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- EMDB-33397: Hemichannel-focused structure of C-terminal truncated connexin43/... -

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Entry
Database: EMDB / ID: EMD-33397
TitleHemichannel-focused structure of C-terminal truncated connexin43/Cx43/GJA1 gap junction intercellular channel in POPE nanodiscs (FIN conformation)
Map dataunsharpened map
Sample
  • Complex: Dodecameric complex of C-terminal deletion mutant of human Cx43/GJA1 (Cx43-M257) in lipid nanodiscs (POPE)
    • Protein or peptide: Gap junction alpha-1 protein
KeywordsCx43 / Connexin / Gap junction channel / Gating mechanism / Membrane protein
Function / homology
Function and homology information


gap junction channel activity involved in cardiac conduction electrical coupling / negative regulation of gonadotropin secretion / positive regulation of striated muscle tissue development / milk ejection reflex / positive regulation of morphogenesis of an epithelium / positive regulation of mesodermal cell differentiation / atrial ventricular junction remodeling / cell communication by chemical coupling / cell communication by electrical coupling / columnar/cuboidal epithelial cell maturation ...gap junction channel activity involved in cardiac conduction electrical coupling / negative regulation of gonadotropin secretion / positive regulation of striated muscle tissue development / milk ejection reflex / positive regulation of morphogenesis of an epithelium / positive regulation of mesodermal cell differentiation / atrial ventricular junction remodeling / cell communication by chemical coupling / cell communication by electrical coupling / columnar/cuboidal epithelial cell maturation / neuroblast migration / gap junction hemi-channel activity / negative regulation of trophoblast cell migration / monoatomic ion transmembrane transporter activity / microtubule-based transport / regulation of bone remodeling / regulation of ventricular cardiac muscle cell membrane depolarization / SARS-CoV-2 targets PDZ proteins in cell-cell junction / gap junction channel activity involved in cell communication by electrical coupling / contractile muscle fiber / glutathione transmembrane transporter activity / Oligomerization of connexins into connexons / Transport of connexins along the secretory pathway / Microtubule-dependent trafficking of connexons from Golgi to the plasma membrane / gap junction assembly / atrial cardiac muscle cell action potential / cellular response to pH / Regulation of gap junction activity / cardiac conduction system development / regulation of atrial cardiac muscle cell membrane depolarization / Golgi-associated vesicle membrane / connexin complex / Formation of annular gap junctions / fascia adherens / Gap junction degradation / cell communication by electrical coupling involved in cardiac conduction / cell-cell contact zone / gap junction / bone remodeling / Gap junction assembly / skeletal muscle tissue regeneration / gap junction channel activity / export across plasma membrane / adult heart development / regulation of bone mineralization / regulation of ventricular cardiac muscle cell membrane repolarization / glutamate secretion / tight junction / blood vessel morphogenesis / lens development in camera-type eye / intermediate filament / xenobiotic transport / embryonic digit morphogenesis / maintenance of blood-brain barrier / positive regulation of stem cell proliferation / heart looping / RHOJ GTPase cycle / beta-tubulin binding / RHOQ GTPase cycle / efflux transmembrane transporter activity / establishment of mitotic spindle orientation / intercalated disc / lateral plasma membrane / alpha-tubulin binding / T cell proliferation / positive regulation of vascular associated smooth muscle cell proliferation / protein tyrosine kinase binding / tubulin binding / neuron migration / bone development / negative regulation