK2 channel gating and its modulation by the bee toxin apamin and small molecules
Authors: Cassell, S.J. / Li, W. / Krautwald, S. / Khoshouei, M. / Lee, Y.T. / Hou, J. / Guan, W. / Peukert, S. / Wilhelm, W.A. / Whicher, J.R.
History
DepositionApr 8, 2025Deposition site: RCSB / Processing site: RCSB
Revision 1.0Jul 9, 2025Provider: repository / Type: Initial release
Revision 1.0Jul 9, 2025Data content type: EM metadata / Data content type: EM metadata / Provider: repository / Type: Initial release
Revision 1.0Jul 9, 2025Data content type: Additional map / Part number: 1 / Data content type: Additional map / Provider: repository / Type: Initial release
Revision 1.0Jul 9, 2025Data content type: FSC / Data content type: FSC / Provider: repository / Type: Initial release
Revision 1.0Jul 9, 2025Data content type: Half map / Part number: 1 / Data content type: Half map / Provider: repository / Type: Initial release
Revision 1.0Jul 9, 2025Data content type: Half map / Part number: 2 / Data content type: Half map / Provider: repository / Type: Initial release
Revision 1.0Jul 9, 2025Data content type: Image / Data content type: Image / Provider: repository / Type: Initial release
Revision 1.0Jul 9, 2025Data content type: Primary map / Data content type: Primary map / Provider: repository / Type: Initial release

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Structure visualization

Structure viewerMolecule:
MolmilJmol/JSmol

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Assembly

Deposited unit
A: Intermediate conductance calcium-activated potassium channel protein 4,Small conductance calcium-activated potassium channel protein 2 chimera
B: Intermediate conductance calcium-activated potassium channel protein 4,Small conductance calcium-activated potassium channel protein 2 chimera
C: Intermediate conductance calcium-activated potassium channel protein 4,Small conductance calcium-activated potassium channel protein 2 chimera
D: Intermediate conductance calcium-activated potassium channel protein 4,Small conductance calcium-activated potassium channel protein 2 chimera
E: Calmodulin-1
F: Calmodulin-1
G: Calmodulin-1
H: Calmodulin-1
hetero molecules


Theoretical massNumber of molelcules
Total (without water)265,65826
Polymers264,9398
Non-polymers71918
Water00
1


  • Idetical with deposited unit
  • defined by author
  • Evidence: electron microscopy
TypeNameSymmetry operationNumber
identity operation1_5551

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Components

#1: Protein
Intermediate conductance calcium-activated potassium channel protein 4,Small conductance calcium-activated potassium channel protein 2 chimera / SKCa 4 / SKCa4 / hSK4 / Gardos channel / IKCa1 / hIK1 / KCa3.1 / Putative Gardos channel / hKCa4 / ...SKCa 4 / SKCa4 / hSK4 / Gardos channel / IKCa1 / hIK1 / KCa3.1 / Putative Gardos channel / hKCa4 / SK2 / SKCa 2 / SKCa2 / KCa2.2


Mass: 49382.164 Da / Num. of mol.: 4
Fragment: SK4 residues 1-15 + SK2 residues 124-412 + SK4 residues 306-428
Source method: isolated from a genetically manipulated source
Source: (gene. exp.) Homo sapiens (human) / Gene: KCNN4, IK1, IKCA1, KCA4, SK4, KCNN2 / Production host: Homo sapiens (human) / References: UniProt: O15554, UniProt: Q9H2S1
#2: Protein
Calmodulin-1


Mass: 16852.545 Da / Num. of mol.: 4
Source method: isolated from a genetically manipulated source
Source: (gene. exp.) Homo sapiens (human) / Gene: CALM1, CALM, CAM, CAM1 / Production host: Homo sapiens (human) / References: UniProt: P0DP23
#3: Chemical ChemComp-K / POTASSIUM ION


Mass: 39.098 Da / Num. of mol.: 2 / Source method: obtained synthetically / Formula: K
#4: Chemical
ChemComp-CA / CALCIUM ION


Mass: 40.078 Da / Num. of mol.: 16 / Source method: obtained synthetically / Formula: Ca
Has ligand of interestN
Has protein modificationY

