Structural elements of the gamma-aminobutyric acid type A receptor conferring subtype selectivity for benzodiazepine site ligands

gamma-aminobutyric acid type A (GABA(A)) receptors comprise a subfamily of ligand-gated ion channels whose activity can be modulated by ligands acting at the benzodiazepine binding site on the receptor. The benzodiazepine bin ding site was characterized using a site-directed mutagenesis strategy in w hich amino acids of the alpha(5) subunit were substituted by their correspo nding alpha(1) residues. Given the high affinity and selectivity of alpha(1 )-containing compared with alpha(5)-containing GABA(A) receptors for zolpid em, mutated alpha(5) subunits were co-expressed with beta(2) and gamma(2) s ubunits, and the affinity of recombinant receptors for zolpidem was measure d. One alpha(5) mutant (bearing P162T, E200G, and T204S) exhibited properti es similar to that of the alpha(1) subunit, notably high affinity zolpidem binding and potentiation by zolpidem of GABA-induced chloride current. Two of these mutations, alpha(5)P162T and alpha(5)E200G, might alter binding po cket conformation, whereas alpha(5)T204S probably permits formation of a hy drogen bond with a proton acceptor in zolpidem, These three amino acid subs titutions also influenced receptor affinity for CL218872, Our data thus sug gest that corresponding amino acids of the alpha(1) subunit, particularly a lpha(1)-Ser(204), are the crucial residues influencing ligand selectivity a t the binding pocket of alpha(1)-containing receptors, and a model of this binding pocket is presented.