1. Department of Molecular Biology, Princeton University, Lewis Thomas Laboratory, Washington Road , Princeton, New Jersey 08544, USA
2. 310 Aycock Dorm, University of North Carolina at Chapel Hill, Chapel Hill, North Carolina 27514, USA
3. Department of Biochemistry and Howard Hughes Medical Institute, University of Texas Southwestern Medical Center , Dallas, Texas 75235, USA
4. These authors contributed equally to this work

Correspondence to: Yigong Shi¹ Correspondence and requests for materials should be addressed to Y.S. (e-mail: Email: yshi@molbio.princeton.edu). Atomic coordinates for the Smac/BIR3 complex have been deposited at the Protein Data Bank with accession number 1G73.

Abstract

Apoptosis is an essential process in the development and homeostasis of all metazoans^{1, 2, 3, 4}. The inhibitor-of-apoptosis (IAP) proteins suppress cell death by inhibiting the activity of caspases; this inhibition is performed by the zinc-binding BIR domains^{5, 6} of the IAP proteins. The mitochondrial protein Smac/DIABLO promotes apoptosis by eliminating the inhibitory effect of IAPs through physical interactions^{7, 8, 9}. Amino-terminal sequences in Smac/DIABLO are required for this function, as mutation of the very first amino acid leads to loss of interaction with IAPs and concomitant loss of Smac/DIABLO function⁹. Here we report the high-resolution crystal structure of Smac/DIABLO complexed with the third BIR domain (BIR3) of XIAP. Our results show that the N-terminal four residues (Ala-Val-Pro-Ile) in Smac/DIABLO recognize a surface groove on BIR3, with the first residue Ala binding a hydrophobic pocket and making five hydrogen bonds to neighbouring residues on BIR3. These observations provide a structural explanation for the roles of the Smac N terminus as well as the conserved N-terminal sequences in the Drosophila proteins Hid/Grim/Reaper. In conjunction with other observations, our results reveal how Smac may relieve IAP inhibition of caspase-9 activity. In addition to explaining a number of biological observations, our structural analysis identifies potential targets for drug screening.