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《第十届全国化学生物学学术会议报告摘要集(女性科学家论坛)》2017年
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Fluorescent Probes for Single-Step Detection and Proteomic Profiling of HDACs

XIE Yusheng  SUN Hongyan  
【摘要】:HDACs play important roles in regulating various physiological and pathological processes.Developing fluorescent probes capable of detecting HDAC activity can help further elucidate the roles of HDACs in biology. In this study, we first developed a set of activity-based fluorescent probes by incorporating Kac residue and O-NBD group. Upon enzymatic removal of the acetyl group in Kac residue, the released free amine reacted intramolecularly with the O-NBD moiety, resulting in turn-on of fluorescence. These designed probes are capable of detecting HDAC activity in a continuous fashion, thereby eliminating the extra step of fluorescence development. Remarkably, the amount of turn-on fluorescence can be as high as50 fold, which is superior to the existing one-step HDAC fluorescent probes. Inhibition experiments further proved that the probes can serve as useful tools for screening HDAC inhibitors. Building on these results, we moved on and designed a dual-purpose fluorescent probe by introducing a diazirine photo-crosslinker into the probe. The resulting probe was not only capable of reporting enzymatic activity, but also able to directly identify and capture the protein targets from complex cellular environment. By combining fluorometric method and in-gel fluorescence scanning technique, we found that epigenetic readers and erasers can be readily identified and differentiated using a single probe. This is not achievable with traditional photoaffinity probes. In light of the prominent properties and the diverse functions of this newly developed probe, we envision that it can provide a robust tool for functional analysis of HDACs and facilitate future drug discovery in epigenetics.

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