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- PDB-7zk3: Structure of 1PBC- and calcium-bound mTMEM16A(ac) chloride channe... -

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Basic information

Entry
Database: PDB / ID: 7zk3
TitleStructure of 1PBC- and calcium-bound mTMEM16A(ac) chloride channel at 2.85 A resolution
ComponentsAnoctamin-1
KeywordsMEMBRANE PROTEIN / calcium-activated chloride channel / anoctamin-1
Function / homology
Function and homology information


glial cell projection elongation / trachea development / iodide transmembrane transporter activity / iodide transport / mucus secretion / intracellularly calcium-gated chloride channel activity / cellular response to peptide / Stimuli-sensing channels / voltage-gated chloride channel activity / chloride transport ...glial cell projection elongation / trachea development / iodide transmembrane transporter activity / iodide transport / mucus secretion / intracellularly calcium-gated chloride channel activity / cellular response to peptide / Stimuli-sensing channels / voltage-gated chloride channel activity / chloride transport / chloride channel activity / positive regulation of insulin secretion involved in cellular response to glucose stimulus / detection of temperature stimulus involved in sensory perception of pain / chloride channel complex / chloride transmembrane transport / regulation of membrane potential / cell projection / establishment of localization in cell / presynaptic membrane / cellular response to heat / phospholipase C-activating G protein-coupled receptor signaling pathway / apical plasma membrane / external side of plasma membrane / signaling receptor binding / glutamatergic synapse / protein homodimerization activity / identical protein binding / metal ion binding / plasma membrane
Similarity search - Function
Anoctamin, dimerisation domain / Anoctamin, dimerisation domain / Anoctamin / : / Calcium-activated chloride channel
Similarity search - Domain/homology
Chem-JRF / Anoctamin-1
Similarity search - Component
Biological speciesMus musculus (house mouse)
MethodELECTRON MICROSCOPY / single particle reconstruction / cryo EM / Resolution: 2.85 Å
AuthorsLam, A.K.M. / Rutz, S. / Dutzler, R.
Funding supportEuropean Union, Switzerland, 2items
OrganizationGrant numberCountry
European Research Council (ERC)339116European Union
University of ZurichFK-18-048 Switzerland
CitationJournal: Nat Commun / Year: 2022
Title: Inhibition mechanism of the chloride channel TMEM16A by the pore blocker 1PBC.
Authors: Andy K M Lam / Sonja Rutz / Raimund Dutzler /
Abstract: TMEM16A, a calcium-activated chloride channel involved in multiple cellular processes, is a proposed target for diseases such as hypertension, asthma, and cystic fibrosis. Despite these therapeutic ...TMEM16A, a calcium-activated chloride channel involved in multiple cellular processes, is a proposed target for diseases such as hypertension, asthma, and cystic fibrosis. Despite these therapeutic promises, its pharmacology remains poorly understood. Here, we present a cryo-EM structure of TMEM16A in complex with the channel blocker 1PBC and a detailed functional analysis of its inhibition mechanism. A pocket located external to the neck region of the hourglass-shaped pore is responsible for open-channel block by 1PBC and presumably also by its structural analogs. The binding of the blocker stabilizes an open-like conformation of the channel that involves a rearrangement of several pore helices. The expansion of the outer pore enhances blocker sensitivity and enables 1PBC to bind at a site within the transmembrane electric field. Our results define the mechanism of inhibition and gating and will facilitate the design of new, potent TMEM16A modulators.
History
DepositionApr 12, 2022Deposition site: PDBE / Processing site: PDBE
Revision 1.0May 25, 2022Provider: repository / Type: Initial release
Revision 1.1Jun 1, 2022Group: 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_PubMed / _citation.title / _citation_author.identifier_ORCID

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Structure visualization

Structure viewerMolecule:
MolmilJmol/JSmol

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Assembly

Deposited unit
A: Anoctamin-1
B: Anoctamin-1
hetero molecules


Theoretical massNumber of molelcules
Total (without water)222,91310
Polymers222,1182
Non-polymers7958
Water00
1


  • Idetical with deposited unit
  • defined by author&software
  • Evidence: electron microscopy
TypeNameSymmetry operationNumber
identity operation1_5551
Buried area2120 Å2
ΔGint-91 kcal/mol
Surface area72970 Å2
MethodPISA
Noncrystallographic symmetry (NCS)NCS domain:
IDEns-IDDetails
d_1ens_1chain "A"
d_2ens_1chain "B"

