The basic property of Lys385 is important for potentiation of the human ?1 glycine receptor by ethanol.

ABSTRACT: Ethanol alters the function of several members of the Cys-loop ligand-gated ion channel superfamily. Recent studies have shown that the sensitivity of the ?1 glycine receptor (GlyR) to ethanol can be affected by the state of G protein activation mediated by the interaction of G?? with intracellular amino acids in the GlyR. Here, we evaluated the physicochemical property of Lys385 that contributes to ethanol modulation by using mutagenesis, patch-clamp, and biochemical techniques. A conserved substitution (K385R) did not affect either the apparent glycine EC?? (40 ± 1 versus 41 ± 0.5 ?M) or the ethanol-induced potentiation (53 ± 5 versus 46 ± 5%) of the human ?1 GlyR. On the other hand, replacement of this residue with glutamic acid (K385E), an acidic amino acid, reduced the potentiation of the GlyR to 10 ± 1%. Furthermore, mutations with a hydrophobic leucine (K385L), a hydrogen bond donor glutamine (K385Q), or a neutral residue (K385A) also reduced ethanol modulation. Finally, substitution by a large and hydrophobic residue (K385F) and deletion of 385 (Lys385_) reduced ethanol modulation to 10 ± 4 and 17 ± 0.4%, respectively. Experiments using dynamic cysteine substitution with a methanethiosulfonate reagent and homology modeling indicate that the basic property and the position of Lys385, probably because of its interaction with G??, is critical for ethanol potentiation of the receptor.

SUBMITTER: Castro PA 

PROVIDER: S-EPMC3263957 | biostudies-literature | 2012 Feb

REPOSITORIES: biostudies-literature