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- PDB-6pby: Single particle cryo-EM structure of the voltage-gated K+ channel... -

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Basic information

Entry
Database: PDB / ID: 6pby
TitleSingle particle cryo-EM structure of the voltage-gated K+ channel Eag1 3-13 deletion mutant bound to calmodulin (conformation 1)
Components
  • Calmodulin-1
  • Potassium voltage-gated channel subfamily H member 1
KeywordsTRANSPORT PROTEIN/CALCIUM BINDING PROTEIN / Voltage-gated potassium channel / ion channel / calmodulin / TRANSPORT PROTEIN-CALCIUM BINDING PROTEIN complex
Function / homology
Function and homology information


Voltage gated Potassium channels / potassium channel complex / regulation of presynaptic cytosolic calcium ion concentration / delayed rectifier potassium channel activity / voltage-gated monoatomic ion channel activity involved in regulation of presynaptic membrane potential / parallel fiber to Purkinje cell synapse / CaM pathway / Cam-PDE 1 activation / Sodium/Calcium exchangers / nuclear inner membrane ...Voltage gated Potassium channels / potassium channel complex / regulation of presynaptic cytosolic calcium ion concentration / delayed rectifier potassium channel activity / voltage-gated monoatomic ion channel activity involved in regulation of presynaptic membrane potential / parallel fiber to Purkinje cell synapse / CaM pathway / Cam-PDE 1 activation / Sodium/Calcium exchangers / nuclear inner membrane / Calmodulin induced events / Reduction of cytosolic Ca++ levels / Activation of Ca-permeable Kainate Receptor / CREB1 phosphorylation through the activation of CaMKII/CaMKK/CaMKIV cascasde / phosphatidylinositol bisphosphate binding / Loss of phosphorylation of MECP2 at T308 / CREB1 phosphorylation through the activation of Adenylate Cyclase / PKA activation / negative regulation of high voltage-gated calcium channel activity / regulation of synaptic vesicle exocytosis / CaMK IV-mediated phosphorylation of CREB / Glycogen breakdown (glycogenolysis) / negative regulation of calcium ion export across plasma membrane / organelle localization by membrane tethering / Activation of RAC1 downstream of NMDARs / regulation of cardiac muscle cell action potential / mitochondrion-endoplasmic reticulum membrane tethering / CLEC7A (Dectin-1) induces NFAT activation / autophagosome membrane docking / positive regulation of ryanodine-sensitive calcium-release channel activity / Negative regulation of NMDA receptor-mediated neuronal transmission / regulation of cell communication by electrical coupling involved in cardiac conduction / startle response / Unblocking of NMDA receptors, glutamate binding and activation / negative regulation of peptidyl-threonine phosphorylation / Synthesis of IP3 and IP4 in the cytosol / Phase 0 - rapid depolarisation / protein phosphatase activator activity / RHO GTPases activate PAKs / positive regulation of cyclic-nucleotide phosphodiesterase activity / positive regulation of phosphoprotein phosphatase activity / Long-term potentiation / Ion transport by P-type ATPases / Uptake and function of anthrax toxins / Calcineurin activates NFAT / Regulation of MECP2 expression and activity / catalytic complex / DARPP-32 events / axolemma / detection of calcium ion / negative regulation of ryanodine-sensitive calcium-release channel activity / Smooth Muscle Contraction / RHO GTPases activate IQGAPs / regulation of cardiac muscle contraction / calcium channel inhibitor activity / cellular response to interferon-beta / regulation of cardiac muscle contraction by regulation of the release of sequestered calcium ion / Protein methylation / voltage-gated potassium channel complex / Activation of AMPK downstream of NMDARs / eNOS activation / regulation of release of sequestered calcium ion into cytosol by sarcoplasmic reticulum / regulation of calcium-mediated signaling / positive regulation of protein dephosphorylation / titin binding / regulation of ryanodine-sensitive calcium-release channel activity / Tetrahydrobiopterin (BH4) synthesis, recycling, salvage and regulation / Ion homeostasis / positive regulation of protein autophosphorylation / potassium ion transmembrane transport / sperm midpiece / calcium channel complex / substantia nigra development / cellular response to calcium ion / adenylate cyclase activator activity / Ras activation upon Ca2+ influx through NMDA receptor / regulation of membrane potential / regulation of heart rate / monoatomic ion transmembrane transport / protein serine/threonine kinase activator activity / sarcomere / FCERI mediated Ca+2 mobilization / FCGR3A-mediated IL10 synthesis / 14-3-3 protein binding / VEGFR2 mediated vascular permeability / positive regulation of peptidyl-threonine phosphorylation / Antigen activates B Cell Receptor (BCR) leading to generation of second messengers / VEGFR2 mediated cell proliferation / regulation of cytokinesis / Translocation of SLC2A4 (GLUT4) to the plasma membrane / spindle microtubule / postsynaptic density membrane / RAF activation / positive regulation of receptor signaling pathway via JAK-STAT / positive regulation of protein serine/threonine kinase activity / Transcriptional activation of mitochondrial biogenesis / Stimuli-sensing channels / spindle pole / cellular response to type II interferon / response to calcium ion
