A strategy for the synthesis of dehydroalanine based diubiquitin activity probes is described.
The site-specific introduction of dehydroalanine was achieved from diubiquitin bearing Cys
residue near the scissile bond between two ubiquitins linked through Lys48, Lys63 or in a
head to tail fashion. The probes were characterized for their activities with various
deubiquitinases, which open new opportunities in studying deubiquitinases in various
settings.