Received for publication August 7, 2006.
Revised September 27, 2006.
Accepted for publication September 28, 2006.

Structural determinants for high affinity zolpidem binding to GABA-A receptors

Abstract

The imidazopyridine zolpidem (Ambien®) is one of the most commonly prescribed sleep aids in the nation (Rush, 1998). Similar to classical benzodiazepines (BZDs), zolpidem binds at the extracellular N-terminal / interface of the GABA-A receptor (GABAR). However, zolpidem differs significantly from classical BZDs in chemical structure and neuropharmacological properties. Thus, classical BZDs and zolpidem likely have different requirements for high affinity binding to GABARs. To date, three residues 2M57, 2F77 and 2M130, have been identified as necessary for high affinity zolpidem binding (Wingrove et al, 1997; Buhr and Sigel, 1997). Here, we used radioligand binding techniques, 2/1 chimeric subunits (), site-directed mutagenesis, and molecular modeling to identify additional 2 subunit residues important for high affinity zolpidem binding. While 12 receptors containing only the first 161 amino-terminal residues of the 2 subunit bind the classical BZD flunitrazepam with wild-type affinity, zolpidem affinity is decreased ~8 fold. By incrementally restoring 2 subunit sequence, we identified a 7 amino acid stretch in the 2 subunit Loop F region (a.a. 186-192) that is required to confer high affinity zolpidem binding to GABARs. When mapped to a homology model, these seven amino acids comprise part of loop F located at the / interface. Based on in silico zolpidem docking, three residues within loop F, 2E189, 2T193, and 2R194, emerge as being important for stabilizing zolpidem in the BZD binding pocket and likely interact with other loop F residues to maintain the structural integrity of the BZD binding site.

Key words: GABAA, GABAC, Structure-activity relationships and modeling, Func. analysis receptor/ion channel mutants, Benzodiazepines

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