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

ABSTRACT

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.

KEYWORDS:

• Connexin26
• post-translational modification
• not-standard residues
• molecular dynamics
• ScrewFit

Acknowledgments

The authors acknowledge funding by Università degli Studi di Padova (CPDA127392/12) and Fondazione della Cassa di Risparmio di Padova e Rovigo (Project Modeling and Monitoring Motions in Proteins-M3PC). The calculations were carried out on Eurora at CINECA (Casalecchio di Reno, Italy) thanks to the ISCRA grant NAMASTE ´ (PI: L. O.) and on the computational facilities of the Department of Chemical Sciences (Università degli Studi di Padova).

Disclosure statement

No potential conflict of interest was reported by the authors.

ORCID

Paolo Calligari http://orcid.org/0000-0001-7614-8931

Mauro Torsello http://orcid.org/0000-0003-3123-2296

Marco Bortoli http://orcid.org/0000-0001-5506-6347

Laura Orian http://orcid.org/0000-0002-1673-5111

Antonino Polimeno http://orcid.org/0000-0001-6441-3689