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- EMDB-25384: Structure of the human proton-activated chloride channel ASOR in ... -

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Basic information

Entry
Database: EMDB / ID: EMD-25384
TitleStructure of the human proton-activated chloride channel ASOR in resting conformation
Map datafinal map
Sample
  • Complex: proton activated chloride channel TMEM206 in resting conformation
    • Protein or peptide: Proton-activated chloride channel
  • Ligand: 2-acetamido-2-deoxy-beta-D-glucopyranose
Function / homologypH-gated chloride channel activity / TMEM206 protein / TMEM206 protein family / chloride transport / chloride channel complex / cell surface / plasma membrane / Proton-activated chloride channel
Function and homology information
Biological speciesHomo sapiens (human)
Methodsingle particle reconstruction / cryo EM / Resolution: 2.6 Å
AuthorsLong SB / Wang C / Delgado B
Funding support United States, 2 items
OrganizationGrant numberCountry
National Institutes of Health/National Institute of General Medical Sciences (NIH/NIGMS)R35GM131921 United States
National Institutes of Health/National Cancer Institute (NIH/NCI)P30CA008748 United States
CitationJournal: Sci Adv / Year: 2022
Title: Gating choreography and mechanism of the human proton-activated chloride channel ASOR.
Authors: Chongyuan Wang / Maya M Polovitskaya / Bryce D Delgado / Thomas J Jentsch / Stephen B Long /
Abstract: The proton-activated chloride channel ASOR (TMEM206/PAC) permeates anions across cellular membranes in response to acidification, thereby enhancing acid-induced cell death and regulating endocytosis. ...The proton-activated chloride channel ASOR (TMEM206/PAC) permeates anions across cellular membranes in response to acidification, thereby enhancing acid-induced cell death and regulating endocytosis. The molecular mechanisms of pH-dependent control are not understood, in part because structural information for an activated conformation of ASOR is lacking. Here, we reconstitute function from purified protein and present a 3.1-Å-resolution cryo-electron microscopy structure of human ASOR at acidic pH in an activated conformation. The work contextualizes a previous acidic pH structure as a desensitized conformation. Combined with electrophysiological studies and high-resolution structures of resting and desensitized states, the work reveals mechanisms of proton sensing and ion pore gating. Clusters of extracellular acidic residues function as pH sensors and coalesce when protonated. Ensuing conformational changes induce metamorphosis of transmembrane helices to fashion an ion conduction pathway unique to the activated conformation. The studies identify a new paradigm of channel gating in this ubiquitous ion channel.
History
DepositionNov 5, 2021-
Header (metadata) releaseFeb 23, 2022-
Map releaseFeb 23, 2022-
UpdateFeb 23, 2022-
Current statusFeb 23, 2022Processing site: RCSB / Status: Released

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Structure visualization

Movie
  • Surface view with section colored by density value
  • Surface level: 0.1
  • Imaged by UCSF Chimera
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  • Surface view colored by cylindrical radius
  • Surface level: 0.1
  • Imaged by UCSF Chimera
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  • Surface view with fitted model
  • Atomic models: PDB-7sqg
  • Surface level: 0.1
  • Imaged by UCSF Chimera
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Structure viewerEM map:
SurfViewMolmilJmol/JSmol
Supplemental images

emd_25384.png

Downloads & links

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Map

FileDownload / File: emd_25384.map.gz / Format: CCP4 / Size: 216 MB / Type: IMAGE STORED AS FLOATING POINT NUMBER (4 BYTES)
Annotationfinal map
Voxel sizeX=Y=Z: 0.826 Å
Density
Contour LevelBy AUTHOR: 0.1 / Movie #1: 0.1
Minimum - Maximum-0.0017951481 - 1.9678134
Average (Standard dev.)0.00043959302 (±0.015616784)
SymmetrySpace group: 1
Details

EMDB XML:

Map geometry
Axis orderXYZ
Origin000
Dimensions384384384
Spacing384384384
CellA=B=C: 317.184 Å
α=β=γ: 90.0 °

CCP4 map header:

modeImage stored as Reals
Å/pix. X/Y/Z0.8260.8260.826
M x/y/z384384384
origin x/y/z0.0000.0000.000
length x/y/z317.184317.184317.184
α/β/γ90.00090.00090.000
MAP C/R/S123
start NC/NR/NS000
NC/NR/NS384384384
D min/max/mean-0.0021.9680.000

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Supplemental data

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Sample components

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Entire : proton activated chloride channel TMEM206 in resting conformation

EntireName: proton activated chloride channel TMEM206 in resting conformation
Components
  • Complex: proton activated chloride channel TMEM206 in resting conformation
    • Protein or peptide: Proton-activated chloride channel
  • Ligand: 2-acetamido-2-deoxy-beta-D-glucopyranose

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Supramolecule #1: proton activated chloride channel TMEM206 in resting conformation

SupramoleculeName: proton activated chloride channel TMEM206 in resting conformation
type: complex / ID: 1 / Parent: 0 / Macromolecule list: #1
Source (natural)Organism: Homo sapiens (human)
Recombinant expressionOrganism: Homo sapiens (human) / Recombinant cell: Expi293, Invitrogen

