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- EMDB-18291: Cryo-EM structure of Cx26 solubilised in LMNG - hemichannel class... -
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Open data
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Basic information
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Title | Cryo-EM structure of Cx26 solubilised in LMNG - hemichannel classification - NConst conformation | |||||||||
![]() | Full map locally sharpened in phenix; classification and masked refinement over A-F hemichannel | |||||||||
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![]() | Gap junction Large Pore Channel Carbon dioxide sensitive / MEMBRANE PROTEIN | |||||||||
Function / homology | ![]() Transport of connexons to the plasma membrane / response to human chorionic gonadotropin / gap junction-mediated intercellular transport / epididymis development / gap junction channel activity involved in cell communication by electrical coupling / Oligomerization of connexins into connexons / Transport of connexins along the secretory pathway / gap junction assembly / connexin complex / gap junction ...Transport of connexons to the plasma membrane / response to human chorionic gonadotropin / gap junction-mediated intercellular transport / epididymis development / gap junction channel activity involved in cell communication by electrical coupling / Oligomerization of connexins into connexons / Transport of connexins along the secretory pathway / gap junction assembly / connexin complex / gap junction / Gap junction assembly / gap junction channel activity / astrocyte projection / cellular response to glucagon stimulus / inner ear development / decidualization / endoplasmic reticulum-Golgi intermediate compartment / lateral plasma membrane / response to retinoic acid / cellular response to dexamethasone stimulus / response to ischemia / response to progesterone / sensory perception of sound / transmembrane transport / response to estradiol / cell-cell signaling / cellular response to oxidative stress / cell body / response to lipopolysaccharide / calcium ion binding / perinuclear region of cytoplasm / identical protein binding / plasma membrane Similarity search - Function | |||||||||
Biological species | ![]() | |||||||||
Method | single particle reconstruction / cryo EM / Resolution: 2.3 Å | |||||||||
![]() | Brotherton DH / Savva CG / Cameron AD | |||||||||
Funding support | ![]()
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![]() | ![]() Title: Structures of wild-type and a constitutively closed mutant of connexin26 shed light on channel regulation by CO. Authors: Deborah H Brotherton / Sarbjit Nijjar / Christos G Savva / Nicholas Dale / Alexander David Cameron / ![]() Abstract: Connexins allow intercellular communication by forming gap junction channels (GJCs) between juxtaposed cells. Connexin26 (Cx26) can be regulated directly by CO. This is proposed to be mediated ...Connexins allow intercellular communication by forming gap junction channels (GJCs) between juxtaposed cells. Connexin26 (Cx26) can be regulated directly by CO. This is proposed to be mediated through carbamylation of K125. We show that mutating K125 to glutamate, mimicking the negative charge of carbamylation, causes Cx26 GJCs to be constitutively closed. Through cryo-EM we observe that the K125E mutation pushes a conformational equilibrium towards the channel having a constricted pore entrance, similar to effects seen on raising the partial pressure of CO. In previous structures of connexins, the cytoplasmic loop, important in regulation and where K125 is located, is disordered. Through further cryo-EM studies we trap distinct states of Cx26 and observe density for the cytoplasmic loop. The interplay between the position of this loop, the conformations of the transmembrane helices and the position of the N-terminal helix, which controls the aperture to the pore, provides a mechanism for regulation. #1: ![]() Title: Structures of wild-type and a constitutively closed mutant of connexin26 shed light on channel regulation by CO2 Authors: Brotherton DH / Nijjar S / Savva CG / Dale N / Cameron AD | |||||||||
History |
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Structure visualization
Supplemental images |
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Downloads & links
-EMDB archive
Map data | ![]() | 288.7 MB | ![]() | |
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Header (meta data) | ![]() ![]() | 22.5 KB 22.5 KB | Display Display | ![]() |
FSC (resolution estimation) | ![]() | 15.4 KB | Display | ![]() |
Images | ![]() | 135 KB | ||
Masks | ![]() | 316.2 MB | ![]() | |
Filedesc metadata | ![]() | 6.7 KB | ||
Others | ![]() ![]() ![]() | 250.3 MB 250.8 MB 250.8 MB | ||
Archive directory | ![]() ![]() | HTTPS FTP |
-Related structure data
Related structure data | ![]() 8qa0MC ![]() 8q9zC ![]() 8qa1C ![]() 8qa2C ![]() 8qa3C M: atomic model generated by this map C: citing same article ( |
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Similar structure data | Similarity search - Function & homology ![]() |
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Links
EMDB pages | ![]() ![]() |
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Map
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Annotation | Full map locally sharpened in phenix; classification and masked refinement over A-F hemichannel | ||||||||||||||||||||
Voxel size | X=Y=Z: 0.835 Å | ||||||||||||||||||||
Density |
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Symmetry | Space group: 1 | ||||||||||||||||||||
Details | EMDB XML:
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-Supplemental data
-Mask #1
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Density Histograms |
-Additional map: Full map unsharpened; classification and masked refinement over...
File | emd_18291_additional_1.map | ||||||||||||
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Annotation | Full map unsharpened; classification and masked refinement over A-F hemichannel | ||||||||||||
Projections & Slices |
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Density Histograms |
-Half map: Half map (unsharpened); classification and masked refinement over...
