A recent paper in Nature Communications describes a collaborative effort between scientists at UbiQ, Genentech, Boston Biochem (Bio-Techne) and Stanford University. Here a reactive-site-centric chemoproteomics protocol is presented which allows evaluating activity and probe reactivity of deubiquitinating enzymes (DUBs). The chemoproteomics protocol makes use of a new type of bio-orthogonally tagged activity-based probe (ABP) and sequential on-bead digestions to enhance the identification of probe-labeling sites on DUBs (Figure 1). The bio-orthogonally tagged DUB activity-based probes are functionalized with a C-terminal vinyl pentynyl sulfone (VPS) electrophile (Figure 1). By using click chemistry, the VPS building block allows post-labeling functionalization of probe bound DUBs with azide modified molecules (such as azide-biotin). Ultimately, ZUFSP (ZUP1) was identified as a previously unannotated DUB with high selectivity toward cleaving K63-linked ubiquitin chains.*

Figure 1.

UbiQ now offers the following VPS activity-based probes:

• UbiQ-193: HA-Ahx-Ahx-Ub-VPS
• UbiQ-237: His6-Ahx-Ahx-SUMO2-VPS 
• UbiQ-262: Ac-ISG15prox-VPS (mouse ISG15 C-terminal domain)
• UbiQ-311: Ac-hISG15prox-VPS (human ISG15 C-terminal domain)

1. Discovery Oncology, Genentech, South San Francisco, California 94080, USA
2. Early Discovery Biochemistry, Genentech, South San Francisco, California 94080, USA
3. Discovery Chemistry, Genentech, South San Francisco, California 94080, USA
4. Department of Pathology, Stanford University School of Medicine, Stanford, California 94305, USA
5. Microchemistry, Proteomics and Lipidomics, Genentech, South San Francisco, CA 94080, USA.
6. UbiQ Bio BV, Science Park 408, 1098 XH Amsterdam, The Netherlands.
7. Boston Biochem Inc., 840 Memorial Drive, Cambridge, Massachussetts 02139, USA.
8. Structural Biology, Genentech, South San Francisco, California 94080, USA.
9. Bioinformatics, Genentech, South San Francisco, California 94080, USA. 10
10. Present address: Merck, 630 Gateway Boulevard, South San Francisco, California 94080, USA.

These authors contributed equally: Taylur P. Ma and Andrew P. AhYoung.

* three other groups also identified ZUFSP (ZUP1) at the same: (1) Kwasna et al. Mol Cell 2018, 70, 150. (2) Haahr et al. Mol Cell 2018, 70, 165. (3) Hermanns et al. Nat Comm 2018, 9, 799.