Conjugation of Synthetic N-Acetyl-Lactosamine to Azide-Containing Proteins Using the Staudinger Ligation

Abstract

Generation of defined glycoconjugates is necessary for the study of glycoprotein function, as well as the development of therapeutics. The biosynthesis of glycoproteins produces multiple glycoforms, proteins which differ only in the structure of the attached glycan. This inherent heterogeneity complicates the study of isolated glycans and, in particular, could obscure the role of individual glycan epitopes in biological function. We present a general strategy based on the Staudinger ligation to introduce specific glycan epitopes onto azide-containing proteins. The use of a phosphane-based Staudinger reagent allows for extremely mild reaction conditions which can be applied to aqueous proteins or cells. We demonstrate that multiple carbohydrate epitopes can be incorporated onto a protein backbone, and that the resulting glycans are competent for recognition by lectins. We propose that this general strategy will allow for testing the role of specific glycan epitopes in cellular and biochemical assays and increasing the stability of protein conjugates.