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- EMDB-27774: Structure of LRRC8C-LRRC8A(IL125) Chimera, Class 5 -

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Basic information

Entry
Database: EMDB / ID: EMD-27774
TitleStructure of LRRC8C-LRRC8A(IL125) Chimera, Class 5
Map data
Sample
  • Complex: LRRC8C-LRRC8A(IL125) chimera
    • Protein or peptide: Volume-regulated anion channel subunit LRRC8C,Volume-regulated anion channel subunit LRRC8A
KeywordsLRRC8C / LRRC8A / SWELL / VRAC / Chimera / TRANSPORT PROTEIN
Function / homology
Function and homology information


pre-B cell differentiation / Miscellaneous transport and binding events / volume-sensitive anion channel activity / taurine transmembrane transport / aspartate transmembrane transport / cyclic-GMP-AMP transmembrane transporter activity / cyclic-GMP-AMP transmembrane import across plasma membrane / monoatomic anion transmembrane transport / cellular response to osmotic stress / protein hexamerization ...pre-B cell differentiation / Miscellaneous transport and binding events / volume-sensitive anion channel activity / taurine transmembrane transport / aspartate transmembrane transport / cyclic-GMP-AMP transmembrane transporter activity / cyclic-GMP-AMP transmembrane import across plasma membrane / monoatomic anion transmembrane transport / cellular response to osmotic stress / protein hexamerization / cell volume homeostasis / monoatomic anion transport / response to osmotic stress / monoatomic ion channel complex / fat cell differentiation / intracellular glucose homeostasis / positive regulation of myoblast differentiation / chloride transmembrane transport / positive regulation of insulin secretion / spermatogenesis / lysosomal membrane / endoplasmic reticulum 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, 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
Volume-regulated anion channel subunit LRRC8A / Volume-regulated anion channel subunit LRRC8C
Similarity search - Component
Biological speciesHomo sapiens (human)
Methodsingle particle reconstruction / cryo EM / Resolution: 4.0 Å
AuthorsTakahashi H / Yamada T / Denton JS / Strange K / Karakas E
Funding support United States, 1 items
OrganizationGrant numberCountry
National Institutes of Health/National Institute of Diabetes and Digestive and Kidney Disease (NIH/NIDDK)DK51610 United States
CitationJournal: Elife / Year: 2023
Title: Cryo-EM structures of an LRRC8 chimera with native functional properties reveal heptameric assembly.
Authors: Hirohide Takahashi / Toshiki Yamada / Jerod S Denton / Kevin Strange / Erkan Karakas /
Abstract: Volume-regulated anion channels (VRACs) mediate volume regulatory Cl and organic solute efflux from vertebrate cells. VRACs are heteromeric assemblies of LRRC8A-E proteins with unknown ...Volume-regulated anion channels (VRACs) mediate volume regulatory Cl and organic solute efflux from vertebrate cells. VRACs are heteromeric assemblies of LRRC8A-E proteins with unknown stoichiometries. Homomeric LRRC8A and LRRC8D channels have a small pore, hexameric structure. However, these channels are either non-functional or exhibit abnormal regulation and pharmacology, limiting their utility for structure-function analyses. We circumvented these limitations by developing novel homomeric LRRC8 chimeric channels with functional properties consistent with those of native VRAC/LRRC8 channels. We demonstrate here that the LRRC8C-LRRC8A(IL1) chimera comprising LRRC8C and 25 amino acids unique to the first intracellular loop (IL1) of LRRC8A has a heptameric structure like that of homologous pannexin channels. Unlike homomeric LRRC8A and LRRC8D channels, heptameric LRRC8C-LRRC8A(IL1) channels have a large-diameter pore similar to that estimated for native VRACs, exhibit normal DCPIB pharmacology, and have higher permeability to large organic anions. Lipid-like densities are located between LRRC8C-LRRC8A(IL1) subunits and occlude the channel pore. Our findings provide new insights into VRAC/LRRC8 channel structure and suggest that lipids may play important roles in channel gating and regulation.
History
DepositionAug 2, 2022-
Header (metadata) releaseMar 22, 2023-
Map releaseMar 22, 2023-
UpdateJun 12, 2024-
Current statusJun 12, 2024Processing site: RCSB / Status: Released

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Structure visualization

Supplemental images

emd_27774.png

Downloads & links

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Map

FileDownload / File: emd_27774.map.gz / Format: CCP4 / Size: 178 MB / Type: IMAGE STORED AS FLOATING POINT NUMBER (4 BYTES)
Projections & slices

Image control

Size
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AxesZ (Sec.)Y (Row.)X (Col.)
1.1 Å/pix.
x 360 pix.
= 396. Å		1.1 Å/pix.
x 360 pix.
= 396. Å		1.1 Å/pix.
x 360 pix.
= 396. Å

Surface

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Images are generated by Spider.

Voxel sizeX=Y=Z: 1.1 Å
Density

Histogram

Histogram (log scale)
Contour LevelBy AUTHOR: 0.15
Minimum - Maximum-0.5502354 - 0.97780293
Average (Standard dev.)-0.00074188836 (±0.020346355)
SymmetrySpace group: 1
Details

EMDB XML:

Map geometry
Axis orderXYZ
Origin000
Dimensions360360360
Spacing360360360
CellA=B=C: 396.0 Å
α=β=γ: 90.0 °

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Supplemental data

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Additional map: Unsharpened map.

