Structures of BIR domains from human NAIP and cIAP2

Abstract

The inhibitor of apoptosis (IAP) family of proteins contains key modulators of apoptosis and inflammation that interact with caspases through baculovirus IAP-repeat (BIR) domains. Overexpression of IAP proteins frequently occurs in cancer cells, thus counteracting the activated apoptotic program. The IAP proteins have therefore emerged as promising targets for cancer therapy. In this work, X-ray crystallography was used to determine the first structures of BIR domains from human NAIP and cIAP2. Both structures harbour an N-terminal tetrapeptide in the conserved peptide-binding groove. The structures reveal that these two proteins bind the tetrapeptides in a similar mode as do other BIR domains. Detailed interactions are described for the P1'-P4' side chains of the peptide, providing a structural basis for peptide-specific recognition. An arginine side chain in the P3' position reveals favourable interactions with its hydrophobic moiety in the binding pocket, while hydrophobic residues in the P2' and P4' pockets make similar interactions to those seen in other BIR domain-peptide complexes. The structures also reveal how a serine in the P1' position is accommodated in the binding pockets of NAIP and cIAP2. In addition to shedding light on the specificity determinants of these two proteins, the structures should now also provide a framework for future structure-based work targeting these proteins.

Figure 1

(a) Schematic representation of the structure of the cIAP2 BIR3 domain. An experimentally achieved dimer constitutes the asymmetric unit. The N-termini bind in the peptide-binding pockets of the BIR domains, forming an extension of the central β-sheet. (b) Schematic representation of the NAIP BIR2-domain structure. The N-terminus, shown in yellow, binds in the peptide-binding pocket similarly as in the cIAP2 BIR3 structure.

Figure 2

Sequence alignment of BIR domains. The sequences of the described BIR3 and BIR2 domains from human cIAP2 and NAIP are compared with the corresponding BIR domains from XIAP. The ML-IAP BIR domain structure was used as search model for molecular replacement. Colour markers: red triangles, Zn²⁺-chelating residues; green triangles, residues of high importance for IBM interaction (see text for details); blue triangles, conserved residues lining a hydrophobic cavity at the P1′ binding position.

Figure 3

Representation of the IBM-binding site of the BIR domains. (a) Superposition of cIAP2 BIR3 (green/yellow) with NAIP BIR2 (blue/light blue). (b) Stereo diagram showing the IBM-binding pocket of cIAP2 BIR3 with the N-terminal peptide SMRY in yellow.

Figure 4

Comparison of interactions in the IBM-binding cleft. (a) Superposition of cIAP2 BIR3 (green/yellow) with XIAP BIR3 (brown/white). (b) Superposition of NAIP BIR2 (blue/light blue) with XIAP BIR2 (pink) bound to the N-terminus of the processed caspase-3 small domain (sand).