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- PDB-5twv: Cryo-EM structure of the pancreatic ATP-sensitive K+ channel SUR1... -

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Entry
Database: PDB / ID: 5twv
TitleCryo-EM structure of the pancreatic ATP-sensitive K+ channel SUR1/Kir6.2 in the presence of ATP and glibenclamide
Components
  • ATP-binding cassette sub-family C member 8
  • ATP-sensitive inward rectifier potassium channel 11
KeywordsTRANSPORT PROTEIN / katp / kir6.2 / sur1 / potassium channel
Function / homology
Function and homology information


inward rectifying potassium channel / ATP-activated inward rectifier potassium channel activity / sulfonylurea receptor activity / cell body fiber / inorganic cation transmembrane transport / ATPase-coupled cation transmembrane transporter activity / inward rectifier potassium channel activity / positive regulation of cation channel activity / ankyrin binding / potassium ion import across plasma membrane ...inward rectifying potassium channel / ATP-activated inward rectifier potassium channel activity / sulfonylurea receptor activity / cell body fiber / inorganic cation transmembrane transport / ATPase-coupled cation transmembrane transporter activity / inward rectifier potassium channel activity / positive regulation of cation channel activity / ankyrin binding / potassium ion import across plasma membrane / nervous system process / response to ATP / axolemma / potassium channel activity / response to testosterone / potassium ion transmembrane transport / intercalated disc / voltage-gated potassium channel activity / ATPase-coupled transmembrane transporter activity / regulation of ion transmembrane transport / negative regulation of insulin secretion / regulation of insulin secretion / heat shock protein binding / T-tubule / acrosomal vesicle / regulation of membrane potential / response to ischemia / potassium ion transport / cellular response to glucose stimulus / positive regulation of protein localization to plasma membrane / sarcolemma / nuclear envelope / cellular response to nicotine / glucose metabolic process / cellular response to tumor necrosis factor / myelin sheath / response to estradiol / ion channel binding / protein C-terminus binding / endosome / ATPase activity / intracellular membrane-bounded organelle / neuronal cell body / response to drug / endoplasmic reticulum / mitochondrion / integral component of membrane / ATP binding / plasma membrane / cytosol
ABC transporter transmembrane region / AAA+ ATPase domain / Potassium channel, inwardly rectifying, Kir, cytoplasmic / ABC transporter-like / Immunoglobulin E-set / Potassium channel, inwardly rectifying, Kir / ABC transporter, conserved site / P-loop containing nucleoside triphosphate hydrolase / ABC transporter type 1, transmembrane domain superfamily / Potassium channel, inwardly rectifying, transmembrane domain ...ABC transporter transmembrane region / AAA+ ATPase domain / Potassium channel, inwardly rectifying, Kir, cytoplasmic / ABC transporter-like / Immunoglobulin E-set / Potassium channel, inwardly rectifying, Kir / ABC transporter, conserved site / P-loop containing nucleoside triphosphate hydrolase / ABC transporter type 1, transmembrane domain superfamily / Potassium channel, inwardly rectifying, transmembrane domain / Inward rectifier potassium channel, C-terminal / Inward rectifier potassium channel transmembrane domain / Inward rectifier potassium channel C-terminal domain / ABC transporter / Potassium channel, inwardly rectifying, Kir6.2 / ATP-binding cassette subfamily C member 8 / Sulphonylurea receptor / ABC transporter type 1, transmembrane domain
ATP-sensitive inward rectifier potassium channel 11 / ATP-binding cassette sub-family C member 8
Biological speciesRattus norvegicus (Norway rat)
Cricetus cricetus (black-bellied hamster)
MethodELECTRON MICROSCOPY / single particle reconstruction / cryo EM / Resolution: 6.3 Å
AuthorsMartin, G.M. / Yoshioka, C. / Chen, J.Z. / Shyng, S.L.
Funding support United States, 2items
OrganizationGrant numberCountry
National Institutes of Health/National Institute of Diabetes and Digestive and Kidney Disease (NIH/NIDDK)R01DK066485 United States
National Institutes of Health/National Institute of Diabetes and Digestive and Kidney Disease (NIH/NIDDK)F31DK105300 United States
CitationJournal: Elife / Year: 2017
Title: Cryo-EM structure of the ATP-sensitive potassium channel illuminates mechanisms of assembly and gating.
Authors: Gregory M Martin / Craig Yoshioka / Emily A Rex / Jonathan F Fay / Qing Xie / Matthew R Whorton / James Z Chen / Show-Ling Shyng /
Abstract: K channels are metabolic sensors that couple cell energetics to membrane excitability. In pancreatic β-cells, channels formed by SUR1 and Kir6.2 regulate insulin secretion and are the targets of ...K channels are metabolic sensors that couple cell energetics to membrane excitability. In pancreatic β-cells, channels formed by SUR1 and Kir6.2 regulate insulin secretion and are the targets of antidiabetic sulfonylureas. Here, we used cryo-EM to elucidate structural basis of channel assembly and gating. The structure, determined in the presence of ATP and the sulfonylurea glibenclamide, at ~6 Å resolution reveals a closed Kir6.2 tetrameric core with four peripheral SUR1s each anchored to a Kir6.2 by its N-terminal transmembrane domain (TMD0). Intricate interactions between TMD0, the loop following TMD0, and Kir6.2 near the proposed PIP binding site, and where ATP density is observed, suggest SUR1 may contribute to ATP and PIP binding to enhance Kir6.2 sensitivity to both. The SUR1-ABC core is found in an unusual inward-facing conformation whereby the two nucleotide binding domains are misaligned along a two-fold symmetry axis, revealing a possible mechanism by which glibenclamide inhibits channel activity.
Validation Report
SummaryFull reportAbout validation report
History
DepositionNov 14, 2016Deposition site: RCSB / Processing site: RCSB
Revision 1.0Jan 25, 2017Provider: repository / Type: Initial release
Revision 1.1May 31, 2017Group: Structure summary
Revision 1.2Sep 13, 2017Group: Author supporting evidence / Data collection / Category: em_image_scans / em_software / pdbx_audit_support
Item: _em_software.name / _pdbx_audit_support.funding_organization
Revision 1.3Dec 25, 2019Group: Author supporting evidence / Category: pdbx_audit_support / Item: _pdbx_audit_support.funding_organization

