E1-catalyzed ubiquitin C-terminal amidation for the facile synthesis of deubiquitinase substrates

Abstract

Will Ub my partner? The ubiquitin (Ub)-activating enzyme (E1) was used to catalyze an amidation reaction to functionalize the C terminus of Ub with unique functional groups, such as thiol, azide, alkyne, and alkene groups, with high efficiency and yield (>90 %). These groups were then applied for the facile synthesis of fluorophore-conjugated ubiquitin and specifically conjugated diubiquitin substrates for deubiquitinases.

Keywords: deubiquitinases; diubiquitin; protein engineering; synthetic biology; ubiquitin functionalization.