Protein:protein interactions (PPIs) are essential to almost every process in a cell, so understanding PPIs is crucial for understanding cell biology in normal and disease states. There are numerous methods used to characterize proteins interactions that utilize cell extracts or live cells to express potential targets.
In a pull-down assay, one protein partner is expressed as a fusion protein (e.g., bait protein) in E. coli and then immobilized using an affinity ligand specific for the fusion tag. The immobilized bait protein can then be incubated with the prey protein.
Coimmunoprecipitation is another option to study protein:protein interactions. To perform coimmunoprecipitation, an antibody against a target protein is coupled to Sepharose beads through protein A or G, then the complexes containing the target protein are immunoprecipitated with the antibody-coupled beads.
However, these traditional methods for studying interactions between proteins do not provide data directly in the context of a cellular environment. Using Bioluminescence Resonance Energy Transfer (BRET) or complementation assays, it is possible to quantitatively measure the interaction between proteins in live cells.