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- PDB-7uaa: CryoEM structure of the pancreatic ATP-sensitive potassium channe... -

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Basic information

Entry
Database: PDB / ID: 7uaa
TitleCryoEM structure of the pancreatic ATP-sensitive potassium channel in the ATP-bound state with Kir6.2-CTD in the up conformation
Components
  • ATP-binding cassette sub-family C member 8
  • ATP-sensitive inward rectifier potassium channel 11
KeywordsMEMBRANE PROTEIN / KATP channel / SUR1 / Kir6.2 / repaglinide / RPG / sulfonylurea receptor / potassium transport
Function / homology
Function and homology information


Regulation of insulin secretion / ATP sensitive Potassium channels / ABC-family proteins mediated transport / response to resveratrol / ATP-activated inward rectifier potassium channel activity / inward rectifying potassium channel / sulfonylurea receptor activity / cell body fiber / ventricular cardiac muscle tissue development / CAMKK-AMPK signaling cascade ...Regulation of insulin secretion / ATP sensitive Potassium channels / ABC-family proteins mediated transport / response to resveratrol / ATP-activated inward rectifier potassium channel activity / inward rectifying potassium channel / sulfonylurea receptor activity / cell body fiber / ventricular cardiac muscle tissue development / CAMKK-AMPK signaling cascade / voltage-gated monoatomic ion channel activity involved in regulation of presynaptic membrane potential / ATPase-coupled monoatomic cation transmembrane transporter activity / inward rectifier potassium channel activity / regulation of monoatomic ion transmembrane transport / nervous system process / inorganic cation transmembrane transport / response to stress / ankyrin binding / Ion homeostasis / response to ATP / action potential / potassium ion import across plasma membrane / response to testosterone / voltage-gated potassium channel activity / intercalated disc / potassium channel activity / axolemma / ABC-type transporter activity / negative regulation of insulin secretion / cellular response to nutrient levels / heat shock protein binding / T-tubule / potassium ion transmembrane transport / regulation of insulin secretion / acrosomal vesicle / regulation of membrane potential / determination of adult lifespan / response to ischemia / positive regulation of protein localization to plasma membrane / cellular response to glucose stimulus / potassium ion transport / sarcolemma / cellular response to nicotine / glucose metabolic process / cellular response to tumor necrosis factor / nuclear envelope / response to estradiol / presynaptic membrane / transmembrane transporter binding / response to hypoxia / endosome / response to xenobiotic stimulus / neuronal cell body / glutamatergic synapse / apoptotic process / ATP hydrolysis activity / protein-containing complex / ATP binding / plasma membrane / cytoplasm
Similarity search - Function
Potassium channel, inwardly rectifying, Kir6.2 / ATP-binding cassette subfamily C member 8 / Sulphonylurea receptor / Potassium channel, inwardly rectifying, transmembrane domain / Inward rectifier potassium channel transmembrane domain / Potassium channel, inwardly rectifying, Kir, cytoplasmic / Potassium channel, inwardly rectifying, Kir / Inward rectifier potassium channel, C-terminal / Inward rectifier potassium channel C-terminal domain / : ...Potassium channel, inwardly rectifying, Kir6.2 / ATP-binding cassette subfamily C member 8 / Sulphonylurea receptor / Potassium channel, inwardly rectifying, transmembrane domain / Inward rectifier potassium channel transmembrane domain / Potassium channel, inwardly rectifying, Kir, cytoplasmic / Potassium channel, inwardly rectifying, Kir / Inward rectifier potassium channel, C-terminal / Inward rectifier potassium channel C-terminal domain / : / ABC transporter transmembrane region / ABC transporter type 1, transmembrane domain / ABC transporter integral membrane type-1 fused domain profile. / ABC transporter type 1, transmembrane domain superfamily / ABC transporter-like, conserved site / ABC transporters family signature. / ABC transporter / ABC transporter-like, ATP-binding domain / ATP-binding cassette, ABC transporter-type domain profile. / Immunoglobulin E-set / ATPases associated with a variety of cellular activities / AAA+ ATPase domain / P-loop containing nucleoside triphosphate hydrolase
Similarity search - Domain/homology
ADENOSINE-5'-TRIPHOSPHATE / ATP-sensitive inward rectifier potassium channel 11 / ATP-binding cassette sub-family C member 8