File | emd_18291_half_map_1.map | ||||||||||||
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Annotation | Half map (unsharpened); classification and masked refinement over A-F hemichannel | ||||||||||||
Projections & Slices |
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Density Histograms |
-Half map: Half map (unsharpened); classification and masked refinement over...
File | emd_18291_half_map_2.map | ||||||||||||
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Annotation | Half map (unsharpened); classification and masked refinement over A-F hemichannel | ||||||||||||
Projections & Slices |
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Density Histograms |
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Sample components
-Entire : Connexin 26, 90mmHg PCO2, pH7.4, solubilised in LMNG.
Entire | Name: Connexin 26, 90mmHg PCO2, pH7.4, solubilised in LMNG. |
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Components |
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-Supramolecule #1: Connexin 26, 90mmHg PCO2, pH7.4, solubilised in LMNG.
Supramolecule | Name: Connexin 26, 90mmHg PCO2, pH7.4, solubilised in LMNG. / type: complex / ID: 1 / Parent: 0 / Macromolecule list: #1 |
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Source (natural) | Organism: ![]() |
-Macromolecule #1: Gap junction beta-2 protein
Macromolecule | Name: Gap junction beta-2 protein / type: protein_or_peptide / ID: 1 / Details: Connexin 26 subunit / Number of copies: 12 / Enantiomer: LEVO |
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Source (natural) | Organism: ![]() |
Molecular weight | Theoretical: 26.713674 KDa |
Recombinant expression | Organism: ![]() ![]() |
Sequence | String: MDWGTLQTIL GGVNKHSTSI GKIWLTVLFI FRIMILVVAA KEVWGDEQAD FVCNTLQPGC KNVCYDHYFP ISHIRLWALQ LIFVSTPAL LVAMHVAYRR HEKKRKFIKG EIKSEFKDIE EIKTQKVRIE GSLWWTYTSS IFFRVIFEAA FMYVFYVMYD G FSMQRLVK ...String: MDWGTLQTIL GGVNKHSTSI GKIWLTVLFI FRIMILVVAA KEVWGDEQAD FVCNTLQPGC KNVCYDHYFP ISHIRLWALQ LIFVSTPAL LVAMHVAYRR HEKKRKFIKG EIKSEFKDIE EIKTQKVRIE GSLWWTYTSS IFFRVIFEAA FMYVFYVMYD G FSMQRLVK CNAWPCPNTV DCFVSRPTEK TVFTVFMIAV SGICILLNVT ELCYLLIRYC SGKSKKPVLV PR UniProtKB: Gap junction beta-2 protein |
-Macromolecule #2: PHOSPHATIDYLETHANOLAMINE
Macromolecule | Name: PHOSPHATIDYLETHANOLAMINE / type: ligand / ID: 2 / Number of copies: 24 / Formula: PTY |
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Molecular weight | Theoretical: 734.039 Da |
Chemical component information | ![]() ChemComp-PTY: |
-Macromolecule #3: water
Macromolecule | Name: water / type: ligand / ID: 3 / Number of copies: 342 / Formula: HOH |
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Molecular weight | Theoretical: 18.015 Da |
Chemical component information | ![]() ChemComp-HOH: |
-Experimental details
-Structure determination
Method | cryo EM |
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![]() | single particle reconstruction |
Aggregation state | particle |
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Sample preparation
Concentration | 3.7 mg/mL | |||||||||||||||||||||||||||
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Buffer | pH: 7.4 Component:
Details: The buffer, except LMNG and DTT, was prepared fresh from 10x stock on day of used. The basal buffer was filtered and degassed and DDM and DTT added. The buffer was pH corrected at point of ...Details: The buffer, except LMNG and DTT, was prepared fresh from 10x stock on day of used. The basal buffer was filtered and degassed and DDM and DTT added. The buffer was pH corrected at point of use to pH 7.4 using 15% CO2 in 85% N2. | |||||||||||||||||||||||||||
Grid | Model: UltrAuFoil / Material: GOLD / Pretreatment - Type: GLOW DISCHARGE | |||||||||||||||||||||||||||
Vitrification | Cryogen name: ETHANE-PROPANE / Instrument: FEI VITROBOT MARK I Details: Blot 3 seconds, force 10, 1 blot, skip transfer. 3ul sample, in 15% CO2/85%N2 atmosphere. |
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Electron microscopy
Microscope | FEI TITAN KRIOS |
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Specialist optics | Energy filter - Name: GIF Bioquantum |
Image recording | Film or detector model: GATAN K3 (6k x 4k) / Number real images: 11362 / Average electron dose: 43.0 e/Å2 |
Electron beam | Acceleration voltage: 300 kV / Electron source: ![]() |
Electron optics | Illumination mode: FLOOD BEAM / Imaging mode: BRIGHT FIELD / Nominal defocus max: 2.3000000000000003 µm / Nominal defocus min: 0.8 µm / Nominal magnification: 105000 |
Sample stage | Specimen holder model: FEI TITAN KRIOS AUTOGRID HOLDER |
Experimental equipment | ![]() Model: Titan Krios / Image courtesy: FEI Company |