Thursday, March 20, 2025
3/20/2025
Project Summary Sulfation is the most abundant and diverse glycan modification which tunes the physical properties and biological activity of residing glycans and glycoproteins. Other than sulfated glycosaminoglycans (GAGs), sulfation is also commonly identified on N-glycans and O-glycans but is often overlooked. These glycans are believed to play critical roles in many physiological/pathological processes and glycan-protein interactions, the first example of which was the discovery of sulfated glycan ligand (on both N-glycans and O-glycans) for L- selectin in lymphocyte homing and recruitment. Many Siglec-glycan ligand interactions are also found to be “sulfation dependent” or enhance by sulfation. Nevertheless, current knowledge of the structures and functions of these sulfated glycans is still limited, and mainly concluded from genetic studies and the use of simple sulfated glycan determinants, which may not be complete or accurate as underlying structures significantly affect glycan- protein interactions. Despite the significance, systematic synthesis of sulfated N-glycans and O-glycans has never been achieved, which represents a major obstacle in understanding glycan sulfation. A major challenge is the great diversity which is derived from multiple branches and cores of N-glycans and O-glycans, as well as many potential locations of sulfate groups on them. The presence of keratan sulfate chains on N-glycans and O- glycans further expands the sulfated glycan repository. To completely solve this problem, we propose to develop a chemoenzymatic modular synthesis platform, by integrating sulfation modules into our current modular synthesis platform for non-sulfated N-glycans and O-glycans. Mammalian sulfotransferases will be expressed, fully characterized in terms of activity and acceptor specificity, and used to construct efficient sulfation modules. Optimized sulfation modules will then be combined with glycosylation modules to synthesize sulfated structures. The modular synthetic process will be well-programmed based on the substrate specificity of involved enzymes. A comprehensive library (300+) of sulfated N-glycan, O-glycan, and keratan sulfates will be prepared to showcase the powerfulness of the modular synthesis platform. Multiple chemical or enzymatic modulations will be introduced to achieve regio- and branch-selective sulfation. The resulting compounds are important standards for glycomics analysis and probes to understand the functions of sulfated glycans as well as many biological processes. The modular synthesis process is programmable and can be readily adopted for automated synthesis.
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