LEC–BiFC: a new method for rapid assay of protein interaction

Abstract

Abstract Protein-protein interactions play fundamental roles in most biological processes. Bimolecular fluorescence complementation (BiFC) is a promising method for its simplicity and direct visualization of protein-protein interactions in cells. This method, however, is limited by background fluorescence that appears without specific interaction between the proteins. We report here a point mutation (V150L) in one Venus BiFC fragment that efficiently decreases background fluorescence of BiFC assay. Furthermore, by combining this modified BiFC and linear expression cassette (LEC), we develop a simple and rapid method (LEC-BiFC) for protein interaction analysis that is demonstrated by a case study of the interaction between Bcl-X(L) and Bak BH3 peptide. The total analysis procedure can be completed in two days for screening tens of mutants. LEC-BiFC can be applied easily in any lab equipped with a fluorescence microscope.