Backbone amides are determinants of Cl− selectivity in CLC ion channels

• Published in: Nature communications Vol. 13; no. 1; pp. 7508 - 11
• Main Authors: Leisle, Lilia, Lam, Kin, Dehghani-Ghahnaviyeh, Sepehr, Fortea, Eva, Galpin, Jason D., Ahern, Christopher A., Tajkhorshid, Emad, Accardi, Alessio
• Format: Journal Article
• Language: English
• Published: London Nature Publishing Group UK 06.12.2022; Nature Publishing Group; Nature Portfolio
• Subjects: 119/118; 631/45/612/1237; 631/57/2270/1140; 631/57/2283; 631/92/269/1147; 9/74; Amides; Amino acids; Anions; Biological membranes; Chains; Chloride channels; Electrophysiology; Homeostasis; Humanities and Social Sciences; Hydrogen bonding; Hydroxy acids; Ion Channels; Ions; Membranes; Molecular dynamics; multidisciplinary; Science; Science (multidisciplinary); Selectivity
• ISSN: 2041-1723; 2041-1723
• DOI: 10.1038/s41467-022-35279-1

Chloride homeostasis is regulated in all cellular compartments. CLC-type channels selectively transport Cl − across biological membranes. It is proposed that side-chains of pore-lining residues determine Cl − selectivity in CLC-type channels, but their spatial orientation and contributions to selectivity are not conserved. This suggests a possible role for mainchain amides in selectivity. We use nonsense suppression to insert α-hydroxy acids at pore-lining positions in two CLC-type channels, CLC-0 and bCLC-k, thus exchanging peptide-bond amides with ester-bond oxygens which are incapable of hydrogen-bonding. Backbone substitutions functionally degrade inter-anion discrimination in a site-specific manner. The presence of a pore-occupying glutamate side chain modulates these effects. Molecular dynamics simulations show backbone amides determine ion energetics within the bCLC-k pore and how insertion of an α-hydroxy acid alters selectivity. We propose that backbone-ion interactions are determinants of Cl − specificity in CLC channels in a mechanism reminiscent of that described for K + channels. CLC-type channels selectively transport Cl− across biological membranes, but it is unclear how discrimination between anions is maintained. Here, authors use a combination of non-natural amino acid substitutions, electrophysiology, and molecular dynamics simulations to determine Cl − specificity within this family of ion channels.