Enzyme-catalyzed expressed protein ligation.

Abstract

Expressed protein ligation is a valuable method for protein semisynthesis that involves the reaction of recombinant protein C-terminal thioesters with N-Cys containing peptides but the requirement of a Cys residue at the ligation junction can limit its use. Here we employ subtiligase variants to efficiently ligate Cys-free peptides to protein thioesters. Using this method, we have more accurately determined the effect of C-terminal phosphorylation on the tumor suppressor protein PTEN.