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- PDB-7xqg: Hemichannel-focused structure of C-terminal truncated connexin43/... -

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Basic information

Entry
Database: PDB / ID: 7xqg
TitleHemichannel-focused structure of C-terminal truncated connexin43/Cx43/GJA1 gap junction intercellular channel in POPE nanodiscs (GCN conformation)
ComponentsGap junction alpha-1 protein
KeywordsMEMBRANE PROTEIN / Cx43 / Connexin / Gap junction channel / Gating mechanism
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 / columnar/cuboidal epithelial cell maturation / cell communication by electrical coupling ...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 / columnar/cuboidal epithelial cell maturation / cell communication by electrical coupling / neuroblast migration / gap junction hemi-channel activity / gap junction channel activity involved in cell communication by electrical coupling / negative regulation of trophoblast cell migration / regulation of bone remodeling / microtubule-based transport / monoatomic ion transmembrane transporter activity / SARS-CoV-2 targets PDZ proteins in cell-cell junction / glutathione transmembrane transporter activity / regulation of ventricular cardiac muscle cell membrane depolarization / Oligomerization of connexins into connexons / Transport of connexins along the secretory pathway / contractile muscle fiber / gap junction assembly / Microtubule-dependent trafficking of connexons from Golgi to the plasma membrane / cellular response to pH / atrial cardiac muscle cell action potential / Regulation of gap junction activity / connexin complex / skeletal muscle tissue regeneration / cardiac conduction system development / regulation of atrial cardiac muscle cell membrane depolarization / Formation of annular gap junctions / Golgi-associated vesicle membrane / Gap junction degradation / fascia adherens / Gap junction assembly / bone remodeling / gap junction channel activity / gap junction / cell-cell contact zone / export across plasma membrane / adult heart development / regulation of bone mineralization / glutamate secretion / regulation of ventricular cardiac muscle cell membrane repolarization / xenobiotic transport / tight junction / blood vessel morphogenesis / lens development in camera-type eye / intermediate filament / cell communication by electrical coupling involved in cardiac conduction / embryonic digit morphogenesis / maintenance of blood-brain barrier / beta-tubulin binding / positive regulation of stem cell proliferation / heart looping / RHOJ GTPase cycle / RHOQ GTPase cycle / efflux transmembrane transporter activity / establishment of mitotic spindle orientation / alpha-tubulin binding / intercalated disc / lateral plasma membrane / T cell proliferation / positive regulation of vascular associated smooth muscle cell proliferation / protein tyrosine kinase binding / tubulin binding / monoatomic ion transmembrane transport / neuron migration / bone development / protein localization / negative regulation of cell growth / beta-catenin binding / osteoblast differentiation / cellular response to amyloid-beta / male gonad development / cell-cell signaling / cell junction / positive regulation of cold-induced thermogenesis / heart development / 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 / signaling receptor binding / focal adhesion / intracellular membrane-bounded organelle / 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
PHOSPHATIDYLETHANOLAMINE / CHOLESTEROL HEMISUCCINATE / Gap junction alpha-1 protein
Similarity search - Component
Biological speciesHomo sapiens (human)
MethodELECTRON MICROSCOPY / single particle reconstruction / cryo EM / Resolution: 3.8 Å
AuthorsLee, H.J. / Cha, H.J. / Jeong, H. / Lee, S.N. / Lee, C.W. / Woo, J.S.
Funding support Korea, Republic Of, 1items
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, 2022Deposition site: PDBJ / Processing site: PDBJ
Revision 1.0Jan 25, 2023Provider: repository / Type: Initial release
Revision 1.1Aug 2, 2023Group: Database references / Category: citation / citation_author
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.year

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Structure visualization

Structure viewerMolecule:
MolmilJmol/JSmol

Downloads & links

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Assembly

Deposited unit
A: Gap junction alpha-1 protein
B: Gap junction alpha-1 protein
C: Gap junction alpha-1 protein
D: Gap junction alpha-1 protein
E: Gap junction alpha-1 protein
F: Gap junction alpha-1 protein
hetero molecules


Theoretical massNumber of molelcules
Total (without water)183,70418
Polymers176,3796
Non-polymers7,32512
Water0
1


  • Idetical with deposited unit
  • defined by author
  • Evidence: microscopy
TypeNameSymmetry operationNumber
identity operation1_5551
Noncrystallographic symmetry (NCS)NCS domain:
IDEns-IDDetails
d_1ens_1chain "A"
d_2ens_1chain "B"
d_3ens_1chain "C"
d_4ens_1chain "D"
d_5ens_1chain "E"
d_6ens_1chain "F"

