Bioorthogonal Ligation and Cleavage by Reactions of Chloroquinoxalines with ortho‐Dithiophenols

Abstract

A bioorthogonal ligation and cleavage method via reactions of chloroquinoxalines (CQ) and ortho‐dithiophenols (DT) is presented. Double nucleophilic substitutions of ortho‐dithiophenols to chloroquinoxalines provide conjugates containing tetracyclic benzo[5,6][1,4]dithiino[2,3‐b]quinoxaline with strong built‐in fluorescence together with release of the other functional molecules. Three cleavable linkers were designed and successfully used in release of the molecules containing biotin from the protein conjugates. The CQ‐DT bioorthogonal reactions can be applied for the bioorthogonal ligations, bioorthogonal cleavages, and trans‐tagging of proteins, and show advantages of readily accessible unnatural orthogonal groups, appealing reaction kinetics (k2≈1.3 m−1 s−1), excellent biocompatibility of orthogonal groups, and high stability of conjugates. This complements previous bioorthogonal reactions and is a new route for protein‐fishing applications and in‐gel fluorescence analysis.