Abstract

To date, glycosyltransferases (GTs) are an essential family of enzymes poorly characterized both structurally and mechanistically which is being a major bottleneck in Glycoscience. These enzymes play a crucial role in living organisms, catalysing the stereo- and regiospecific transfer of an activated donor sugar to an acceptor moiety to build up complex oligosaccharides onto the cell surface. New analytical tools are required to screen enzyme-glycan interactions in high-throughput manner. In this context, Surface Plasmon Resonance imaging (SPRi) has spread as a versatile methodology to overcome this lack in the glycoscience field, for ligands screening and biomolecular interaction monitoring in a real time and without any labels.

Therefore, this multidisciplinary research project is organized in three main axes:  first, we address the heterologous expression and purification of a fucosyltransferase from the plant Arabidopsis thaliana (AtFUT1) that participates in the last step in the biosynthesis of xyloglucan (XyG) ligand. Then, specific on-surface immobilization approaches motivate the design of chemoselective strategies to conjugate XyG building-blocks to DNA scaffolds in good yields. The versatility of having ODN structures conjugated to glycans provides powerful insights for glycan purification, characterization and on-surface immobilization. Finally, the third axis is dedicated towards the conception and construction of a versatile XyG glycochip for its specific immobilization approach by DNA-Directed Immobilization (DDI). This strategy is envisioned through the powerful SPRi technique that allows the simultaneous visualization and characterization of biomolecular interactions in real time measurements, without using any label.