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Macromolecule #1: Proton-activated chloride channel

MacromoleculeName: Proton-activated chloride channel / type: protein_or_peptide / ID: 1 / Number of copies: 3 / Enantiomer: LEVO
Source (natural)Organism: Homo sapiens (human)
Molecular weightTheoretical: 40.092047 KDa
Recombinant expressionOrganism: Homo sapiens (human)
SequenceString: MIRQERSTSY QELSEELVQV VENSELADEQ DKETVRVQGP GILPGLDSES ASSSIRFSKA CLKNVFSVLL IFIYLLLMAV AVFLVYRTI TDFREKLKHP VMSVSYKEVD RYDAPGIALY PGQAQLLSCK HHYEVIPPLT SPGQPGDMNC TTQRINYTDP F SNQTVKSA ...String:
MIRQERSTSY QELSEELVQV VENSELADEQ DKETVRVQGP GILPGLDSES ASSSIRFSKA CLKNVFSVLL IFIYLLLMAV AVFLVYRTI TDFREKLKHP VMSVSYKEVD RYDAPGIALY PGQAQLLSCK HHYEVIPPLT SPGQPGDMNC TTQRINYTDP F SNQTVKSA LIVQGPREVK KRELVFLQFR LNKSSEDFSA IDYLLFSSFQ EFLQSPNRVG FMQACESAYS SWKFSGGFRT WV KMSLVKT KEEDGREAVE FRQETSVVNY IDQRPAAKKS AQLFFVVFEW KDPFIQKVQD IVTANPWNTI ALLCGAFLAL FKA AEFAKL SIKWMIKIRK RYLKRRGQAT SHIS

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Macromolecule #2: 2-acetamido-2-deoxy-beta-D-glucopyranose

MacromoleculeName: 2-acetamido-2-deoxy-beta-D-glucopyranose / type: ligand / ID: 2 / Number of copies: 12 / Formula: NAG
Molecular weightTheoretical: 221.208 Da
Chemical component information

ChemComp-NAG:
2-acetamido-2-deoxy-beta-D-glucopyranose / N-Acetylglucosamine

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Experimental details

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Structure determination

Methodcryo EM
Processingsingle particle reconstruction
Aggregation stateparticle

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Sample preparation

Concentration5.0 mg/mL
BufferpH: 7.5
Component:
ConcentrationNameFormula
20.0 mMHEPES
150.0 mMsodium chlorideNaClSodium chloride
GridModel: UltrAuFoil R1.2/1.3 / Support film - Material: CARBON / Support film - topology: HOLEY ARRAY
VitrificationCryogen name: ETHANE / Chamber humidity: 100 % / Chamber temperature: 298 K / Instrument: FEI VITROBOT MARK IV / Details: 2 second blot, blot force of 0.
DetailsMonodisperse sample

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Electron microscopy

MicroscopeFEI TITAN KRIOS
Electron beamAcceleration voltage: 300 kV / Electron source: FIELD EMISSION GUN
Electron opticsIllumination mode: FLOOD BEAM / Imaging mode: BRIGHT FIELDBright-field microscopy / Nominal defocus max: -3.0 µm / Nominal defocus min: -1.0 µm / Nominal magnification: 22500
Sample stageSpecimen holder model: FEI TITAN KRIOS AUTOGRID HOLDER / Cooling holder cryogen: NITROGEN
Image recordingFilm or detector model: GATAN K3 BIOQUANTUM (6k x 4k) / Detector mode: SUPER-RESOLUTION / Digitization - Frames/image: 1-40 / Number grids imaged: 1 / Number real images: 60374 / Average exposure time: 1.8 sec. / Average electron dose: 48.0 e/Å2
Experimental equipment
Model: Titan Krios / Image courtesy: FEI Company

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Image processing

Particle selectionNumber selected: 612580
CTF correctionSoftware - Name: Gctf (ver. 1.06)
Startup modelType of model: INSILICO MODEL
In silico model: Ab initio model created using CryoSPARC v3.1.0
Initial angle assignmentType: RANDOM ASSIGNMENT
Software: (Name: cryoSPARC (ver. v3.1.0), RELION (ver. 3.1))
Final 3D classificationNumber classes: 1 / Software - Name: cryoSPARC (ver. v3.1.0)
Final angle assignmentType: MAXIMUM LIKELIHOOD / Software - Name: cryoSPARC (ver. v3.1.0)
Final reconstructionResolution.type: BY AUTHOR / Resolution: 2.6 Å / Resolution method: FSC 0.143 CUT-OFF / Software - Name: cryoSPARC (ver. v3.1.0) / Number images used: 60374

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Atomic model buiding 1

RefinementSpace: REAL / Protocol: AB INITIO MODEL / Overall B value: 30 / Target criteria: Correlation coefficient
Output model

PDB-7sqg:
Structure of the human proton-activated chloride channel ASOR in resting conformation

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