Fig. 1.

Designing a deregulated Bax mutant. (A) Sequence alignments illustrate key conserved residues. The BH3 domains from selected proapoptotic Bcl-2 family members (Left) show the invariant aspartate (red D) and the conserved hydrophobic residues in bold, whereas the sequences from the hydrophobic receptor grooves of the mammalian prosurvival relatives (Right) show the invariant arginine (R) in blue. (B) The D68R mutation compromises Bax binding to prosurvival proteins. Lysates prepared from 293T cells overexpressing HA-tagged human Bax or the D68R mutant, and FLAG-tagged prosurvival Bcl-2 proteins were immunoprecipitated with mouse monoclonal antibodies recognizing the HA, FLAG (FL), or an irrelevant control (C) tag. The immunoprecipitates were subjected to SDS-PAGE, transferred onto membranes, and the blots probed with rat anti-HA or -FLAG antibodies.