Efficient semi-synthesis of ubiquitin-7-amino-4-methylcoumarin

Ubiquitin (Ub)-based probe Ub-7-amino-4-methylcoumarin is widely used to detect deubiquitinase (DUB) enzymatic activity, but its current synthesis procedures are cumbersome. In this work, we reported a new highly efficient semi-synthetic strategy to prepare Ub-AMC through two facile steps: first, preparation of Ub-thioester through hydrozinolysis-thioesterification of expressed Ub (1-G76C) mutant; second, direct aminolysis of Ub-thioester in a mixed-solvent system which exhibits very low hydrolysis.