Using BRET to study chemical compound-induced disruptions of the p53-HDM2 interactions in live cells.
Résumé
Modification of protein-protein interactions (PPIs) holds promise for novel rational drug design. Disrupting or modifying protein interactions offer new challenges in terms of chemical compound libraries and techniques for compound validation. As proteins interact with several partners in different allosteric conformation in pathological and tissue specific fashions it is difficult to predict the in vivo effect of PPI acting compounds identified by in vitro screening assays. It is therefore desirable to develop techniques that rapidly allow cell-based validation of protein interacting compounds. The binding of the p53 tumour suppressor to the HDM2 E3 ubiquitin ligase is important for controlling p53 activity and several compounds, such as Nutlin-3, have been designed to bind a hydrophobic pocket in the N-terminus of HDM2 to prevent the interaction with p53 in order to stabilize and activate downstream p53 pathways. We have used the p53-HDM2 interaction as a model system to explore the Bioluminescence Resonance Energy Transfer (BRET) technique for validating compounds that disrupt protein-protein interactions in living cells.
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