Activity-based protein profiling (ABPP) is a proteomics tool used to measure functionally active proteins from a complex biological sample, typically by irreversible inhibition of enzyme active sites. Coupled to quantitative LC-MS based technology, probe-protein reactivity, measured as protein abundance, can be compared between proteome samples.  We are interested in combining LC-MS ABPP technology with organelle targeting probes in order to elucidate spatial factors involved in protein reactivity of disease states, such as infection of host by microbial pathogens.  Herein, we present preliminary data on the design and synthesis of a copper-catalyzed click chemistry compatible suite of lysosome targeting probes.  Using a convergent synthesis, a lysosomal targeting moiety was conjugated to known cathepsin targeting active site and general cysteine reactive irreversible inhibitors.  Appending a fluorophore reporter or a biotin affinity enrichment moiety, via click chemistry allows down-stream analysis of lysosomal protein reactivity by SDS-PAGE, in situ cell imaging, and quantitative LC-MS.

Lysosomal directing activity-based probes for the analysis of acid hydrolases by cell imaging and mass spectrometry

245th National Meeting (2013)

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