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Application: Enzyme kinetics studies and screening small molecule inhibitors for drug discovery and high-throughput screening (HTS) applications.
Background: Ubiquitin specific peptidase 15 (USP15), belongs to a large group of ubiquitin-specific proteases capable of cleaving ubiquitin from other proteins. These enzymes are also referred to as deubiquitinating peptidases, deubiquitinases (DUBs), ubiquitin proteases, ubiquitin hydrolases or ubiquitin isopeptidases. They contribute to the ubiquitin signaling pathway by countering the signal induced by ubiquitin conjugating enzymes and ligases. In humans, USP15 is found mainly in endocrine tissues, liver, bone marrow, lymphoid tissues, amongst others. It is involved in cell proliferation, apoptosis, autophagy, immune response, etc. It can be found at high levels in certain types of cancer, such as MM (multiple myeloma) and prostate cancer, but it is also absent in a percentage of pancreatic cancers and glioblastoma. Thus, USP15 may have a tumor suppressive or a tumorigenic role dependent on the cancer type. Studies with the small molecule inhibitor USP15-Inh resulted in deleterious effects on leukemia cells, indicating that the use of USP15 inhibitors may open new avenues of treatment for these diseases.
Description: The USP15 Inhibitor Screening Assay Kit is a 96-well format fluorogenic assay kit designed to measure the activity of the deubiquitinating (DUB) enzyme USP15 (ubiquitin specific peptidase 15) for screening and profiling applications. The kit contains enough purified USP15, Ubiquitinated-AMC substrate, and assay buffer for 100 reactions. To determine the effect of an inhibitor on USP15 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 the DUB subfamilies Ubiquitin C-terminal Hydrolases (UCHs), Ubiquitin-Specific Proteases (USPs), Ovarian Tumor Proteases (OTU), and Machado-Josephin Domain (MJD) proteases.Ubiquitin-AMC is a fluorogenic substrate for ubiquitin hydrolases, with a C-terminus derivatization of ubiquitin with 7-amido-4-methylcoumarin (AMC). In the conjugated form, the energy emitted from fluorochrome AMC is quenched. Upon proteolysis, AMC is no longer quenched and emits fluorescence with an λexcitation/λemission 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 USP15? Check out our Protease Screening Services or Ubiquitination Screening Services.
Storage Stability: This assay kit will perform optimally for up to 6 months from date of receipt when the materials are stored as directed.
Target: USP15
Uniprot: Q9Y4E8
Warnings: Avoid freeze/thaw cycles.
Biosafety Level: Not applicable (BSL-1)
References: An T., et al., 2022 Front Pharmacol. 13:944989.
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