Abstract
Deubiquitinating enzymes (DUBs) function to remove or cleave ubiquitin from post-translationally modified protein substrates. There are about 100 known DUBs in the proteome, and their dysregulation has been implicated a number of disease states, but the specific function of many subclass members remains poorly understood. Activity-based probes (ABPs) react covalently with an active site residue to report on specific enzyme activity, and thus represent a powerful method to evaluate cellular and physiological enzyme function and dynamics. Ubiquitin-based ABPs, such as HA-Ub-VME, an epitope-tagged ubiquitin carrying a C-terminal reactive warhead, are the leading tool for “DUBome” activity profiling. However, these probes are generally cell membrane impermeable, limiting their use to isolated enzymes or lysates. Development of cell-permeable ABPs would allow engagement of DUB enzymes directly within the context of an intact live cell or organism, refining our understanding of physiological and pathological function, and greatly enhancing opportunities for translational research, including target engagement, imaging and biomarker discovery. This mini-review discusses recent developments in small molecule activity-based probes that target DUBs in live cells, and the unique applications of cell-permeable DUB activity-based probes vs. their traditional ubiquitin-based counterparts.
Highlights
The Ubiquitin-Proteasome SystemThe ubiquitin-proteasome system (UPS) has attracted more excitement, scope and promise as a therapeutic target than any system since the rise of the kinome as a druggable protein family
Comprehensive reviews of Ub-based Activity-based probes (ABPs) have recently been published and the present review focuses on recent work toward cell permeable deubiquitinating enzymes (DUBs) ABPs (Fleury and Walker, 2015; Hewings et al, 2017; Leestemaker and Ovaa, 2017)
Ubiquitin based ABPs have significantly advanced our knowledge of DUB structure, dynamics and function
Summary
The ubiquitin-proteasome system (UPS) has attracted more excitement, scope and promise as a therapeutic target than any system since the rise of the kinome as a druggable protein family. In 2016, Ward et al published the first small-molecule based DUB ABP, based on a chloroacetylpyrrole scaffold (1), which was originally identified from a high-throughput screening (HTS) campaign at Mission Therapeutics (Figure 1F; Ward et al, 2016) This compound exhibited potent USP4 and USP11 biochemical activity, and so the authors employed a competitive activitybased protein profiling (ABPP) method to assess whether an alkyne-tagged analog 2 could be used as a live cell DUB ABP for quantitative target engagement. The authors demonstrate imaging of probe-labeled proteins in zebrafish embryos, strong off-target labeling of DJ-1 and ALDH may limit the utility of these probes in living systems
Sign-in/Register to view highlights & summary Sign-in/Register to access full text options 
Free) 
Talk to us
Join us for a 30 min session where you can share your feedback and ask us any queries you have
Schedule a call
