Discovery of Specific Nonpeptide Probe for Chymotrypsin via Molecular Docking-Based Virtual Screening and the Application.

Chymotrypsin is a proteolytic enzyme associated with numerous biological processes. Moreover, it has been reported to be significantly involved in pancreatic diseases. Thus, its rapid and sensitive detection is of great significance for early diagnosis and related drug discovery. Herein, a new strategy of molecular docking-based virtual screening (MDVS) was developed for the identification of a new nonpeptide-based biosensor targeting chymotrypsin. The newly discovered probe, numbered probe 20, exhibits about 45-fold specificity toward chymotrypsin over the similar enzyme trypsin and produces about 250-fold higher increase of fluorescence intensity under the catalysis of chymotrypsin. Furthermore, the probe successfully allowed the characterization of the kinetics of chymotrypsin inhibitors. More importantly, the endogenous chymotrypsin in zebrafish was visualized by nonpeptide probe for the first time, demonstrating the potential of the probe 20 for future application in the mechanistic study or clinic diagnosis for pancreatic diseases. Accordingly, the MDVS strategy could be generally applied in the identification of specific fluorescent probe for a particular enzyme.