Similarity search - Function
Potassium channel, voltage-dependent, EAG / Potassium channel, voltage-dependent, EAG/ELK/ERG / PAS-associated, C-terminal / PAC domain profile. / PAS domain / PAC motif / Motif C-terminal to PAS motifs (likely to contribute to PAS structural domain) / Cyclic nucleotide-monophosphate binding domain / Cyclic nucleotide-binding domain / cAMP/cGMP binding motif profile. ...Potassium channel, voltage-dependent, EAG / Potassium channel, voltage-dependent, EAG/ELK/ERG / PAS-associated, C-terminal / PAC domain profile. / PAS domain / PAC motif / Motif C-terminal to PAS motifs (likely to contribute to PAS structural domain) / Cyclic nucleotide-monophosphate binding domain / Cyclic nucleotide-binding domain / cAMP/cGMP binding motif profile. / Cyclic nucleotide-binding domain / Cyclic nucleotide-binding domain superfamily / PAS repeat profile. / PAS domain / RmlC-like jelly roll fold / PAS domain superfamily / 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. / Ion transport domain / Ion transport protein / EF-hand domain / EF-hand domain pair
Similarity search - Domain/homology
Calmodulin-1 / Potassium voltage-gated channel subfamily H member 1
Similarity search - Component
Biological speciesRattus norvegicus (Norway rat)
Homo sapiens (human)
MethodELECTRON MICROSCOPY / single particle reconstruction / cryo EM / Resolution: 3.67 Å
AuthorsWhicher, J.R. / MacKinnon, R.
Funding support United States, 3items
OrganizationGrant numberCountry
National Institutes of Health/National Institute of General Medical Sciences (NIH/NIGMS)GM43949 United States
Howard Hughes Medical Institute (HHMI) United States
Damon Runyon Cancer Research FoundationDRG-2212-15 United States
CitationJournal: Elife / Year: 2019
Title: Regulation of Eag1 gating by its intracellular domains.
Authors: Jonathan R Whicher / Roderick MacKinnon /
Abstract: Voltage-gated potassium channels (Ks) are gated by transmembrane voltage sensors (VS) that move in response to changes in membrane voltage. K10.1 or Eag1 also has three intracellular domains: PAS, C- ...Voltage-gated potassium channels (Ks) are gated by transmembrane voltage sensors (VS) that move in response to changes in membrane voltage. K10.1 or Eag1 also has three intracellular domains: PAS, C-linker, and CNBHD. We demonstrate that the Eag1 intracellular domains are not required for voltage-dependent gating but likely interact with the VS to modulate gating. We identified specific interactions between the PAS, CNBHD, and VS that modulate voltage-dependent gating and provide evidence that VS movement destabilizes these interactions to promote channel opening. Additionally, mutation of these interactions renders Eag1 insensitive to calmodulin inhibition. The structure of the calmodulin insensitive mutant in a pre-open conformation suggests that channel opening may occur through a rotation of the intracellular domains and calmodulin may prevent this rotation by stabilizing interactions between the VS and intracellular domains. Intracellular domains likely play a similar modulatory role in voltage-dependent gating of the related K11-12 channels.
History
DepositionJun 14, 2019Deposition site: RCSB / Processing site: RCSB
Revision 1.0Sep 18, 2019Provider: repository / Type: Initial release
Revision 1.1Sep 25, 2019Group: Author supporting evidence / Data collection / Category: pdbx_audit_support / Item: _pdbx_audit_support.funding_organization
Revision 1.2Nov 20, 2019Group: Author supporting evidence / Category: pdbx_audit_support / Item: _pdbx_audit_support.funding_organization
Revision 1.3Mar 20, 2024Group: Data collection / Database references / Refinement description
Category: chem_comp_atom / chem_comp_bond ...chem_comp_atom / chem_comp_bond / database_2 / em_3d_fitting_list / pdbx_initial_refinement_model
Item: _database_2.pdbx_DOI / _database_2.pdbx_database_accession ..._database_2.pdbx_DOI / _database_2.pdbx_database_accession / _em_3d_fitting_list.accession_code / _em_3d_fitting_list.initial_refinement_model_id / _em_3d_fitting_list.source_name / _em_3d_fitting_list.type

