NMR structure and mutagenesis of the third Bir domain of the inhibitor of apoptosis protein XIAP

The inhibitor of apoptosis proteins (IAPs) regulate the caspase family of c ysteine proteases, which play an important role in the execution of program med cell death. Human X-linked inhibitor of apoptosis protein (XIAP) is a p otent inhibitor of caspases-3, -7, and -9, Here we show that the Bir3 domai n is the minimal region of XIAP that is needed for potent caspase-9 inhibit ion. The three-dimensional structure of the Bir3 domain of XIAP, determined by NMR spectroscopy, resembles a classical zinc finger and consists of fiv e cu-helices, a three-stranded beta -sheet, and a zinc atom chelated to thr ee cysteines and one histidine, The structure of the Bir3 domain is similar to that of the Bir2 domain of XIAP but differs from the previously determi ned structure of the Bir3 domain of MIHB. Based on site-directed mutagenesi s, tee have identified the regions of the Bir3 domain of XIAP that are impo rtant for inhibiting caspase-9. Despite the structural similarities of the Bir2 and Bir3 domain of XIAP, a different set of residues were found to be critical for inhibiting the individual caspases, These results suggest that XIAP inhibits caspase-3 and caspase-9 in a different manner.