. Department of Chemistry, City University of Hong Kong, Hong Kong, China.. Key Laboratory of Biochip Technology, Biotech and Health Centre, City University of Hong Kong Shenzhen Research Institute, Shenzhen 518057, China.. School of Biomedical Sciences, University of Hong Kong, Hong Kong, China.. Department of Biomedical Science, College of Veterinary Medicine and Life Sciences, City University of Hong Kong, Hong Kong, China
Lysine lipoylation plays vital roles in cell metabolism and redox processes. For example, removal of lipoylation will decrease pyruvate dehydrogenase activity and affect the citric acid cycle. Despite the important functions of lysine lipoylation, the mechanisms for the addition and removal of this modification remain largely unexplored. Very few useful chemical tools are available to study the interactions of lysine lipoylation with its regulatory delipoylation proteins. For example, immunoaffinity purification-mass spectrometry is one of such tools, which highly relies on antibody efficiency and purification techniques. Single-step activity based fluorogenic probes developed by our groups and others is also an efficient method to study the deacylation activity. Affinity-based labeling probe using photo-cross-linker is a powerful platform to study the transient and dynamic interactions of peptide ligands with the interacting proteins. Herein, we have designed and synthesized a dual-function probe KTLlip for studying enzymatic delipoylation (eraser) activity and interaction of lysine lipoylation with the eraser at the same time. We show that KTLlip can be used as a useful tool to detect delipoylation as demonstrated by its ability to fluorescently label the eraser activity of recombinant Sirt2. We envision that the probe will help delineate the roles of delipoylation enzyme in biology.