of cell growth / beta-catenin binding / osteoblast differentiation / male gonad development / cellular response to amyloid-beta / protein localization / cell junction / cell-cell signaling / positive regulation of cold-induced thermogenesis / heart development / monoatomic ion transmembrane transport / scaffold protein binding / spermatogenesis / positive regulation of canonical NF-kappaB signal transduction / in utero embryonic development / membrane raft / apical plasma membrane / Golgi membrane / negative regulation of gene expression / intracellular membrane-bounded organelle / signaling receptor binding / focal adhesion / endoplasmic reticulum membrane / positive regulation of gene expression / Golgi apparatus / signal transduction / mitochondrion / nucleoplasm / nucleus / plasma membrane
Similarity search - Function
Gap junction alpha-1 protein (Cx43) / Gap junction alpha-1 protein (Cx43), C-terminal / Gap junction alpha-1 protein (Cx43), alpha helix domain superfamily / Gap junction alpha-1 protein (Cx43) / Connexin, C-terminal / Connexin / Connexin, N-terminal / Connexin, conserved site / Gap junction protein, cysteine-rich domain / Connexin, N-terminal domain superfamily ...Gap junction alpha-1 protein (Cx43) / Gap junction alpha-1 protein (Cx43), C-terminal / Gap junction alpha-1 protein (Cx43), alpha helix domain superfamily / Gap junction alpha-1 protein (Cx43) / Connexin, C-terminal / Connexin / Connexin, N-terminal / Connexin, conserved site / Gap junction protein, cysteine-rich domain / Connexin, N-terminal domain superfamily / Connexin / Connexins signature 1. / Connexins signature 2. / Connexin homologues / Gap junction channel protein cysteine-rich domain
Similarity search - Domain/homology
Gap junction alpha-1 protein
Similarity search - Component
Biological speciesHomo sapiens (human)
Methodsingle particle reconstruction / cryo EM / Resolution: 3.7 Å
AuthorsLee HJ / Cha HJ / Jeong H / Lee SN / Lee CW / Woo JS
Funding support Korea, Republic Of, 1 items
OrganizationGrant numberCountry
National Research Foundation (NRF, Korea)NRF-2018R1C1B6004447 Korea, Republic Of
CitationJournal: Nat Commun / Year: 2023
Title: Conformational changes in the human Cx43/GJA1 gap junction channel visualized using cryo-EM.
Authors: Hyuk-Joon Lee / Hyung Jin Cha / Hyeongseop Jeong / Seu-Na Lee / Chang-Won Lee / Minsoo Kim / Jejoong Yoo / Jae-Sung Woo /
Abstract: Connexin family proteins assemble into hexameric hemichannels in the cell membrane. The hemichannels dock together between two adjacent membranes to form gap junction intercellular channels (GJIChs). ...Connexin family proteins assemble into hexameric hemichannels in the cell membrane. The hemichannels dock together between two adjacent membranes to form gap junction intercellular channels (GJIChs). We report the cryo-electron microscopy structures of Cx43 GJICh, revealing the dynamic equilibrium state of various channel conformations in detergents and lipid nanodiscs. We identify three different N-terminal helix conformations of Cx43-gate-covering (GCN), pore-lining (PLN), and flexible intermediate (FIN)-that are randomly distributed in purified GJICh particles. The conformational equilibrium shifts to GCN by cholesteryl hemisuccinates and to PLN by C-terminal truncations and at varying pH. While GJIChs that mainly comprise GCN protomers are occluded by lipids, those containing conformationally heterogeneous protomers show markedly different pore sizes. We observe an α-to-π-helix transition in the first transmembrane helix, which creates a side opening to the membrane in the FIN and PLN conformations. This study provides basic structural information to understand the mechanisms of action and regulation of Cx43 GJICh.
History
DepositionMay 7, 2022-
Header (metadata) releaseJan 25, 2023-
Map releaseJan 25, 2023-
UpdateOct 30, 2024-
Current statusOct 30, 2024Processing site: PDBj / Status: Released