Similarity search - Component
Biological speciesRattus norvegicus (Norway rat)
Cricetus cricetus (black-bellied hamster)
MethodELECTRON MICROSCOPY / single particle reconstruction / cryo EM / Resolution: 5.7 Å
AuthorsShyng, S.L. / Sung, M.W. / Driggers, C.M.
Funding support United States, 1items
OrganizationGrant numberCountry
National Institutes of Health/National Institute of Diabetes and Digestive and Kidney Disease (NIH/NIDDK)R01 DK066485-13 United States
Citation
Journal: J Mol Biol / Year: 2022
Title: Ligand-mediated Structural Dynamics of a Mammalian Pancreatic K Channel.
Authors: Min Woo Sung / Camden M Driggers / Barmak Mostofian / John D Russo / Bruce L Patton / Daniel M Zuckerman / Show-Ling Shyng /
Abstract: Regulation of pancreatic K channels involves orchestrated interactions of their subunits, Kir6.2 and SUR1, and ligands. Previously we reported K channel cryo-EM structures in the presence and absence ...Regulation of pancreatic K channels involves orchestrated interactions of their subunits, Kir6.2 and SUR1, and ligands. Previously we reported K channel cryo-EM structures in the presence and absence of pharmacological inhibitors and ATP, focusing on the mechanisms by which inhibitors act as pharmacological chaperones of K channels (Martin et al., 2019). Here we analyzed the same cryo-EM datasets with a focus on channel conformational dynamics to elucidate structural correlates pertinent to ligand interactions and channel gating. We found pharmacological inhibitors and ATP enrich a channel conformation in which the Kir6.2 cytoplasmic domain is closely associated with the transmembrane domain, while depleting one where the Kir6.2 cytoplasmic domain is extended away into the cytoplasm. This conformational change remodels a network of intra- and inter-subunit interactions as well as the ATP and PIP binding pockets. The structures resolved key contacts between the distal N-terminus of Kir6.2 and SUR1's ABC module involving residues implicated in channel function and showed a SUR1 residue, K134, participates in PIP binding. Molecular dynamics simulations revealed two Kir6.2 residues, K39 and R54, that mediate both ATP and PIP binding, suggesting a mechanism for competitive gating by ATP and PIP.
#1: Journal: Elife / Year: 2019
Title: Mechanism of pharmacochaperoning in a mammalian K channel revealed by cryo-EM.
Authors: Gregory M Martin / Min Woo Sung / Zhongying Yang / Laura M Innes / Balamurugan Kandasamy / Larry L David / Craig Yoshioka / Show-Ling Shyng /
Abstract: ATP-sensitive potassium (K) channels composed of a pore-forming Kir6.2 potassium channel and a regulatory ABC transporter sulfonylurea receptor 1 (SUR1) regulate insulin secretion in pancreatic β- ...ATP-sensitive potassium (K) channels composed of a pore-forming Kir6.2 potassium channel and a regulatory ABC transporter sulfonylurea receptor 1 (SUR1) regulate insulin secretion in pancreatic β-cells to maintain glucose homeostasis. Mutations that impair channel folding or assembly prevent cell surface expression and cause congenital hyperinsulinism. Structurally diverse K inhibitors are known to act as pharmacochaperones to correct mutant channel expression, but the mechanism is unknown. Here, we compare cryoEM structures of a mammalian K channel bound to pharmacochaperones glibenclamide, repaglinide, and carbamazepine. We found all three drugs bind within a common pocket in SUR1. Further, we found the N-terminus of Kir6.2 inserted within the central cavity of the SUR1 ABC core, adjacent the drug binding pocket. The findings reveal a common mechanism by which diverse compounds stabilize the Kir6.2 N-terminus within SUR1's ABC core, allowing it to act as a firm 'handle' for the assembly of metastable mutant SUR1-Kir6.2 complexes.
History
DepositionMar 11, 2022Deposition site: RCSB / Processing site: RCSB
Revision 1.0Aug 31, 2022Provider: repository / Type: Initial release
Revision 1.1Sep 14, 2022Group: Database references / Category: citation / citation_author
Item: _citation.journal_volume / _citation.title / _citation_author.name
Revision 1.2Jun 12, 2024Group: Data collection / Refinement description
Category: chem_comp_atom / chem_comp_bond ...chem_comp_atom / chem_comp_bond / em_3d_fitting_list / pdbx_initial_refinement_model
Item: _em_3d_fitting_list.accession_code / _em_3d_fitting_list.initial_refinement_model_id ..._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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Structure visualization

