An unconventional IAP‐binding motif revealed by target‐assisted iterative screening (TAIS) of the BIR3‐cIAP1 domain

Target‐assisted iterative screening (TAIS) has been applied to a random phage‐displayed peptide library in a search for novel ligands of the third b aculovirus I AP (‘inhibitors of apoptosis’) r epeat (BIR) domain of cIAP1. The peptides selected in the screen fall into two distinct specificity groups, one that conforms to a known IAP‐binding motif (IBM) and another one that reveals a novel BIR domain interacting motif, NH 2 ‐SR(V/P)W. The biochemical profiling of selected sequences with synthetic peptides, which included alanine scanning and N‐ and C‐terminal truncations as well as competition with the Smac peptide, suggests a major energetic contribution of tryptophan at the +4 position of peptide ligands to binding and identifies the latter together with the respective pocket on the BIR domain surface as a ‘hot spot’ of the interaction. A peptide featuring the novel motif selectively binds the full‐length cIAP1 protein in cell lysates. A ‘two‐pocket’ model of BIR domain recognition mechanism is proposed as the basis of differential BIR domain interactions with different IBMs. Copyright © 2006 John Wiley & Sons, Ltd.