Quantifying Protein-Glycan Interactions Using Native Mass Spectrometry

Interactions between glycan-binding proteins (GBPs) and carbohydrates (glycans) are essential to many biological processes relevant to human health and disease. For most GBPs, however, their glycan interactome-the repertoire of glycans recognized and their specificities-is poorly defined. The structural diversity of biologically relevant glycans and their limited availability in purified form, as well as their varied presentation, often as glycoconjugates, and weak affinities are key challenges hindering comprehensive glycan interaction mapping. Native mass spectrometry (nMS), a versatile, sensitive and label-free tool for the discovery of GBP-glycan interactions and quantifying their stoichiometry and thermodynamic parameters, is poised to play a leading role in defining the glycan interactome of GBPs. Here, we review established nMS methodologies, as well as important experimental and instrumental considerations, for detecting GBP-glycan interactions in vitro, and reliably measuring their stoichiometry and affinity. Recent advances in nMS methods for high-throughput library screening, including shotgun glycomics, and quantifying GBP interactions with glycoproteins and glycosphingolipids, are also described.