Structure viewerMolecule:
MolmilJmol/JSmol

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Assembly

Deposited unit
A: ATP-sensitive inward rectifier potassium channel 11
B: ATP-sensitive inward rectifier potassium channel 11
C: ATP-sensitive inward rectifier potassium channel 11
D: ATP-sensitive inward rectifier potassium channel 11
E: ATP-binding cassette sub-family C member 8
hetero molecules


Theoretical massNumber of molelcules
Total (without water)354,51710
Polymers351,9815
Non-polymers2,5365
Water00
1


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

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Components

#1: Protein
ATP-sensitive inward rectifier potassium channel 11 / BIR / Inward rectifier K(+) channel Kir6.2 / Potassium channel / inwardly rectifying subfamily J member 11


Mass: 43661.762 Da / Num. of mol.: 4
Source method: isolated from a genetically manipulated source
Source: (gene. exp.) Rattus norvegicus (Norway rat) / Gene: Kcnj11 / Cell line (production host): INS - 1 CELLS CLONE 832/13 / Production host: Rattus norvegicus (Norway rat) / References: UniProt: P70673
#2: Protein ATP-binding cassette sub-family C member 8 / SUR1 / Sulfonylurea receptor 1


Mass: 177333.578 Da / Num. of mol.: 1
Source method: isolated from a genetically manipulated source
Source: (gene. exp.) Cricetus cricetus (black-bellied hamster)
Gene: ABCC8, SUR / Cell line (production host): INS - 1 CELLS CLONE 832/13 / Production host: Rattus norvegicus (Norway rat) / References: UniProt: Q09427
#3: Chemical
ChemComp-ATP / ADENOSINE-5'-TRIPHOSPHATE


Mass: 507.181 Da / Num. of mol.: 5 / Source method: obtained synthetically / Formula: C10H16N5O13P3 / Feature type: SUBJECT OF INVESTIGATION / Comment: ATP, energy-carrying molecule*YM
Has ligand of interestY

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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: KATP-ATP-SUR1/Kir6.2-CTD-up / Type: COMPLEX
Details: Adenovirus-based co-infection of INS-1 cells with Kir6.2 and SUR1
Entity ID: #1-#2 / Source: MULTIPLE SOURCES
Source (natural)
IDEntity assembly-IDOrganismNcbi tax-ID
21Rattus norvegicus (Norway rat)10116
31Cricetus cricetus (black-bellied hamster)10034
Source (recombinant)
IDEntity assembly-IDOrganismNcbi tax-ID
21Rattus norvegicus (Norway rat)10116
31Rattus norvegicus (Norway rat)10116
Buffer solutionpH: 7.5
SpecimenEmbedding applied: NO / Shadowing applied: NO / Staining applied: NO / Vitrification applied: YES
VitrificationInstrument: FEI VITROBOT MARK III / Cryogen name: ETHANE / Humidity: 95 % / Chamber temperature: 279 K

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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 / Nominal defocus max: 2500 nm / Nominal defocus min: 1000 nm / Alignment procedure: COMA FREE
Image recordingElectron dose: 40 e/Å2 / Detector mode: SUPER-RESOLUTION / Film or detector model: GATAN K2 SUMMIT (4k x 4k)
Image scansMovie frames/image: 50

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Processing

Software
NameVersionClassificationNB
phenix.real_space_refine1.20.1_4487refinement
PHENIX1.20.1_4487refinement
EM software
IDNameCategory
2SerialEMimage acquisition
7Cootmodel fitting
12RELION3D reconstruction
13PHENIXmodel refinement
CTF correctionType: NONE
SymmetryPoint symmetry: C1 (asymmetric)
3D reconstructionResolution: 5.7 Å / Resolution method: FSC 0.143 CUT-OFF / Num. of particles: 90000 / Symmetry type: POINT
Atomic model buildingPDB-ID: 7TYT

7tyt


Accession code: 7TYT / Source name: PDB / Type: experimental model
RefinementCross valid method: NONE

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