Solubilization and partial purification of the GABA receptor from mouse brain and a binding assay for the solubilized receptor.

Abstract

Abstract— The GABA receptor from mouse brain was solubilized with lysolecithin. A 56‐fold overall purification and activation were achieved by discontinuous sucrose gradient centrifugation and solubilization. Activation of binding by both procedures was observed. The solubilized receptor has the following binding constants: KD1= 3.5 nM, KD2= 52 nM, Bmax 1= 2.8 pmol/mg protein and Bmax 2= 14 pmol/mg protein for muscimol; KD1= 12 nM, KD2= 470 nM, Bmax 1= 1.4 pmol/mg protein and Bmax 2= 17 pmol/mg protein for GABA. Specific GABA binding was inhibited by imidazoleacetic acid and bicuculline with IC50 values of 250nM and 1 μM respectively. A rapid and sensitive filtration binding assay for the solubilized receptor has been developed. Lysolecithin was also found suitable for the solubilization of acetylcholine receptor from T. californica electroplaques.