Mechanism of Cl- selection by a glutamate-gated chloride (GluCl) receptor revealed through mutations in the selectivity filter

To learn about the mechanism of ion charge selectivity by invertebrate glutamate-gated chloride ( GluCl) channels, we swapped segments between the GluCl beta receptor of Caenorhabditis elegans and the vertebrate cationic alpha 7-acetylcholine receptor and monitored anionic/cationic permeability ratios. Complete conversion of the ion charge selectivity in a set of receptor microchimeras indicates that the selectivity filter of the GluCl beta receptor is created by a sequence connecting the first with the second transmembrane segments. A single substitution of a negatively charged residue within this sequence converted the selectivity of the GluCl beta receptor's pore from anionic to cationic. Unexpectedly, elimination of the charge of each basic residue of the selectivity filter, one at a time or concomitantly, moderately reduced the P-Cl/P-Na ratios, but the GluCl beta receptor's mutants retained high capacity to select Cl- over Na+. These results indicate that, unlike the proposed case of anionic Gly- and gamma-aminobutyric acid-gated ion channels, positively charged residues do not play the key role in the selection of ionic charge by the GluCl beta receptor. Taken together with measurements of the effective open pore diameter and with structural modeling, the study presented here collectively indicates that in the most constricted part of the open GluCl beta receptor's channel, Cl- interacts with backbone amides, where it undergoes partial dehydration necessary for traversing the pore.