NCS domain segments:
Dom-IDComponent-IDEns-IDBeg label comp-IDEnd label comp-IDLabel asym-IDLabel seq-ID
d_11ens_1GLYASPA1 - 201
d_12ens_1PTYPTYB
d_13ens_1Y01Y01C
d_21ens_1GLYASPD1 - 201
d_22ens_1PTYPTYE
d_23ens_1Y01Y01F
d_31ens_1GLYASPG1 - 201
d_32ens_1PTYPTYH
d_33ens_1Y01Y01I
d_41ens_1GLYASPJ1 - 201
d_42ens_1PTYPTYK
d_43ens_1Y01Y01L
d_51ens_1GLYASPM1 - 201
d_52ens_1PTYPTYN
d_53ens_1Y01Y01O
d_61ens_1GLYASPP1 - 201
d_62ens_1PTYPTYQ
d_63ens_1Y01Y01R

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Components

#1: Protein
Gap junction alpha-1 protein / Connexin-43 / Cx43 / Gap junction 43 kDa heart protein


Mass: 29396.547 Da / Num. of mol.: 6
Source method: isolated from a genetically manipulated source
Source: (gene. exp.) Homo sapiens (human) / Gene: GJA1, GJAL / Production host: Homo sapiens (human) / References: UniProt: P17302
#2: Chemical
ChemComp-PTY / PHOSPHATIDYLETHANOLAMINE / Phosphatidylethanolamine


Mass: 734.039 Da / Num. of mol.: 6 / Source method: obtained synthetically / Formula: C40H80NO8P / Comment: phospholipid*YM
#3: Chemical
ChemComp-Y01 / CHOLESTEROL HEMISUCCINATE


Mass: 486.726 Da / Num. of mol.: 6 / Source method: obtained synthetically / Formula: C31H50O4 / 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: Dodecameric complex of C-terminal deletion mutant of human Cx43/GJA1 (Cx43-M257) in lipid nanodiscs (POPE)
Type: COMPLEX
Details: Hemichannel of Human Cx43-M257 gap junction channel with gate-covering NTH conformation
Entity ID: #1 / Source: RECOMBINANT
Source (natural)Organism: Homo sapiens (human)
Source (recombinant)Organism: Homo sapiens (human)
Buffer solutionpH: 8
SpecimenEmbedding applied: NO / Shadowing applied: NO / Staining applied: NO / Vitrification applied: YES
VitrificationCryogen name: ETHANE

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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 FIELDBright-field microscopy / Nominal defocus max: 2400 nm / Nominal defocus min: 900 nm
Image recordingElectron dose: 72.42 e/Å2 / Film or detector model: GATAN K3 BIOQUANTUM (6k x 4k)

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Processing

Software
NameVersionClassification
phenix.real_space_refine1.20.1_4487refinement
PHENIX1.20.1_4487refinement
CTF correctionType: PHASE FLIPPING AND AMPLITUDE CORRECTION
3D reconstructionResolution: 3.8 Å / Resolution method: FSC 0.143 CUT-OFF / Num. of particles: 18108 / Symmetry type: POINT
RefinementCross valid method: NONE
Stereochemistry target values: GeoStd + Monomer Library + CDL v1.2
Displacement parametersBiso mean: 176.75 Å2
Refine LS restraints
Refine-IDTypeDev idealNumber
ELECTRON MICROSCOPYf_bond_d0.001910476
ELECTRON MICROSCOPYf_angle_d0.450714190
ELECTRON MICROSCOPYf_chiral_restr0.03171602
ELECTRON MICROSCOPYf_plane_restr0.00241674
ELECTRON MICROSCOPYf_dihedral_angle_d11.18963744
Refine LS restraints NCS
Ens-IDDom-IDAuth asym-IDRefine-IDTypeRms dev position (Å)
ens_1d_2AELECTRON MICROSCOPYNCS constraints2.62908714935E-13
ens_1d_3AELECTRON MICROSCOPYNCS constraints3.67607201342E-13
ens_1d_4AELECTRON MICROSCOPYNCS constraints2.86818915313E-13
ens_1d_5AELECTRON MICROSCOPYNCS constraints3.17459677995E-13
ens_1d_6AELECTRON MICROSCOPYNCS constraints2.82402204052E-13

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