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- PDB-7xzh: Cryo-EM structure of human LRRC8A -

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Basic information

Entry
Database: PDB / ID: 7xzh
TitleCryo-EM structure of human LRRC8A
ComponentsVolume-regulated anion channel subunit LRRC8A
KeywordsMEMBRANE PROTEIN / Volume Regulated Anion Channel / LRRC8A / Cryo-EM / VRAC / N terminal extention
Function / homology
Function and homology information


pre-B cell differentiation / Miscellaneous transport and binding events / volume-sensitive anion channel activity / aspartate transmembrane transport / taurine transmembrane transport / cyclic-GMP-AMP transmembrane transporter activity / cyclic-GMP-AMP transmembrane import across plasma membrane / monoatomic anion transmembrane transport / protein hexamerization / cell volume homeostasis ...pre-B cell differentiation / Miscellaneous transport and binding events / volume-sensitive anion channel activity / aspartate transmembrane transport / taurine transmembrane transport / cyclic-GMP-AMP transmembrane transporter activity / cyclic-GMP-AMP transmembrane import across plasma membrane / monoatomic anion transmembrane transport / protein hexamerization / cell volume homeostasis / monoatomic anion transport / response to osmotic stress / intracellular glucose homeostasis / monoatomic ion channel complex / positive regulation of myoblast differentiation / chloride transmembrane transport / positive regulation of insulin secretion / spermatogenesis / intracellular signal transduction / lysosomal membrane / cell surface / identical protein binding / membrane / plasma membrane / cytoplasm
Similarity search - Function
LRRC8, pannexin-like TM region / Pannexin-like TM region of LRRC8 / : / : / Leucine-rich repeat region / Leucine-rich repeat, SDS22-like subfamily / Leucine rich repeat / Leucine-rich repeat, typical subtype / Leucine-rich repeats, typical (most populated) subfamily / Leucine-rich repeat profile. ...LRRC8, pannexin-like TM region / Pannexin-like TM region of LRRC8 / : / : / Leucine-rich repeat region / Leucine-rich repeat, SDS22-like subfamily / Leucine rich repeat / Leucine-rich repeat, typical subtype / Leucine-rich repeats, typical (most populated) subfamily / Leucine-rich repeat profile. / Leucine-rich repeat / Leucine-rich repeat domain superfamily
Similarity search - Domain/homology
CHOLESTEROL / Chem-P5S / Chem-POV / Volume-regulated anion channel subunit LRRC8A
Similarity search - Component
Biological speciesHomo sapiens (human)
MethodELECTRON MICROSCOPY / single particle reconstruction / cryo EM / Resolution: 2.78 Å
AuthorsLiu, H. / Liao, J.
Funding support China, 2items
OrganizationGrant numberCountry
Other government2017YFA0504800 China
Other government2018ZX09711003-003-003 China
CitationJournal: Cell Rep / Year: 2023
Title: Structural insights into anion selectivity and activation mechanism of LRRC8 volume-regulated anion channels.
Authors: Heng Liu / Maya M Polovitskaya / Linlin Yang / Meiling Li / Hongyue Li / Zhen Han / Jianguo Wu / Qiansen Zhang / Thomas J Jentsch / Jun Liao /
Abstract: Volume-regulated anion channels (VRACs) are hexamers of LRRC8 proteins that are crucial for cell volume regulation. N termini (NTs) of the obligatory LRRC8A subunit modulate VRACs activation and ion ...Volume-regulated anion channels (VRACs) are hexamers of LRRC8 proteins that are crucial for cell volume regulation. N termini (NTs) of the obligatory LRRC8A subunit modulate VRACs activation and ion selectivity, but the underlying mechanisms remain poorly understood. Here, we report a 2.8-Å cryo-electron microscopy structure of human LRRC8A that displays well-resolved NTs. Amino-terminal halves of NTs fold back into the pore and constrict the permeation path, thereby determining ion selectivity together with an extracellular selectivity filter with which it works in series. They also interact with pore-surrounding helices and support their compact arrangement. The C-terminal halves of NTs interact with intracellular loops that are crucial for channel activation. Molecular dynamics simulations indicate that low ionic strength increases NT mobility and expands the radial distance between pore-surrounding helices. Our work suggests an unusual pore architecture with two selectivity filters in series and a mechanism for VRAC activation by cell swelling.
History
DepositionJun 2, 2022Deposition site: PDBJ / Processing site: PDBJ
Revision 1.0Jun 7, 2023Provider: repository / Type: Initial release
Revision 1.1Nov 13, 2024Group: Data collection / Structure summary
Category: chem_comp_atom / chem_comp_bond ...chem_comp_atom / chem_comp_bond / em_admin / pdbx_entry_details / pdbx_modification_feature
Item: _em_admin.last_update / _pdbx_entry_details.has_protein_modification
Revision 1.2Dec 18, 2024Group: Data collection / Database references / Category: citation / citation_author / em_admin
Item: _citation.country / _citation.journal_abbrev ..._citation.country / _citation.journal_abbrev / _citation.journal_id_CSD / _citation.journal_id_ISSN / _citation.journal_volume / _citation.page_first / _citation.page_last / _citation.pdbx_database_id_DOI / _citation.pdbx_database_id_PubMed / _citation.title / _citation.year / _em_admin.last_update