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Assembly

Deposited unit
A: Potassium voltage-gated channel subfamily H member 1
B: Calmodulin-1
D: Calmodulin-1
F: Calmodulin-1
H: Calmodulin-1
C: Potassium voltage-gated channel subfamily H member 1
E: Potassium voltage-gated channel subfamily H member 1
G: Potassium voltage-gated channel subfamily H member 1


Theoretical massNumber of molelcules
Total (without water)452,4568
Polymers452,4568
Non-polymers00
Water0
1


  • Idetical with deposited unit
  • defined by author
  • Evidence: gel filtration
TypeNameSymmetry operationNumber
identity operation1_5551
Buried area41940 Å2
ΔGint-327 kcal/mol
Surface area149610 Å2

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Components

#1: Protein
Potassium voltage-gated channel subfamily H member 1 / Ether-a-go-go potassium channel 1 / r-eag / Voltage-gated potassium channel subunit Kv10.1


Mass: 96261.430 Da / Num. of mol.: 4 / Fragment: UNP residues 14-773,888-962
Source method: isolated from a genetically manipulated source
Source: (gene. exp.) Rattus norvegicus (Norway rat) / Gene: Kcnh1, Eag / Production host: Homo sapiens (human) / References: UniProt: Q63472
#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

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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
1Voltage-gated potassium channel Eag1 3-13 deletion mutant bound to calmodulinCOMPLEXall0MULTIPLE SOURCES
2Voltage-gated potassium channel Eag1 3-13 deletion mutantCOMPLEX#11RECOMBINANT
3calmodulinCOMPLEX#21RECOMBINANT
Source (natural)
IDEntity assembly-IDOrganismNcbi tax-ID
12Rattus norvegicus (Norway rat)10116
23Homo sapiens (human)9606
Source (recombinant)
IDEntity assembly-IDOrganismNcbi tax-ID
12Homo sapiens (human)9606
23Homo sapiens (human)9606
Buffer solutionpH: 8
SpecimenEmbedding applied: NO / Shadowing applied: NO / Staining applied: NO / Vitrification applied: YES
Specimen supportDetails: unspecified
VitrificationCryogen name: ETHANE

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Electron microscopy imaging

Experimental equipment
Model: Titan Krios / Image courtesy: FEI Company
MicroscopyModel: FEI TITAN KRIOS
Electron gunElectron source: FIELD EMISSION GUN / Accelerating voltage: 300 kV / Illumination mode: FLOOD BEAM
Electron lensMode: BRIGHT FIELDBright-field microscopy
Image recordingElectron dose: 1.6 e/Å2 / Film or detector model: GATAN K2 SUMMIT (4k x 4k)

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Processing

SoftwareName: PHENIX / Version: 1.14_3260: / Classification: refinement
EM software
IDNameVersionCategory
1RELION3particle selection
4CTFFIND4CTF correction
7Cootmodel fitting
9RELION3initial Euler assignment
10RELION3final Euler assignment
12RELION33D reconstruction
13PHENIXmodel refinement
CTF correctionType: PHASE FLIPPING AND AMPLITUDE CORRECTION
SymmetryPoint symmetry: C4 (4 fold cyclic)
3D reconstructionResolution: 3.67 Å / Resolution method: FSC 0.143 CUT-OFF / Num. of particles: 43137 / Symmetry type: POINT
Atomic model buildingProtocol: FLEXIBLE FIT / Space: REAL
Atomic model buildingPDB-ID: 5K7L

5k7l


Accession code: 5K7L / Source name: PDB / Type: experimental model

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