NCS domain segments:
Dom-IDComponent-IDEns-IDBeg label comp-IDEnd label comp-IDLabel asym-IDLabel seq-ID
d_11ens_1METARGA1 - 721
d_12ens_1CACAB
d_13ens_1CACAC
d_14ens_111D
d_21ens_1METARGF1 - 721
d_22ens_1CACAG
d_23ens_1CACAH
d_24ens_111I

NCS oper: (Code: givenMatrix: (-0.999980068895, -0.0038673588816, -0.00499052590843), (0.0038441086586, -0.999981753054, 0.004660084584), (-0.00500845706608, 0.00464080757951, 0.99997668886)Vector: ...NCS oper: (Code: given
Matrix: (-0.999980068895, -0.0038673588816, -0.00499052590843), (0.0038441086586, -0.999981753054, 0.004660084584), (-0.00500845706608, 0.00464080757951, 0.99997668886)
Vector: 397.132612679, 393.950871491, 0.26164429329)

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Components

#1: Protein Anoctamin-1 / Transmembrane protein 16A


Mass: 111058.992 Da / Num. of mol.: 2
Source method: isolated from a genetically manipulated source
Source: (gene. exp.) Mus musculus (house mouse) / Gene: Ano1, Tmem16a / Cell line (production host): GnTI- HEK293S / Production host: Homo sapiens (human) / References: UniProt: Q8BHY3
#2: Chemical
ChemComp-CA / CALCIUM ION


Mass: 40.078 Da / Num. of mol.: 6 / Source method: obtained synthetically / Formula: Ca / Feature type: SUBJECT OF INVESTIGATION
#3: Chemical ChemComp-JRF / 1-Hydroxy-3-(trifluoromethyl)pyrido[1,2-a]benzimidazole-4-carbonitrile / 1-Hydroxy-3-trifluoromethyl-benzo[4,5]imidazo[1,2-a]pyridine-4-carbonitrile


Mass: 277.201 Da / Num. of mol.: 2 / Source method: obtained synthetically / Formula: C13H6F3N3O / 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: mouse TMEM16A ac splice variant in complex with 1PBC and calcium
Type: ORGANELLE OR CELLULAR COMPONENT / Entity ID: #1 / Source: RECOMBINANT
Source (natural)Organism: Mus musculus (house mouse)
Source (recombinant)Organism: Homo sapiens (human) / Cell: GnTI- HEK293S
Buffer solutionpH: 7.4
SpecimenEmbedding applied: NO / Shadowing applied: NO / Staining applied: NO / Vitrification applied: YES
Specimen supportGrid type: Quantifoil R1.2/1.3
VitrificationInstrument: FEI VITROBOT MARK IV / Cryogen name: ETHANE-PROPANE

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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 magnification: 130000 X / Nominal defocus max: 2400 nm / Nominal defocus min: 1000 nm / Alignment procedure: COMA FREE
Specimen holderCryogen: NITROGEN / Specimen holder model: FEI TITAN KRIOS AUTOGRID HOLDER
Image recordingElectron dose: 61 e/Å2 / Film or detector model: GATAN K3 BIOQUANTUM (6k x 4k)

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Processing

Software
NameVersionClassificationNB
phenix.real_space_refine1.19.2_4158refinement
PHENIX1.19.2_4158refinement
CTF correctionType: PHASE FLIPPING AND AMPLITUDE CORRECTION
3D reconstructionResolution: 2.85 Å / Resolution method: FSC 0.143 CUT-OFF / Num. of particles: 101813 / Symmetry type: POINT
RefinementCross valid method: NONE
Stereochemistry target values: GeoStd + Monomer Library + CDL v1.2
Displacement parametersBiso mean: 36.56 Å2
Refine LS restraints
Refine-IDTypeDev idealNumber
ELECTRON MICROSCOPYf_bond_d0.002112198
ELECTRON MICROSCOPYf_angle_d0.503316506
ELECTRON MICROSCOPYf_chiral_restr0.03581802
ELECTRON MICROSCOPYf_plane_restr0.00322060
ELECTRON MICROSCOPYf_dihedral_angle_d3.70531594
Refine LS restraints NCSType: NCS constraints / Rms dev position: 0.000706321995647 Å

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