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Structure visualization

Structure viewerMolecule:
MolmilJmol/JSmol

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Assembly

Deposited unit
A: Volume-regulated anion channel subunit LRRC8A
B: Volume-regulated anion channel subunit LRRC8A
C: Volume-regulated anion channel subunit LRRC8A
D: Volume-regulated anion channel subunit LRRC8A
E: Volume-regulated anion channel subunit LRRC8A
F: Volume-regulated anion channel subunit LRRC8A
hetero molecules


Theoretical massNumber of molelcules
Total (without water)578,66930
Polymers564,7166
Non-polymers13,95324
Water00
1


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

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Components

#1: Protein
Volume-regulated anion channel subunit LRRC8A / Leucine-rich repeat-containing protein 8A / HsLRRC8A / Swelling protein 1


Mass: 94119.312 Da / Num. of mol.: 6
Source method: isolated from a genetically manipulated source
Source: (gene. exp.) Homo sapiens (human) / Gene: LRRC8A, KIAA1437, LRRC8, SWELL1, UNQ221/PRO247 / Cell line (production host): HEK293 / Production host: Homo sapiens (human) / Variant (production host): GNTI- / References: UniProt: Q8IWT6
#2: Chemical
ChemComp-P5S / O-[(R)-{[(2R)-2,3-bis(octadecanoyloxy)propyl]oxy}(hydroxy)phosphoryl]-L-serine / phosphatidyl serine


Mass: 792.075 Da / Num. of mol.: 6 / Source method: obtained synthetically / Formula: C42H82NO10P / Feature type: SUBJECT OF INVESTIGATION
#3: Chemical
ChemComp-POV / (2S)-3-(hexadecanoyloxy)-2-[(9Z)-octadec-9-enoyloxy]propyl 2-(trimethylammonio)ethyl phosphate / POPC


Mass: 760.076 Da / Num. of mol.: 6 / Source method: obtained synthetically / Formula: C42H82NO8P / Comment: phospholipid*YM
#4: Chemical
ChemComp-CLR / CHOLESTEROL


Mass: 386.654 Da / Num. of mol.: 12 / Source method: obtained synthetically / Formula: C27H46O / Feature type: SUBJECT OF INVESTIGATION
Has ligand of interestY
Has protein modificationY

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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: Volume Regulated Anion Channel / Type: COMPLEX / Entity ID: #1 / Source: RECOMBINANT
Molecular weightExperimental value: NO
Source (natural)Organism: Homo sapiens (human)
Source (recombinant)Organism: Homo sapiens (human) / Cell: HEK293-GNTI- / Plasmid: pEGMam
Buffer solutionpH: 8
SpecimenConc.: 16 mg/ml / Embedding applied: NO / Shadowing applied: NO / Staining applied: NO / Vitrification applied: YES
VitrificationCryogen name: ETHANE / Humidity: 100 % / Chamber temperature: 4 K

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Electron microscopy imaging

Experimental equipment
Model: Titan Krios / Image courtesy: FEI Company
MicroscopyModel: FEI TITAN KRIOS
Electron gunElectron source: LAB6 / Accelerating voltage: 300 kV / Illumination mode: FLOOD BEAM
Electron lensMode: DARK FIELD / Nominal defocus max: 2000 nm / Nominal defocus min: 1000 nm / Cs: 2.7 mm / C2 aperture diameter: 70 µm
Image recordingElectron dose: 60 e/Å2 / Film or detector model: GATAN K3 (6k x 4k)

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Processing

CTF correctionType: PHASE FLIPPING AND AMPLITUDE CORRECTION
3D reconstructionResolution: 2.78 Å / Resolution method: FSC 0.143 CUT-OFF / Num. of particles: 31640 / Symmetry type: POINT

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