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- EMDB-33270: Human Cx36/GJD2 (BRIL-fused mutant) gap junction channel in deter... -

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Basic information

Entry
Database: EMDB / ID: EMD-33270
TitleHuman Cx36/GJD2 (BRIL-fused mutant) gap junction channel in detergents at 2.2 Angstroms resolution
Map datasharpened_map
Sample
  • Complex: Cx36-BRIL
    • Protein or peptide: Gap junction delta-2 protein,Soluble cytochrome b562
  • Ligand: 1,2-DIMYRISTOYL-RAC-GLYCERO-3-PHOSPHOCHOLINE
  • Ligand: CHOLESTEROL HEMISUCCINATE
  • Ligand: 1,2-DIPALMITOYL-PHOSPHATIDYL-GLYCEROLE
  • Ligand: water
Keywordsconnexin 36 / Cx36 / Gap Junction Channel / BRIL / MEMBRANE PROTEIN
Function / homology
Function and homology information


Electric Transmission Across Gap Junctions / connexin complex / Gap junction assembly / gap junction channel activity / neuronal action potential / visual perception / electron transport chain / cell-cell signaling / chemical synaptic transmission / periplasmic space ...Electric Transmission Across Gap Junctions / connexin complex / Gap junction assembly / gap junction channel activity / neuronal action potential / visual perception / electron transport chain / cell-cell signaling / chemical synaptic transmission / periplasmic space / electron transfer activity / iron ion binding / heme binding / synapse / plasma membrane
Similarity search - Function
Gap junction delta-2 protein (Cx36) / 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 delta-2 protein (Cx36) / 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 / Cytochrome b562 / Cytochrome b562 / Cytochrome c/b562
Similarity search - Domain/homology
Soluble cytochrome b562 / Gap junction delta-2 protein
Similarity search - Component
Biological speciesHomo sapiens (human)
Methodsingle particle reconstruction / cryo EM / Resolution: 2.2 Å
AuthorsCho HJ / Lee SN / Jeong H / Ryu B / Lee HJ / Woo JS / Lee HH
Funding support Korea, Republic Of, 2 items
OrganizationGrant numberCountry
National Research Foundation (NRF, Korea)2022R1A2B5B02002529 Korea, Republic Of
National Research Foundation (NRF, Korea)2018R1C1B6004447 Korea, Republic Of
CitationJournal: Nat Commun / Year: 2023
Title: Cryo-EM structures of human Cx36/GJD2 neuronal gap junction channel.
Authors: Seu-Na Lee / Hwa-Jin Cho / Hyeongseop Jeong / Bumhan Ryu / Hyuk-Joon Lee / Minsoo Kim / Jejoong Yoo / Jae-Sung Woo / Hyung Ho Lee /
Abstract: Connexin 36 (Cx36) is responsible for signal transmission in electrical synapses by forming interneuronal gap junctions. Despite the critical role of Cx36 in normal brain function, the molecular ...Connexin 36 (Cx36) is responsible for signal transmission in electrical synapses by forming interneuronal gap junctions. Despite the critical role of Cx36 in normal brain function, the molecular architecture of the Cx36 gap junction channel (GJC) is unknown. Here, we determine cryo-electron microscopy structures of Cx36 GJC at 2.2-3.6 Å resolutions, revealing a dynamic equilibrium between its closed and open states. In the closed state, channel pores are obstructed by lipids, while N-terminal helices (NTHs) are excluded from the pore. In the open state with pore-lining NTHs, the pore is more acidic than those in Cx26 and Cx46/50 GJCs, explaining its strong cation selectivity. The conformational change during channel opening also includes the α-to-π-helix transition of the first transmembrane helix, which weakens the protomer-protomer interaction. Our structural analyses provide high resolution information on the conformational flexibility of Cx36 GJC and suggest a potential role of lipids in the channel gating.
History
DepositionApr 20, 2022-
Header (metadata) releaseMar 22, 2023-
Map releaseMar 22, 2023-
UpdateOct 16, 2024-
Current statusOct 16, 2024Processing site: PDBj / Status: Released

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Structure visualization

Supplemental images

emd_33270.png

Downloads & links

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Map

FileDownload / File: emd_33270.map.gz / Format: CCP4 / Size: 600.7 MB / Type: IMAGE STORED AS FLOATING POINT NUMBER (4 BYTES)
Annotationsharpened_map
Projections & slices

Image control

Size
Brightness
Contrast
Others
AxesZ (Sec.)Y (Row.)X (Col.)
0.68 Å/pix.
x 540 pix.
= 364.5 Å		0.68 Å/pix.
x 540 pix.
= 364.5 Å		0.68 Å/pix.
x 540 pix.
= 364.5 Å

Surface

Projections

Slices (1/3)

Slices (1/2)

Slices (2/3)

Images are generated by Spider.

