Molecular requirements for ethanol differential allosteric modulation of ligand-gated ion channels based on selective G beta gamma modulation

Gonzalo E. Yevenes, University of Concepcion
Gustavo Moraga-Cid, University of Concepcion
Ariel Avila, University of Concepcion
Leonardo Guzman, Unviersity of Concepcion
Maximiliano Figueroa, University of Concepcion
Robert Peoples, Marquette University
Luis G. Aguayo, University of Concepcion

Journal of Biological Chemistry, Vol. 285, No. 39 (September 2010): 30203-30213. DOI.


It is now believed that the allosteric modulation produced by ethanol in glycine receptors (GlyRs) depends on alcohol binding to discrete sites within the protein structure. Thus, the differential ethanol sensitivity of diverse GlyR isoforms and mutants was explained by the presence of specific residues in putative alcohol pockets. Here, we demonstrate that ethanol sensitivity in two LGIC members, the GlyR adult alpha1 and embryonic alpha2 subunits, can be modified through selective mutations that rescued or impaired Gbetagamma modulation. Even though that both isoforms were able to physically interact with Gbetagamma, only the alpha1 GlyR was functionally modulated by Gbetagamma and pharmacological ethanol concentrations. Remarkably, the simultaneous switching of two transmembrane and a single extracellular residue in alpha2 GlyRs was enough to generate GlyRs modulated by Gbetagamma and low ethanol concentrations. Interestingly, while we found that these TM residues were different to those in the alcohol binding site, the extracellular residue was recently implicated in conformational changes important to generate a pre-open activated state that precedes ion channel gating. Thus, these results support the idea that the differential ethanol sensitivity of these two GlyR isoforms rests on conformational changes in transmembrane and extracellular residues within the ion channel structure rather than in differences in alcohol binding pockets. Our results describe the molecular basis for the differential ethanol sensitivity of two LGIC members based on selective Gbetagamma modulation and provide a new mechanistic framework for allosteric modulations of abuse drugs.