Enzyme-catalyzed expressed protein ligation.

Henager, Samuel H; Chu, Nam; Chen, Zan; Bolduc, David; Dempsey, Daniel R; Hwang, Yousang; Wells, James; Cole, Philip A · Nat Methods · 2016

basic_science · Level V

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Abstract

Expressed protein ligation is a valuable method for protein semisynthesis that involves the reaction of recombinant protein C-terminal thioesters with N-terminal cysteine (N-Cys)-containing peptides, but the requirement of a Cys residue at the ligation junction can limit the utility of this method. 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.

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