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- PDB-6uxb: MthK N-terminal truncation state 3 bound with calcium -

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Basic information

Entry
Database: PDB / ID: 6uxb
TitleMthK N-terminal truncation state 3 bound with calcium
ComponentsCalcium-gated potassium channel MthK
KeywordsTRANSPORT PROTEIN / MthK
Function / homology
Function and homology information


monoatomic cation transmembrane transporter activity / potassium ion transport / identical protein binding / metal ion binding / plasma membrane
Similarity search - Function
: / Regulator of K+ conductance, N-terminal / TrkA-N domain / Regulator of K+ conductance, C-terminal / Regulator of K+ conductance, C-terminal domain superfamily / TrkA-C domain / RCK C-terminal domain profile. / RCK N-terminal domain profile. / Potassium channel domain / Ion channel / NAD(P)-binding domain superfamily
Similarity search - Domain/homology
Calcium-gated potassium channel MthK
Similarity search - Component
Biological speciesMethanothermobacter thermautotrophicus (archaea)
MethodELECTRON MICROSCOPY / single particle reconstruction / cryo EM / Resolution: 4.9 Å
AuthorsChen, F. / Crina, N.
CitationJournal: Nature / Year: 2020
Title: Ball-and-chain inactivation in a calcium-gated potassium channel.
Authors: Chen Fan / Nattakan Sukomon / Emelie Flood / Jan Rheinberger / Toby W Allen / Crina M Nimigean /
Abstract: Inactivation is the process by which ion channels terminate ion flux through their pores while the opening stimulus is still present. In neurons, inactivation of both sodium and potassium channels is ...Inactivation is the process by which ion channels terminate ion flux through their pores while the opening stimulus is still present. In neurons, inactivation of both sodium and potassium channels is crucial for the generation of action potentials and regulation of firing frequency. A cytoplasmic domain of either the channel or an accessory subunit is thought to plug the open pore to inactivate the channel via a 'ball-and-chain' mechanism. Here we use cryo-electron microscopy to identify the molecular gating mechanism in calcium-activated potassium channels by obtaining structures of the MthK channel from Methanobacterium thermoautotrophicum-a purely calcium-gated and inactivating channel-in a lipid environment. In the absence of Ca, we obtained a single structure in a closed state, which was shown by atomistic simulations to be highly flexible in lipid bilayers at ambient temperature, with large rocking motions of the gating ring and bending of pore-lining helices. In Ca-bound conditions, we obtained several structures, including multiple open-inactivated conformations, further indication of a highly dynamic protein. These different channel conformations are distinguished by rocking of the gating rings with respect to the transmembrane region, indicating symmetry breakage across the channel. Furthermore, in all conformations displaying open channel pores, the N terminus of one subunit of the channel tetramer sticks into the pore and plugs it, with free energy simulations showing that this is a strong interaction. Deletion of this N terminus leads to functionally non-inactivating channels and structures of open states without a pore plug, indicating that this previously unresolved N-terminal peptide is responsible for a ball-and-chain inactivation mechanism.
History
DepositionNov 7, 2019Deposition site: RCSB / Processing site: RCSB
Revision 1.0Mar 18, 2020Provider: repository / Type: Initial release
Revision 1.1Apr 22, 2020Group: Database references / Category: citation / citation_author
Item: _citation.journal_volume / _citation.page_first ..._citation.journal_volume / _citation.page_first / _citation.page_last / _citation.pdbx_database_id_DOI / _citation.pdbx_database_id_PubMed / _citation.title / _citation_author.identifier_ORCID
Revision 1.2Mar 6, 2024Group: Data collection / Database references / Category: chem_comp_atom / chem_comp_bond / database_2
Item: _database_2.pdbx_DOI / _database_2.pdbx_database_accession

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Structure visualization

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Assembly

Deposited unit
A: Calcium-gated potassium channel MthK
E: Calcium-gated potassium channel MthK
C: Calcium-gated potassium channel MthK
G: Calcium-gated potassium channel MthK
B: Calcium-gated potassium channel MthK
F: Calcium-gated potassium channel MthK
D: Calcium-gated potassium channel MthK
H: Calcium-gated potassium channel MthK


Theoretical massNumber of molelcules
Total (without water)298,8498
Polymers298,8498
Non-polymers00
Water00
1


  • Idetical with deposited unit
  • defined by author
  • Evidence: gel filtration
TypeNameSymmetry operationNumber
identity operation1_5551

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Components

#1: Protein
Calcium-gated potassium channel MthK


Mass: 37356.160 Da / Num. of mol.: 8
Source method: isolated from a genetically manipulated source
Source: (gene. exp.) Methanothermobacter thermautotrophicus (archaea)
Gene: mthK, MTH_1520 / Production host: Escherichia coli (E. coli) / References: UniProt: O27564

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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

ComponentName: MthK N-terminal truncation / Type: COMPLEX / Entity ID: all / Source: RECOMBINANT
Source (natural)Organism: Methanothermobacter thermautotrophicus (archaea)
Source (recombinant)Organism: Escherichia coli (E. coli)
Buffer solutionpH: 8.5
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 FIELD
Image recordingElectron dose: 1.425 e/Å2 / Detector mode: COUNTING / Film or detector model: GATAN K2 SUMMIT (4k x 4k)

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Processing

EM software
IDNameVersionCategory
2Leginonimage acquisition
4CTFFIND4CTF correction
7UCSF Chimeramodel fitting
12RELION33D reconstruction
CTF correctionType: PHASE FLIPPING AND AMPLITUDE CORRECTION
SymmetryPoint symmetry: C1 (asymmetric)
3D reconstructionResolution: 4.9 Å / Resolution method: FSC 0.143 CUT-OFF / Num. of particles: 68286 / Symmetry type: POINT

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