USP7 is a deubiquitinating enzyme that plays a pivotal role in multiple oncogenic pathways and therefore is a desirable target for new anti-cancer therapies. However, the lack of structural information about the USP7-inhibitor interactions has been a critical gap in the development of potent inhibitors. USP7 is unique among USPs in that its active site is catalytically incompetent, and is postulated to rearrange into a productive conformation only upon binding to ubiquitin. Surprisingly, we found that ubiquitin alone does not induce an active conformation in solution. Using a combination of nuclear magnetic resonance, mass spectrometry, computational modeling, and cell-based assays, we found that DUB inhibitors P22077 and P50429 covalently modify the catalytic cysteine of USP7 and induce a conformational switch in the enzyme associated with active site rearrangement. This work represents the first experimental insights into USP7 activation and inhibition and provides a structural basis for rational development of potent anti-cancer therapeutics. Pozhidaeva et al. combine biophysical and biological methods to reveal the mechanism of action of two selective inhibitors of deubiquitinating enzyme USP7, and show that the compounds covalently and irreversibly modify the catalytic cysteine of the enzyme through distinct chemical reactions.
Pozhidaeva, A., Valles, G., Wang, F., Wu, J., Sterner, D. E., Nguyen, P., … Bezsonova, I. (2017). USP7-Specific Inhibitors Target and Modify the Enzyme’s Active Site via Distinct Chemical Mechanisms. Cell Chemical Biology, 24(12), 1501-1512.e5. https://doi.org/10.1016/j.chembiol.2017.09.004