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- PDB-7cn1: Cryo-EM structure of K+-bound hERG channel in the presence of ast... -

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Entry
Database: PDB / ID: 7cn1
TitleCryo-EM structure of K+-bound hERG channel in the presence of astemizole
Componentspotassium channel
KeywordsTRANSPORT PROTEIN / potassium channel
Biological speciesHomo sapiens (human)
MethodELECTRON MICROSCOPY / single particle reconstruction / cryo EM / Resolution: 3.7 Å
AuthorsAsai, T. / Adachi, N. / Moriya, T. / Kawasaki, M. / Suzuki, K. / Senda, T. / Murata, T.
Funding support Japan, 3items
OrganizationGrant numberCountry
Japan Agency for Medical Research and Development (AMED)JP20am0101071 Japan
Japan Agency for Medical Research and Development (AMED)JP20am0101083 Japan
Japan Society for the Promotion of Science (JSPS)18H05425 Japan
CitationJournal: Structure / Year: 2020
Title: Cryo-EM structure of K-bound hERG channel complexed with the blocker astemizole.
Authors: Tatsuki Asai / Naruhiko Adachi / Toshio Moriya / Hideyuki Oki / Takamitsu Maru / Masato Kawasaki / Kano Suzuki / Sisi Chen / Ryohei Ishii / Kazuko Yonemori / Shigeru Igaki / Satoshi Yasuda / ...Authors: Tatsuki Asai / Naruhiko Adachi / Toshio Moriya / Hideyuki Oki / Takamitsu Maru / Masato Kawasaki / Kano Suzuki / Sisi Chen / Ryohei Ishii / Kazuko Yonemori / Shigeru Igaki / Satoshi Yasuda / Satoshi Ogasawara / Toshiya Senda / Takeshi Murata /
Abstract: The hERG channel is a voltage-gated potassium channel involved in cardiac repolarization. Off-target hERG inhibition by drugs has become a critical issue in the pharmaceutical industry. The three- ...The hERG channel is a voltage-gated potassium channel involved in cardiac repolarization. Off-target hERG inhibition by drugs has become a critical issue in the pharmaceutical industry. The three-dimensional structure of the hERG channel was recently reported at 3.8-Å resolution using cryogenic electron microscopy (cryo-EM). However, the drug inhibition mechanism remains unclear because of the scarce structural information regarding the drug- and potassium-bound hERG channels. In this study, we obtained the cryo-EM density map of potassium-bound hERG channel complexed with astemizole, a well-known hERG inhibitor that increases risk of potentially fatal arrhythmia, at 3.5-Å resolution. The structure suggested that astemizole inhibits potassium conduction by binding directly below the selectivity filter. Furthermore, we propose a possible binding model of astemizole to the hERG channel and provide insights into the unusual sensitivity of hERG to several drugs.
Validation Report
SummaryFull reportAbout validation report
History
DepositionJul 29, 2020Deposition site: PDBJ / Processing site: PDBJ
Revision 1.0Jan 20, 2021Provider: repository / Type: Initial release

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Assembly

Deposited unit
A: potassium channel
B: potassium channel
C: potassium channel
D: potassium channel
hetero molecules


Theoretical massNumber of molelcules
Total (without water)367,2867
Polymers367,1684
Non-polymers1173
Water0
1


TypeNameSymmetry operationNumber
identity operation1_5551
Buried area8290 Å2
ΔGint-66 kcal/mol
Surface area44240 Å2

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Components

#1: Protein
potassium channel /


Mass: 91792.086 Da / Num. of mol.: 4
Source method: isolated from a genetically manipulated source
Source: (gene. exp.) Homo sapiens (human) / Plasmid: BacMam expression vector / Production host: Homo sapiens (human) / Strain (production host): Expi293F cells
#2: Chemical ChemComp-K / POTASSIUM ION / Potassium


Mass: 39.098 Da / Num. of mol.: 3 / Source method: obtained synthetically / Formula: K / Feature type: SUBJECT OF INVESTIGATION
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: potassium channel / Type: ORGANELLE OR CELLULAR COMPONENT / Details: homo-tetramer / Entity ID: #1 / Source: RECOMBINANT
Source (natural)Organism: Homo sapiens (human)
Source (recombinant)Organism: Homo sapiens (human) / Cell: Expi293F cells / Plasmid: BacMam expression vector
Buffer solutionpH: 7.4
SpecimenConc.: 2 mg/ml / Embedding applied: NO / Shadowing applied: NO / Staining applied: NO / Vitrification applied: YES / Details: This sample was mono-disperse.
Specimen supportDetails: The grid was washed by acetone prior to use. / Grid material: COPPER / Grid mesh size: 300 divisions/in. / Grid type: Quantifoil R1.2/1.3
VitrificationInstrument: FEI VITROBOT MARK IV / Cryogen name: ETHANE / Humidity: 100 % / Chamber temperature: 291 K / Details: Blotting time was 5 seconds (blot force 10)

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Electron microscopy imaging

Experimental equipment
Model: Talos Arctica / Image courtesy: FEI Company
MicroscopyModel: FEI TALOS ARCTICA
Electron gunElectron source: FIELD EMISSION GUN / Accelerating voltage: 200 kV / Illumination mode: FLOOD BEAM
Electron lensMode: BRIGHT FIELDBright-field microscopy / Nominal magnification: 120000 X / Nominal defocus max: 2000 nm / Nominal defocus min: 800 nm / Cs: 2.7 mm / C2 aperture diameter: 50 µm
Specimen holderCryogen: NITROGEN
Image recordingAverage exposure time: 55.55 sec. / Electron dose: 50 e/Å2 / Detector mode: COUNTING / Film or detector model: FEI FALCON III (4k x 4k) / Num. of grids imaged: 1 / Num. of real images: 1865

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Processing

SoftwareName: PHENIX / Version: 1.16_3549: / Classification: refinement
EM software
IDNameVersionCategory
1RELION3particle selection
2EPUimage acquisition
4CTFFIND4CTF correction
7Coot0.8.9.2model fitting
9PHENIX1.16model refinement
10RELION3initial Euler assignment
11RELION3final Euler assignment
12RELION3classification
13RELION33D reconstruction
CTF correctionType: PHASE FLIPPING AND AMPLITUDE CORRECTION
Particle selectionNum. of particles selected: 930193
SymmetryPoint symmetry: C1 (asymmetric)
3D reconstructionResolution: 3.7 Å / Resolution method: FSC 0.143 CUT-OFF / Num. of particles: 102128 / Symmetry type: POINT
Atomic model buildingProtocol: AB INITIO MODEL / Space: REAL
Refine LS restraints
Refinement-IDTypeDev idealNumber
ELECTRON MICROSCOPYf_bond_d0.0086440
ELECTRON MICROSCOPYf_angle_d0.7648762
ELECTRON MICROSCOPYf_dihedral_angle_d6.8023655
ELECTRON MICROSCOPYf_chiral_restr0.0461053
ELECTRON MICROSCOPYf_plane_restr0.0041028

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