Optimization of the Azobenzene Scaffold for Reductive Cleavage by Dithionite; Development of an Azobenzene Cleavable Linker for Proteomic Applications

In this paper we conducted an extensive reactivity study to determine the key structural features that favour the dithionite-triggered reductive cleavage of the azo-arene group Our stepwise investigation allowed identification of a highly reactive azo arene structure 25 bearing a carboxylic acid at the ortho position of the electron-poor arene and an ortho-O-alkyl-resorcinol as the electron-rich arene. Based on this 2-(2'-alkoxy-4'-hydroxyphenylazo)benzoic acid (HAZA) scaffold, the orthogonally protected difunctional azo-arene cleavable linker 26 was designed and synthesized Selective linker deprotection and derivatization was performed by introducing an alkyne reactive group and a biotin affinity tag. This optimized azo-arene cleavable linker led to a total cleavage in less than 10 s with only 1 mm dithionite Similar results were obtained in biological media