Fileemd_27774_additional_1.map
AnnotationUnsharpened map.
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Half map: #2

Fileemd_27774_half_map_1.map
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Half map: #1

Fileemd_27774_half_map_2.map
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Sample components

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Entire : LRRC8C-LRRC8A(IL125) chimera

EntireName: LRRC8C-LRRC8A(IL125) chimera
Components
  • Complex: LRRC8C-LRRC8A(IL125) chimera
    • Protein or peptide: Volume-regulated anion channel subunit LRRC8C,Volume-regulated anion channel subunit LRRC8A

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Supramolecule #1: LRRC8C-LRRC8A(IL125) chimera

SupramoleculeName: LRRC8C-LRRC8A(IL125) chimera / type: complex / ID: 1 / Parent: 0 / Macromolecule list: all / Details: Heptameric LRRC8C-LRRC8A(IL125) chimera.
Source (natural)Organism: Homo sapiens (human)
Molecular weightTheoretical: 700 KDa

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Macromolecule #1: Volume-regulated anion channel subunit LRRC8C,Volume-regulated an...

MacromoleculeName: Volume-regulated anion channel subunit LRRC8C,Volume-regulated anion channel subunit LRRC8A
type: protein_or_peptide / ID: 1 / Number of copies: 7 / Enantiomer: LEVO
Source (natural)Organism: Homo sapiens (human)
Molecular weightTheoretical: 94.747695 KDa
Recombinant expressionOrganism: Spodoptera frugiperda (fall armyworm)
SequenceString: MIPVTEFRQF SEQQPAFRVL KPWWDVFTDY LSVAMLMIGV FGCTLQVMQD KIICLPKRVQ PAQNHSSLSN VSQAVASTTP LPPPKPSPA NPITVEMKGL KTDLDLQQYS FINQMCYERA LHWYAKYFPY LVLIHTLVFM LCSNFWFKFP GSSSKIEHFI S ILGKCFDS ...String:
MIPVTEFRQF SEQQPAFRVL KPWWDVFTDY LSVAMLMIGV FGCTLQVMQD KIICLPKRVQ PAQNHSSLSN VSQAVASTTP LPPPKPSPA NPITVEMKGL KTDLDLQQYS FINQMCYERA LHWYAKYFPY LVLIHTLVFM LCSNFWFKFP GSSSKIEHFI S ILGKCFDS PWTTRALSEV SGEDSDPKPA FSKMNGSMDK KSSTVSEDVE GSLVNSQSLK SIPEKFVVDK STAGALDKKE GE QAKALFE KVKKFRLHVE EGDILYAMYV RQTVLKVIKF LIIIAYNSAL VSKVQFTVDC NVDIQDMTGY KNFSCNHTMA HLF SKLSFC YLCFVSIYGL TCLYTLYWLF YRSLREYSFE YVRQETGIDD IPDVKNDFAF MLHMIDQYDP LYSKRFAVFL SEVS ENKLK QLNLNNEWTP DKLRQKLQTN AHNRLELPLI MLSGLPDTVF EITELQSLKL EIIKNVMIPA TIAQLDNLQE LSLHQ CSVK IHSAALSFLK ENLKVLSVKF DDMRELPPWM YGLRNLEELY LVGSLSHDIS RNVTLESLRD LKSLKILSIK SNVSKI PQA VVDVSSHLQK MCIHNDGTKL VMLNNLKKMT NLTELELVHC DLERIPHAVF SLLSLQELDL KENNLKSIEE IVSFQHL RK LTVLKLWHNS ITYIPEHIKK LTSLERLSFS HNKIEVLPSH LFLCNKIRYL DLSYNDIRFI PPEIGVLQSL QYFSITCN V ESLPDELYFC KKLKTLKIGK NSLSVLSPKI GNLLFLSYLD VKGNHFEILP PELGDCRALK RAGLVVEDAL FETLPSDVR EQMKTEENLY FQGAAAGDYK DDDDK

UniProtKB: Volume-regulated anion channel subunit LRRC8C, Volume-regulated anion channel subunit LRRC8A, Volume-regulated anion channel subunit LRRC8C

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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

Concentration3.0 mg/mL
BufferpH: 8
Component:
ConcentrationFormulaName
0.15 MNaClSodium chloride
0.05 MTris-HClTris buffer pH 8.0
0.005 %LMNGLMNG detergent
GridModel: UltrAuFoil R1.2/1.3 / Material: GOLD / Mesh: 300 / Support film - Material: GOLD / Support film - topology: HOLEY
VitrificationCryogen name: ETHANE / Chamber humidity: 100 % / Chamber temperature: 281 K / Instrument: FEI VITROBOT MARK IV

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Electron microscopy

MicroscopeFEI TITAN KRIOS
Image recordingFilm or detector model: GATAN K3 BIOQUANTUM (6k x 4k) / Number real images: 3198 / Average electron dose: 54.0 e/Å2
Electron beamAcceleration voltage: 300 kV / Electron source: FIELD EMISSION GUN
Electron opticsIllumination mode: FLOOD BEAM / Imaging mode: BRIGHT FIELD / Nominal defocus max: 2.2 µm / Nominal defocus min: 0.8 µm
Experimental equipment
Model: Titan Krios / Image courtesy: FEI Company

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Image processing

Particle selectionNumber selected: 846122
Startup modelType of model: NONE
Final reconstructionNumber classes used: 1 / Applied symmetry - Point group: C1 (asymmetric) / Resolution.type: BY AUTHOR / Resolution: 4.0 Å / Resolution method: FSC 0.143 CUT-OFF / Software - Name: cryoSPARC / Number images used: 85591
Initial angle assignmentType: ANGULAR RECONSTITUTION
Final angle assignmentType: ANGULAR RECONSTITUTION
Final 3D classificationNumber classes: 6
FSC plot (resolution estimation)

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Atomic model buiding 1

RefinementSpace: REAL / Protocol: FLEXIBLE FIT
Output model

PDB-8dxr:
Structure of LRRC8C-LRRC8A(IL125) Chimera, Class 5

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