The atypical M2 segment of the beta subunit confers picrotoxinin resistance to inhibitory glycine receptor channels.

Abstract

Purified preparations of the inhibitory glycine receptor (GlyR) contain (α and β subunits, which share homologous primary structures and a common transmembrane topology with other members of the ligand-gated ion channel superfamily. Here, a β subunit-specific antiserum was shown to precipitate the [3H]strychnine binding sites localized on α subunits from membrane extracts of both rat spinal cord and mammalian cells co-transfected with α and β cDNAs. Further, inhibition of α homo-oligomeric GlyRs by picrotoxinin, a non-competitive blocker of ion flow, was reduced 50- to 200-fold for α/β hetero-oligomeric receptors generated by cotransfection. Site-directed mutagenesis identified residues within the second predicted transmembrane segment (M2) of the β subunit as major determinants of picrotoxinin resistance. These data implicate the M2 segment in blocker binding to and lining of the GlyR chloride channel.