Article ID | Journal | Published Year | Pages | File Type |
---|---|---|---|---|
8296960 | Biochemical and Biophysical Research Communications | 2014 | 6 Pages |
Abstract
The deubiquitinating enzymes (DUBs) are a family of isopeptidases responsible for removing the ubiquitin from the ubiquitinated proteins. Identification of inhibitors for DUBs is emerging as an efficient way for discovering potential medicines for disease treatment. However, the high throughput screening (HTS) assay is still not available for all USPs, especially OTULIN. Here, we described a novel steadily quantifiable DUBs assay platform using Nanoluc (Nluc) as reporter. We further demonstrated that the Ub-Nluc assay could be used for HTS of DUBs inhibitors. Moreover, we generated a sensitive system for OTULIN inhibitors screening using Nluc as a reporter. In summary, our data indicate that Ub-Nluc and the improved Ub-Ub-GS-Nluc assay are efficient systems for measuring activities and screening inhibitors of USPs and OTULIN.
Keywords
OTULINHTSNanoLucRFUUSPRLUeGFPDTTAmCGSTCBB7-amino-4-methylcoumarinBSACoomassie Brilliant BlueDMSObovine serum albuminDeubiquitinating enzymesFluorescence resonance energy transferFRETstandard deviationdithiothreitolDimethylsulfoxidehigh-throughput screeningrelative fluorescence unitubiquitin-specific proteaseenhanced green fluorescence proteinglutathione S-transferaseUbiquitin
Related Topics
Life Sciences
Biochemistry, Genetics and Molecular Biology
Biochemistry
Authors
Yunfei Chen, Lufan Wang, Xiaomu Cheng, Xin Ge, Ping Wang,