A phage display system designed to detect and study protein–protein interactions |
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Authors: | Catherine L Bair Amos Oppenheim rei Trostel Gali Prag Sankar Adhya |
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Institution: | Laboratory of Molecular Biology, Center for Cancer Research, National Cancer Institute, National Institutes of Health, Bethesda, MD 20892, USA.; Department of Molecular Genetics and Biotechnology, Hebrew University-Hadassah Medical School, Jerusalem 91120, Israel.; Department of Biochemistry, George S. Wise Faculty of Life Sciences Tel Aviv University Tel Aviv, Israel. |
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Abstract: | Analysing 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. |
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