Enzymatic “Click” Ligation: Selective Cysteine Modification in Polypeptides Enabled by Promiscuous Glutathione S‐Transferase

Abstract

Post-translational modifications of biomolecules represent nature's versatile chemical toolbox, allowing for precise and on-demand placement of various functional moieties in vivo.[1] Synthetic chemists have strived to develop a similar reaction toolkit with chemical orthogonality, selectivity and reactivity comparable to those of natural systems.[2] Recently, “click” chemistry has produced several powerful transformations applicable to a wide range of synthetic settings.[3] While the efficiency and chemical orthogonality of such “click” processes are remarkable, regio-control for these transformations has been limited and remains a major challenge.[4] The difficulty of selectively differentiating between two chemically identical sites within a macromolecule is routinely surmounted in natural systems by recognition elements such as binding pockets, directing groups and allosteric components. Although various enzymes have been engineered to catalyze reactions for protein modification and engineering,[5] they often require a long recognition sequence in polypeptides, and their substrate scope is relatively restricted. In certain cases enzymes are capable of recognizing and carrying out efficient transformations on more than one specific substrate, a phenomenon known as enzyme promiscuity. Promiscuous enzymes have been previously engineered to catalyze several synthetically important organic transformations.[6]