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Using BRET to study chemical compound‐induced disruptions of the p53‐HDM2 interactions in live cells
Author(s) -
Mazars Anne,
Fåhraeus Robin
Publication year - 2010
Publication title -
biotechnology journal
Language(s) - English
Resource type - Journals
SCImago Journal Rank - 1.144
H-Index - 84
eISSN - 1860-7314
pISSN - 1860-6768
DOI - 10.1002/biot.200900272
Subject(s) - allosteric regulation , protein–protein interaction , ubiquitin ligase , chemistry , ubiquitin , in vitro , microbiology and biotechnology , computational biology , rational design , biochemistry , biophysics , biology , receptor , genetics , gene
Modification of protein‐protein interactions (PPIs) holds promise for novel rational drug design. Disrupting or modifying protein interactions offers new challenges in terms of chemical compound libraries and techniques for compound validation. As proteins interact with several partners in different allosteric conformation in a pathological and tissue‐specific fashion, 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 tumor 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 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 PPIs in living cells.