Protein‐Protein Interactions: Identification, modulation and validation

Abstract

Protein interaction mapping is key to elucidate protein functions and decipher cellular pathways.We have implemented an optimized yeast two‐hybrid (Y2H) protein interaction screening platform that allows to test on average 100 million interactions per screen. This corresponds to a 10‐fold coverage of any of our highly complex fragment libraries, and ensures exhaustive and reproducible results. Moreover, multiple independent fragments are isolated for each interactant, enabling the immediate delineation of a minimal interacting domain and the computation of a confidence score. Individual amino acids critical for the interaction can be identified using other yeast‐based tools, paving the way for functional studies and drug discovery projects where protein‐protein interactions can be used as drug targets.We have also developed additional methods to probe and characterize protein interactions in vitro with a medium to high throughput, using Surface Plasmon Resonance and a miniaturized time‐resolved FRET assay.Finally, taking advantage of our unique protein fragment libraries and screening technology in yeast, we are currently developing new tools to identify protein partners of DNA, RNA and small molecules.Here, we will present results that illustrate the strength and benefits of our protein interaction screening and validation techniques.