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Experimental details

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Experiment

ExperimentMethod: ELECTRON MICROSCOPY
EM experimentAggregation state: PARTICLE / 3D reconstruction method: single particle reconstruction

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Sample preparation

Component
IDNameTypeEntity IDParent-IDSource
1Ternary complex of human SK2-4 chimera/calmodulin channel complex in the Ca2+ bound stateCOMPLEX#1-#20RECOMBINANT
2Human SK2-4 chimeric channelCOMPLEX#11RECOMBINANT
3Human calmodulinCOMPLEX#21RECOMBINANT
Source (natural)
IDEntity assembly-IDOrganismNcbi tax-ID
22Homo sapiens (human)9606
32Homo sapiens (human)9606
43Homo sapiens (human)9606
Source (recombinant)
IDEntity assembly-IDOrganismNcbi tax-IDStrain
22Homo sapiens (human)9606Expi293F
32Homo sapiens (human)9606Expi293F
43Homo sapiens (human)9606Expi293F
Buffer solutionpH: 8
Details: 20 mM Tris pH 8, 150 mM KCl, 2 mM CaCl2, 0.005% GDN, 0.0005% CHS
SpecimenConc.: 7.5 mg/ml / Embedding applied: NO / Shadowing applied: NO / Staining applied: NO / Vitrification applied: YES
Specimen supportDetails: UltrAuFoil R 1.2/0.4 200 mesh grids were glow-discharged for 30 seconds at 0.38 mBar with 15 mA using Glocube system
Grid material: GOLD / Grid mesh size: 200 divisions/in. / Grid type: UltrAuFoil R1.2/1.3
VitrificationInstrument: FEI VITROBOT MARK IV / Cryogen name: ETHANE / Humidity: 100 % / Chamber temperature: 298 K
Details: 5 uL of sample was applied to grids at 4 degree temperature with 100% humidity. After 30 seconds, grids were blotted for 5 seconds with blot force 25 and plunged into liquid ethane.

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Electron microscopy imaging

Experimental equipment
Model: Titan Krios / Image courtesy: FEI Company
MicroscopyModel: TFS KRIOS
Electron gunElectron source: FIELD EMISSION GUN / Accelerating voltage: 300 kV / Illumination mode: FLOOD BEAM
Electron lensMode: BRIGHT FIELD / Nominal magnification: 75000 X / Nominal defocus max: 1600 nm / Nominal defocus min: 800 nm / Cs: 0.01 mm / C2 aperture diameter: 50 µm
Specimen holderCryogen: NITROGEN / Specimen holder model: FEI TITAN KRIOS AUTOGRID HOLDER
Image recordingAverage exposure time: 3.25 sec. / Electron dose: 50 e/Å2 / Film or detector model: FEI FALCON IV (4k x 4k) / Num. of grids imaged: 1 / Num. of real images: 10123
Details: Images were collected in EER format with 50 fractions
EM imaging opticsSpherical aberration corrector: Microscope has a Cs corrector

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Processing

EM software
IDNameVersionCategory
1cryoSPARCparticle selection
2EPUimage acquisition
7PHENIX1.21.2_5419model fitting
9PHENIX1.21.2_5419model refinement
10cryoSPARCinitial Euler assignment
11cryoSPARCfinal Euler assignment
12cryoSPARCclassification
13cryoSPARC3D reconstruction
CTF correctionType: PHASE FLIPPING AND AMPLITUDE CORRECTION
Particle selectionNum. of particles selected: 8300000
SymmetryPoint symmetry: C4 (4 fold cyclic)
3D reconstructionResolution: 3.1 Å / Resolution method: FSC 0.143 CUT-OFF / Num. of particles: 658368 / Symmetry type: POINT
Atomic model buildingProtocol: AB INITIO MODEL / Space: REAL
RefinementHighest resolution: 3.1 Å
Stereochemistry target values: REAL-SPACE (WEIGHTED MAP SUM AT ATOM CENTERS)
Refine LS restraints
Refine-IDTypeDev idealNumber
ELECTRON MICROSCOPYf_bond_d0.00216368
ELECTRON MICROSCOPYf_angle_d0.31622136
ELECTRON MICROSCOPYf_dihedral_angle_d3.0722212
ELECTRON MICROSCOPYf_chiral_restr0.0342568
ELECTRON MICROSCOPYf_plane_restr0.0022788

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