Daily Method Archive

Unless somehow the two domains could be joined back together. Now consider how this sets the stage for detecting protein-protein interactions. By creating two hybrids, one containing a binding domain fused to a protein of interest (binding domain + protein¹ = bait), the other with the activating domain fused to another protein (activating domain + protein² = prey).

The bait hybrid can bind to the DNA, but cannot activate transcription. The prey hybrid can activate transcription but has little chance to come in close proximity to the appropriate gene segment. However, if protein¹ and protein² interact with each other, then transcription can ensue. And we know when transcription ensues if the cell is manufacturing the protein that the reporter gene codes. The technique has been combined with a number of different reporter genes which can allow selection through a simple colour change or through automatic death of cells in which the interaction does or does not take place. For example, the lacZ reporter gene allows the highlighting of cells in which, β-galactosidase, the protein product of the lacZ gene produces a blue coloration through the metabolism of X-gal.