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Polydisperse molecular architecture of connexin 26/30 heteromeric hemichannels revealed by AFM imaging

ABSTRACT: Gap junctions are arrays of intercellular channels formed by two hemichannels docked end-to-end. Each hemichannel is a hexamer containing the same or different connexin (Cx) isoform. While homomeric Cxs forms have been largely described functionally and structurally, lesser is known about heteromeric Cx channels. To unveil properties of heteromeric Cx channels is challenging considering the high number of potential subunit arrangements and stoichiometries, even for only combining two Cx isoforms. We engineered an HA tag onto Cx26 or Cx30 subunits and imaged hemichannels that were liganded by Fab-epitope antibody fragments. For Cx26-HA/Cx30 or Cx26/Cx30-HA heteromeric channels, the Fab-HA binding distribution followed a binomial distribution with a maximum of 3 Fab-HA bound. Furthermore, imaged Cx26/Cx30-HA triple liganded by Fab-HA showed multiple arrangements which can be derived from the law of total probabilities. Our results indicate a dominant subunit stoichiometry of 3Cx26:3Cx30 with the most abundant subunit arrangement of Cx26-Cx26-Cx30-Cx26-Cx30-Cx30.

INSTRUMENT(S): LTQ Orbitrap XL

ORGANISM(S): Homo Sapiens (human)

TISSUE(S): Cell Culture

SUBMITTER: Marie Alfes

LAB HEAD: Carla Schmidt

PROVIDER: PXD020633 | Pride | 2020-09-04

REPOSITORIES: Pride

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