Chemoenzymatic Synthesis of Sulfated Glycosphingolipids for Probing Antibody specificities, Glycan-Binding Protein Interactions, and Cell Surface Remodeling

Glycosphingolipids (GSLs) are essential components of cell membranes that play key roles in immune recognition, signal transduction, and interactions with pathogens. This work brings together advanced synthetic techniques, targeted cell-surface remodeling, and detailed molecular profiling to improve our understanding of how GSLs function. The study begins with a neochemoenzymatic synthesis platform that uses enzyme-catalyzed glycosylation to create a wide range of well-defined glycans. By carefully controlling stereoselectivity, regioselectivity, and the incorporation of key modifications such as sulfate groups, this method produces GSLs that closely resemble those found in nature. These synthetic glycans overcome traditional challenges in GSL production and provide valuable tools for further study. Next, the synthesized GSLs are incorporated into cell membranes to remodel the cell surface. This approach allows for the examination of glycan-dependent immune responses, particularly through complement-mediated cytotoxicity. The experiments show how specific glycosylation patterns can influence the activation of complement pathways, affecting cell survival. These findings offer new insights into the mechanism of disease formation in anti-MAG neuropathy. In addition, a high-throughput glycan microarray is used to explore the interactions between glycans, serum auto-antibodies, and other glycan-binding proteins. This platform enables a comprehensive analysis that uncovers binding patterns differing from conventional expectations, further clarifying the molecular underpinnings of anti-MAG neuropathy. By integrating sophisticated synthetic methodologies, targeted cell-surface glycoengineering, advanced microarray technology, and cutting-edge analytical techniques, this work not only elucidates the fundamental roles of glycosphingolipids in immune recognition and cell-surface dynamics but also paves the way for novel glycan-based diagnostics and interventions in autoimmune and neurodegenerative diseases.