USP21 Inhibitor Screening Assay Kit

The USP21 Inhibitor Screening Assay Kit is a 96-well format fluorogenic assay designed to measure the activity of the deubiquitinating (DUB) enzyme USP21 (ubiquitin specific peptidase 21) for screening and profiling applications. The kit contains enough purified USP21 protein, Ubiquitinated-AMC substrate, and assay buffer for 100 reactions. To determine the effect of an inhibitor on USP21 activity, the enzyme should be preincubated with or without the test inhibitor prior to adding the Ub-AMC substrate to the reaction. The assay was functionally validated using Ub-Aldehyde, a potent inhibitor of DUB subfamilies ubiquitin C-terminal hydrolases (UCHs), ubiquitin-specific proteases (USPs), Ovarian Tumor Proteases (OTU), and Machado-Josephin domain proteases (MJD). Figure 1: Illustration of the assay principle. Ubiquitin-AMC is a fluorogenic substrate for ubiquitin hydrolases based on the C-terminus derivatization of ubiquitin with 7-amido-4-methylcoumarin (AMC). In conjugated form, the energy emitted from fluorochrome AMC is quenched. Upon proteolysis, AMC is no longer quenched and emits fluorescence with excitation/emission maxima of 350/460 nm. The increase in fluorescence is proportional to the DUB activity. Need us to run inhibitor screens or profile your compounds against USP21? Check out our Protease Screening Services or Ubiquitination Screening Services.