A Yeast Three-Hybrid Method to Clone Ternary Protein Complex Components

Abstract

The yeast two-hybrid system is a powerful technique to detect binary protein interactions. In order to facilitate ternary complex analysis, we have developed the yeast three-hybrid system. Using epidermal growth factor (EGF) receptor, Grb2 and Sos proteins as an example, we demonstrated the use of the three-hybrid system to detect formation of a three-component complex, i.e., the Grb2-mediated interactions between EGF receptor and Sos. We then used the three-hybrid system to screen a library with Gal4–DNA binding domain –EGF receptor and Gal4 activation domain–Sos fusion proteins and isolated a plasmid encoding Grb2. Thus, the three-hybrid system is useful for characterizing and cloning components of ternary protein complexes.