Voxel sizeX=Y=Z: 0.675 Å
Density

Histogram

Histogram (log scale)
Contour LevelBy AUTHOR: 0.45
Minimum - Maximum-3.7389934 - 6.3313518
Average (Standard dev.)0.0000752069 (±0.15548573)
SymmetrySpace group: 1
Details

EMDB XML:

Map geometry
Axis orderXYZ
Origin000
Dimensions540540540
Spacing540540540
CellA=B=C: 364.5 Å
α=β=γ: 90.0 °

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Supplemental data

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Half map: half map A

Fileemd_33270_half_map_1.map
Annotationhalf_map_A
Projections & Slices
AxesZYX

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Slices (1/2)
Density Histograms

Histogram

Histogram (log scale)

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Half map: half map B

Fileemd_33270_half_map_2.map
Annotationhalf_map_B
Projections & Slices
AxesZYX

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Histogram

Histogram (log scale)

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Sample components

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Entire : Cx36-BRIL

EntireName: Cx36-BRIL
Components
  • Complex: Cx36-BRIL
    • Protein or peptide: Gap junction delta-2 protein,Soluble cytochrome b562
  • Ligand: 1,2-DIMYRISTOYL-RAC-GLYCERO-3-PHOSPHOCHOLINE
  • Ligand: CHOLESTEROL HEMISUCCINATE
  • Ligand: 1,2-DIPALMITOYL-PHOSPHATIDYL-GLYCEROLE
  • Ligand: water

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Supramolecule #1: Cx36-BRIL

SupramoleculeName: Cx36-BRIL / type: complex / ID: 1 / Parent: 0 / Macromolecule list: #1
Source (natural)Organism: Homo sapiens (human)

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Macromolecule #1: Gap junction delta-2 protein,Soluble cytochrome b562

MacromoleculeName: Gap junction delta-2 protein,Soluble cytochrome b562 / type: protein_or_peptide / ID: 1
Details: The chimeric protein of Cx36 (UNP residues 1-108), BRIL (UNP residues 23-123 with mutation M29W, H124I, R128L), Cx36 (UNP residues 188-321), linker and tags
Number of copies: 12 / Enantiomer: LEVO
Source (natural)Organism: Homo sapiens (human)
Molecular weightTheoretical: 40.999199 KDa
Recombinant expressionOrganism: Spodoptera frugiperda (fall armyworm)
SequenceString: MGEWTILERL LEAAVQQHST MIGRILLTVV VIFRILIVAI VGETVYDDEQ TMFVCNTLQP GCNQACYDRA FPISHIRYWV FQIIMVCTP SLCFITYSVH QSAKQRERRA DLEDNWETLN DNLKVIEKAD NAAQVKDALT KMRAAALDAQ KATPPKLEDK S PDSPEMKD ...String:
MGEWTILERL LEAAVQQHST MIGRILLTVV VIFRILIVAI VGETVYDDEQ TMFVCNTLQP GCNQACYDRA FPISHIRYWV FQIIMVCTP SLCFITYSVH QSAKQRERRA DLEDNWETLN DNLKVIEKAD NAAQVKDALT KMRAAALDAQ KATPPKLEDK S PDSPEMKD FRHGFDILVG QIDDALKLAN EGKVKEAQAA AEQLKTTRNA YIQKYLKLRR QEGISRFYII QVVFRNALEI GF LVGQYFL YGFSVPGLYE CNRYPCIKEV ECYVSRPTEK TVFLVFMFAV SGICVVLNLA ELNHLGWRKI KLAVRGAQAK RKS IYEIRN KDLPRVSVPN FGRTQSSDSA YVSRDYKDDD DK

UniProtKB: Gap junction delta-2 protein, Soluble cytochrome b562, Gap junction delta-2 protein

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Macromolecule #2: 1,2-DIMYRISTOYL-RAC-GLYCERO-3-PHOSPHOCHOLINE

MacromoleculeName: 1,2-DIMYRISTOYL-RAC-GLYCERO-3-PHOSPHOCHOLINE / type: ligand / ID: 2 / Number of copies: 72 / Formula: MC3
Molecular weightTheoretical: 677.933 Da
Chemical component information

ChemComp-MC3:
1,2-DIMYRISTOYL-RAC-GLYCERO-3-PHOSPHOCHOLINE / phospholipid*YM

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Macromolecule #3: CHOLESTEROL HEMISUCCINATE

MacromoleculeName: CHOLESTEROL HEMISUCCINATE / type: ligand / ID: 3 / Number of copies: 24 / Formula: Y01
Molecular weightTheoretical: 486.726 Da
Chemical component information

ChemComp-Y01:
CHOLESTEROL HEMISUCCINATE

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Macromolecule #4: 1,2-DIPALMITOYL-PHOSPHATIDYL-GLYCEROLE

MacromoleculeName: 1,2-DIPALMITOYL-PHOSPHATIDYL-GLYCEROLE / type: ligand / ID: 4 / Number of copies: 12 / Formula: LHG
Molecular weightTheoretical: 722.97 Da
Chemical component information

ChemComp-LHG:
1,2-DIPALMITOYL-PHOSPHATIDYL-GLYCEROLE / phospholipid*YM

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Macromolecule #5: water

MacromoleculeName: water / type: ligand / ID: 5 / Number of copies: 444 / Formula: HOH
Molecular weightTheoretical: 18.015 Da
Chemical component information

ChemComp-HOH:
WATER

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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: 7.5
VitrificationCryogen name: ETHANE

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Electron microscopy

MicroscopeFEI TITAN KRIOS
Image recordingFilm or detector model: FEI FALCON III (4k x 4k) / Average electron dose: 40.0 e/Å2
Electron beamAcceleration voltage: 300 kV / Electron source: FIELD EMISSION GUN
Electron opticsIllumination mode: FLOOD BEAM / Imaging mode: BRIGHT FIELD / Nominal defocus max: 3.0 µm / Nominal defocus min: 1.0 µm
Experimental equipment
Model: Titan Krios / Image courtesy: FEI Company

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Image processing

Startup modelType of model: NONE
Final reconstructionResolution.type: BY AUTHOR / Resolution: 2.2 Å / Resolution method: FSC 0.143 CUT-OFF / Number images used: 70095
Initial angle assignmentType: MAXIMUM LIKELIHOOD
Final angle assignmentType: MAXIMUM LIKELIHOOD

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