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Assembly

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


Theoretical massNumber of molelcules
Total (without water)889,93412
Polymers887,9058
Non-polymers2,0294
Water0
1


TypeNameSymmetry operationNumber
identity operation1_5551
Buried area36970 Å2
ΔGint-298 kcal/mol
Surface area351960 Å2

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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: 43645.719 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 / Production host: Rattus norvegicus (Norway rat) / References: UniProt: P70673*PLUS
#2: Protein
ATP-binding cassette sub-family C member 8 / Sulfonylurea receptor 1 / SUR1


Mass: 178330.516 Da / Num. of mol.: 4
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 / Production host: Rattus norvegicus (Norway rat) / References: UniProt: Q09427
#3: Chemical
ChemComp-ATP / ADENOSINE-5'-TRIPHOSPHATE / Adenosine triphosphate


Mass: 507.181 Da / Num. of mol.: 4 / Source method: obtained synthetically / Formula: C10H16N5O13P3 / Comment: ATP, energy-carrying molecule*YM
Sequence detailsKir6.2: GenBank reference BAA96239.1

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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
1ATP-sensitive K+ channel complex SUR1/Kir6.2COMPLEX#1-#20MULTIPLE SOURCES
2SUR1ABCC8COMPLEX#21RECOMBINANT
3Kir6.2COMPLEX#11RECOMBINANT
Source (natural)
IDEntity assembly-IDOrganismNcbi tax-ID
11Rattus norvegicus (Norway rat)10116
22Cricetus cricetus (black-bellied hamster)10034
33Rattus norvegicus (Norway rat)10116
Source (recombinant)
IDEntity assembly-IDOrganismNcbi tax-IDCellPlasmid
11Rattus norvegicus (Norway rat)10116INS-1
22Rattus norvegicus (Norway rat)10116INS-1pShuttle-TetR:pAdEasy
33Rattus norvegicus (Norway rat)10116INS-1pShuttle-CMV:pAdEasy
Buffer solutionpH: 7.5
SpecimenEmbedding applied: NO / Shadowing applied: NO / Staining applied: NO / Vitrification applied: YES
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: SPOT SCAN
Electron lensMode: BRIGHT FIELDBright-field microscopy / Nominal magnification: 81000 X / Nominal defocus max: 3000 nm / Nominal defocus min: 1500 nm
Image recordingElectron dose: 40 e/Å2 / Detector mode: COUNTING / Film or detector model: GATAN K2 SUMMIT (4k x 4k)

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Processing

EM software
IDNameVersionCategory
4CTFFIND4CTF correction
13RELION23D reconstruction
CTF correctionType: PHASE FLIPPING AND AMPLITUDE CORRECTION
SymmetryPoint symmetry: C4 (4 fold cyclic)
3D reconstructionResolution: 6.3 Å / Resolution method: FSC 0.143 CUT-OFF / Num. of particles: 20000 / Symmetry type: POINT

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