Characterization of Bcl-2, Bcl-XL, and Bax Pore Formation and Their Role in Apoptosis Regulation

摘要

The Bcl-2 protein family plays an important role in governing a cell's decision to live or die. Dysregulation of these proteins is observed in many breast cancer cases and thus it is important to understand their mechanism in order to develop new treatments. The Bcl-X(L) protein structure showed a strong similarity to pore-forming bacterial toxins, suggesting that Bcl-2 protein family protein may regulate apoptosis by pore formation or membrane insertion. Single cysteine mutants of Bcl-X(L) have been generated to probe the topology of the membrane-inserted state. Preliminary results show that the BH3 domain is present on the membrane surface but it is unclear as to whether this portion of the protein is actually membrane-inserted. Labeling of these cysteine mutants with thiol- specific probes such as BODIPY also yielded results suggesting that residues in both the putative pore-forming fifth a-helix and the BH3 domain were still solvent accessible, as indicated by the ability of a fluorescence- quenching anti-BODIPY antibody to quench fluorescence in the presence and absence of lipid vesicles on which the proteins display channel- forming capability.