A phage display system designed to detect and study protein–protein interactions

Abstract

SummaryAnalysing protein–protein interactions is critical in proteomics and drug discovery. The usage of 2‐Hybrid (2λ) systems is limited to an in vivo environment. We describe a bacteriophage 2‐Hybrid system for studying protein interactions in vitro. Bait and prey are displayed as fusions to the surface of phage λ that are marked with different selectable drug‐resistant markers. An interaction of phages in vitro through displayed proteins allows bacterial infection by two phages resulting in double drug‐resistant bacterial colonies at very low multiplicity of infections. We demonstrate interaction of the protein sorting signal Ubiquitin with the Vps9‐CUE, a Ubiquitin binding domain, and by the interaction of (Gly‐Glu)4 and (Gly‐Arg)4 peptides. Interruptions of the phage interactions by non‐fused (free) bait or prey molecules show how robust and unique our approach is. We also demonstrate the use of Ubiquitin and CUE display phages to find binding partners in a λ‐display library. The unique usefulness to 2λ is also described.