A cell-permeable inhilbitor and activity-based probe for the caspase-like activity of the proteasome

The ubiquitin-proteasome pathway degrades the majority of proteins in mammalian cells and plays an essential role in the generation of antigenic peptides presented by major histocompatibility class I molecules. Proteasome inhibitors are of great interest as research tools and drug candidates. Most work on proteasome inhibitors has focused on the inhibition of the chymotryptic-like (beta 5) sites; little attention has been paid to the inhibition of two other types of active sites, the trypsin-like (beta 2) and the caspase-like (beta 1). We report here the development of the first cell-permeable and highly selective inhibitors (4 and 5) of the proteasome's caspase-like site. The selectivity of the compounds is directly and unambiguously established by Staudinger-Bertozzi labeling of proteasome subunits covalently modified with azide-functionalized inhibitor 5. This labeling reveals that the caspase-like site of the immunoproteasome (beta 1i) is a preferred target of this compound. These compounds can be used as tools to study roles of beta 1 and beta 1i sites in generation of specific antigenic peptides and their potential role as co-targets of anti-cancer drugs. (c) 2007 Elsevier Ltd. All rights reserved.