Comparison of the Smac based peptides and Smac protein in antagonizing caspase‐3 and ‐7 inhibition by the BIR12 and BIR123 domains of XIAP

XIAP prevents apoptosis through inhibition of caspase‐9 by the BIR‐3 and caspases ‐3 and ‐7 through the BIR2 domain. SMAC which is released from the mitochondria competes with caspases in binding to XIAP unleashing caspase activity and causing cell death. SMAC peptides and protein were used to investigate their ability in relieving the executioner caspase activities inhibited by both the BIR12 domains and the BIR123 domains of XIAP. Furthermore, the potency of these peptides was compared to the Smac protein in antagonizing XIAP. AKPD, ANPR, SGVD, AVPI peptides and the SMAC protein were preincubated with the XIAP domains and the activity of caspases was studied in the presence of these mixtures. Moreover, the ability of these peptides in preventing the interaction of BIR12 domain with the large and the small subunits of executioner caspases was studied. It was observed that the Smac protein by far is the most potent agent in reversing caspase inhibition. In addition, Caspase‐3 inhibition by XIAP domains was more sensitive to SMAC peptides than that of caspase‐7. These results indicate that under conditions of extensive XIAP cleavage and involvement of caspase‐7 as the driving force for execution of apoptosis, Smac, and by extension Smac based anticancer agents, cannot be effective in inducing cell death.