Irreversible, or mechanism-based, inhibitors are attractive tools in chemical biology. Attachment of a reporter entity, which can be a biotin, a fluorophore, a bioorthogonal tag, a radiolabel or a combination of these, allows for the identification, visualization, identification and spatiotemporal study of the modified enzyme(s) using various techniques and in complex biological systems ranging from cell extracts, living tissue to animal models. This is the field of activity-based protein profiling (ABPP). Within the group we develop activity-based probes (ABPs) for proteases, glycosidases and a range of other hydrolases and apply these in immunological and glycobiological studies related to health and disease. Together with these ABPs we develop focused libraries of (competitive) inhibitors. Together, this toolset allows us to perform a number of conceptual experiments. In comparative ABPP ABPs are used to discover new enzyme activities and to compare enzyme expression levels in different tissue. In competitive ABPP the focused inhibitor libraries are included to study efficacy and selectivity of these in complex biological samples. In fluorescence polarization ABPP we develop assays amenable for high-throughput screening to discover new chemical entities for inhibiting one of our target enzymes.
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