Purification and characterization of the α1/β1 receptor from mammalian cells.

(A) Preparative size exclusion chromatography of IMAC-purified material isolates homogeneous receptor as suggested by FSEC analysis of four representative elution fractions collected across the SEC peak (B). (C) SDS-PAGE analysis of SEC fractions across the elution peak shown in (A) indicates diffuse subunit bands, consistent with glycosylation. (D) Purified receptor shows specific binding in C₁₂M or L-MNG detergent. Ligand binding experiments by SPA reveal high affinity binding sites for ³H-muscimol (K_D = 24 nM, B_max = 1690 CPM or 0.76 pmol ligand, E) or GABA (K_i = 98 nM, F) as measured by a direct binding isotherm or competition assay, respectively.