Coimmunoprecipitation of GNB-1 with GNG-1. (A) Levels of GNB-1, FLAG-GNG-1, and GNG-1 proteins in plasma membrane fractions. Plasma membrane fractions were prepared from 16-h submerged cultures of gng-1+ (74A), Δgng-1+ FLAG-GNG-1 (5A), and Δgng-1 his-3 (113) strains. Only strain 5A expresses the FLAG-GNG-1 fusion protein (see Materials and Methods). Samples containing 50 μg of total protein were resolved on 10% (GNB-1) or 15% (GNG-1 and FLAG-GNG-1) SDS-PAGE gels. GNB-1, GNG-1, and FLAG antisera were used for Western analysis (see Materials and Methods). Nonspecific bands are indicated by asterisks. (B) Immunoblot analysis after coimmunoprecipitation. The FLAG-GNG-1 protein in extracts from the indicated strains in panel A was immunoprecipitated using anti-FLAG M2-agarose (see Materials and Methods), and the precipitated proteins were examined by immunoblot analysis using anti-FLAG, anti-GNG-1, or anti-GNB-1 antibodies.