Modelling of Ca2+-promoted structural effects in wild type and post-translationally modified Connexin26

Connexins (Cx) are a class of membrane proteins important for auditory function, intercellular signalling and skin biology. Although the presence of concentration of calcium ions is known to work as a trigger for the Cx functionality, the structural changes induced by calcium binding still need to be well elucidated. In this computational study, we have explored the structural effects promoted by Ca2+ on both the wild type (Cx26WT) and on two post-translationally modified Connexin 26 (Cx26): Cx26E42-47γ, which contains two glutamates (E42 and E47) that are γ-carboxylated and Cx26R75m, where a key arginine (R75) is N-monomethylated. These modified amino acids, whose forcefield parameters have been developed in this work, alter Cx26 structure around the Ca2+coordination site. Structural changes were assessed from the analysis of molecular dynamics (MD) simulations. We observed a strict relation between the chemical properties of the post-translational modifications and significantly different responses of Cx26 to Ca2+-binding, while charge-adding modifications have destabilising effects upon calcium coordination, the uncharged ones share the same structural properties of the wild-type counterpart. Overall, these findings suggest the critical role of the electrostatic network flanking the Ca2+ coordination site in maintaining the native tertiary and quaternary structures.