Copper-Catalyzed Alkyne-Azide Cycloaddition and Thiol-Oxanobornadiene Reactions for Bioconjugation

The copper (I)-catalyzed azide-alkyne cycloaddition (CuAAC) reaction has found broad application in myriad fields. Sodium ascorbate is the reductant of choice for CuAAC reactions in organic and materials synthesis, but is avoided in bioconjugation with a few exceptions. Copper and sodium ascorbate have been shown to be detrimental to biological and synthetic polymers due to copper-mediated generation of reactive oxygen species. Moreover, dehydroascorbate and other ascorbate byproducts can react with lysine amine and arginine guanidine groups, leading to covalent modification and potential aggregation of proteins. We have looked into solutions to these problems and provided protocols and guidelines to perform demanding reactions with biomolecules of all types. This has allowed rapid synthesis of small molecules, DNA, RNA, and protein conjugates for numerous studies in our laboratory. In addition, the hunt for fluorogenic CuAAC reactions resulted in the discoveries of a new class of fluorogenic probe for thiols, an Hg (II) probe based on Rhodamine B, and a potential probe for CuI.