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Updated: Jun 29, 2025
Method for Measuring the Activity of Deubiquitinating Enzymes in Cell Lines and Tissue Samples
Published on: May 10, 2015
A cell-permeable Ub-Dha probe for profiling E1-E2-E3 enzymes in live cells
Qiong Xia1, Xianbin Meng2, Yu Wang1
1School of Food and Biological Engineering, Engineering Research Center of Bio-process, Ministry of Education, Hefei University of Technology, Hefei 230009, China. ymli@hfut.edu.cn.
Abstract:
Activity-based ubiquitin probes (Ub-ABPs) have recently been developed as effective tools for studying the capabilities of E1-E2-E3 enzymes, but most of them can only be used in cell lysates. Here, we report the first cell-penetrating Ub-Dha probes based on thiazolidine-protected cysteines, which enable successful delivery into cells confirmed by a fluorophore at the N-terminus of Ub and live-cell fluorescence microscopy. A total of 18 E1-E2-E3 enzymes in live cells were labelled and enriched in combination with label-free quantification (LFQ) mass spectrometry. This work provided a new cell-penetrating Ub tool for studying the activity and function of Ub-related enzymes.
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