Determinants of BH3 Sequence Specificity for the Disruption of Bcl-xL/cBid Complexes in Membranes

The prosurvival Bcl-2 proteins exhibit a specific pattern of interactions with BH3-only proteins that determines the cellular dependence on apoptotic stress. This specificity is crucial for the development of BH3 mimetics, a class of anticancer molecules based on the BH3 domain with promising activity in clinical trials. Although complex formation mainly takes place in the mitochondrial outer membrane, most studies so far addressed the interaction between BH3 peptides and truncated Bcl-2 proteins in solution. As a consequence, quantitative understanding of the sequence specificity determinants of BH3 peptides in the membrane environment is missing. Here, we tackle this issue by systematically quantifying the ability of BH3 peptides to compete for the complexes between cBid and Bcl-xL in giant unilamellar vesicles and compare it with solution and mitochondria. We show that the BH3 peptides derived from Hrk, Bim, Bid, and Bad are the most efficient in disrupting cBid/Bcl-xL complexes in the membrane, which correlates with their activity in mitochondria. Our findings support the targeting to the membrane of small molecules that bind Bcl-2 proteins as a strategy to improve their efficiency