Selective 13C‐Labels on Repeating Glycan Oligomers to Reveal Protein Binding Epitopes through NMR: Polylactosamine Binding to Galectins

A combined chemo‐enzymatic synthesis/NMR‐based methodology is presented to identify, in unambiguous manner, the distinctive binding epitope within repeating sugar oligomers when binding to protein receptors. The concept is based on the incorporation of 13C‐labels at specific monosaccharide units, selected within a repeating glycan oligomeric structure. No new chemical tags are added, and thus the chemical entity remains the same, while the presence of the 13C‐labeled monosaccharide breaks the NMR chemical shift degeneracy that occurs in the non‐labeled compound and allows the unique identification of the different components of the oligomer. The approach is demonstrated by a proof‐of‐concept study dealing with the interaction of a polylactosamine hexasaccharide with five different galectins that display distinct preferences for these entities. Repeating carbohydrate patterns are commonly found in Nature in extended oligosaccharides and polysaccharides. When recognized by protein receptors, dissecting the contribution of each repeating unit to the binding event is not easy. Here, the introduction of 13C‐labels at specific positions of a tri‐LacNAc repeating hexasaccharide has been used in STD‐HSQC NMR experiments to reveal the preferred binding epitopes for a panel of human galectins.