Activity-based protein profiling: The serine hydrolases

With the postgenome era rapidly approaching, new strategies for the functio nal analysis of proteins are needed. To date, proteomics efforts have prima rily been confined to recording variations in efforts have primarily been c onfined to recording variations in protein level rather than activity. The ability to profile classes of proteins on the basis of changes in their act ivity would greatly accelerate both the assignment of protein function and the identification of potential pharmaceutical targets. Here, we describe t he chemical synthesis and utility of an active-site directed probe for visu alizing dynamics in the expression and function of an entire enzyme family, the serine hydrolases. By reacting this probe, a biotinylated fluorophosph onate referred to as FP-biotin, with crude tissue extracts, we quickly and with high sensitivity detect numerous serine hydrolases, many of which disp lay tissue-restricted patterns of expression. Additionally, we show that FP -biotin labels these proteins in an activity-dependent manner that can be f ollowed kinetically, offering a powerful means to monitor dynamics simultan eously in both protein function and expression.