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Structure visualization

Supplemental images

emd_33397.png

Downloads & links

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Map

FileDownload / File: emd_33397.map.gz / Format: CCP4 / Size: 64 MB / Type: IMAGE STORED AS FLOATING POINT NUMBER (4 BYTES)
Annotationunsharpened map
Projections & slices

Image control

Size
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AxesZ (Sec.)Y (Row.)X (Col.)
0.85 Å/pix.
x 256 pix.
= 217.088 Å		0.85 Å/pix.
x 256 pix.
= 217.088 Å		0.85 Å/pix.
x 256 pix.
= 217.088 Å

Surface

Projections

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Images are generated by Spider.

Voxel sizeX=Y=Z: 0.848 Å
Density

Histogram

Histogram (log scale)
Contour LevelBy AUTHOR: 0.005
Minimum - Maximum-0.009938417 - 0.028929545
Average (Standard dev.)0.000118919605 (±0.0011434577)
SymmetrySpace group: 1
Details

EMDB XML:

Map geometry
Axis orderXYZ
Origin000
Dimensions256256256
Spacing256256256
CellA=B=C: 217.088 Å
α=β=γ: 90.0 °

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Supplemental data

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Half map: half map

Fileemd_33397_half_map_1.map
Annotationhalf map
Projections & Slices
AxesZYX

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Slices (1/2)
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Histogram

Histogram (log scale)

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Half map: half map

Fileemd_33397_half_map_2.map
Annotationhalf map
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Sample components

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Entire : Dodecameric complex of C-terminal deletion mutant of human Cx43/G...

EntireName: Dodecameric complex of C-terminal deletion mutant of human Cx43/GJA1 (Cx43-M257) in lipid nanodiscs (POPE)
Components
  • Complex: Dodecameric complex of C-terminal deletion mutant of human Cx43/GJA1 (Cx43-M257) in lipid nanodiscs (POPE)
    • Protein or peptide: Gap junction alpha-1 protein

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Supramolecule #1: Dodecameric complex of C-terminal deletion mutant of human Cx43/G...

SupramoleculeName: Dodecameric complex of C-terminal deletion mutant of human Cx43/GJA1 (Cx43-M257) in lipid nanodiscs (POPE)
type: complex / ID: 1 / Parent: 0 / Macromolecule list: all
Details: Hemichannel of Human Cx43-M257 gap junction channel with flexible-intermediate NTH conformation
Source (natural)Organism: Homo sapiens (human)

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Macromolecule #1: Gap junction alpha-1 protein

MacromoleculeName: Gap junction alpha-1 protein / type: protein_or_peptide / ID: 1 / Number of copies: 6 / Enantiomer: LEVO
Source (natural)Organism: Homo sapiens (human)
Molecular weightTheoretical: 29.396547 KDa
Recombinant expressionOrganism: Homo sapiens (human)
SequenceString: MGDWSALGKL LDKVQAYSTA GGKVWLSVLF IFRILLLGTA VESAWGDEQS AFRCNTQQPG CENVCYDKSF PISHVRFWVL QIIFVSVPT LLYLAHVFYV MRKEEKLNKK EEELKVAQTD GVNVDMHLKQ IEIKKFKYGI EEHGKVKMRG GLLRTYIISI L FKSIFEVA ...String:
MGDWSALGKL LDKVQAYSTA GGKVWLSVLF IFRILLLGTA VESAWGDEQS AFRCNTQQPG CENVCYDKSF PISHVRFWVL QIIFVSVPT LLYLAHVFYV MRKEEKLNKK EEELKVAQTD GVNVDMHLKQ IEIKKFKYGI EEHGKVKMRG GLLRTYIISI L FKSIFEVA FLLIQWYIYG FSLSAVYTCK RDPCPHQVDC FLSRPTEKTI FIIFMLVVSL VSLALNIIEL FYVFFKGVKD RV KGKSDPY HATSGALSPA

UniProtKB: Gap junction alpha-1 protein

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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

BufferpH: 8
VitrificationCryogen name: ETHANE

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Electron microscopy

MicroscopeFEI TITAN KRIOS
Image recordingFilm or detector model: GATAN K3 BIOQUANTUM (6k x 4k) / Average electron dose: 72.42 e/Å2
Electron beamAcceleration voltage: 300 kV / Electron source: FIELD EMISSION GUN
Electron opticsIllumination mode: FLOOD BEAM / Imaging mode: BRIGHT FIELD / Nominal defocus max: 2.4 µm / Nominal defocus min: 0.9 µm
Experimental equipment
Model: Titan Krios / Image courtesy: FEI Company

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Image processing

Startup modelType of model: OTHER
Final reconstructionResolution.type: BY AUTHOR / Resolution: 3.7 Å / Resolution method: FSC 0.143 CUT-OFF / Number images used: 128883
Initial angle assignmentType: MAXIMUM LIKELIHOOD
Final angle assignmentType: MAXIMUM LIKELIHOOD
FSC plot (resolution estimation)

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