Synthesis of N-Hydroxy Isopeptide Containing Proteins

Chemical synthesis of a peptide-ubiquitin conjugate linked by an N-hydroxy isopeptide bond to determine what effect the N-hydroxy group has on the enzymatic hydrolysis of the isopeptide linkage by deubiquitinases is reported. This conjugate was subjected to proteolysis by UCH-L3 in the presence and absence of various metal ions, and no substantive difference in hydrolysis was seen compared to a control lacking the N-hydroxy group. The accessibility of N-hydroxy ubiquitinated substrates may find uses to study other deubiquitinases in particular those which use a zinc ion as a part of their catalytic mechanism.