NMR spectroscopy: an excellent tool to study protein interactions and glycosylation
Dr. Mario Schubert
Institute of Molecular Biology and Biophysics,
ETH Zürich, 8093 Zürich, Switzerland
Protein-carbohydrate interactions play a central role in the discrimination between self and nonself,a prerequisite for any defense mechanism. Such discrimination is crucial for innateimmunity in mammals as well as for defense strategies of plants or fungi. The molecular basis ofprotein-mediated fungal defense is largely unexplored. Here, I demonstrate the power of NMRspectroscopy to study the interaction of a novel fungal defense lectin, CCL2, from the ink capmushroom Coprinopsis cinerea with its in vivo target, the trisaccharideGlcNAcβ1,4[Fucα1,3]GlcNAc. This interaction is essential for toxicity of CCL2 towardsinvertebrates. Since this glycoepitope is characteristic for invertebrates and represents animportant allergen, these results suggest that the same glycoepitope is targeted by both fungaldefense and mammalian immune systems.In addition, I introduce a novel class of N-linked protein glycosylation that is found inpathogenic bacteria. In contrast to typical N-glycosylation that is achieved by membrane-boundoligosaccharyltransferase (OST) the novel N-glycosyltransferase (NGT) is active in thecytoplasm. Glycosylation is crucially important for the maturation of a bacterial adhesin andultimatively for adherence to human epithelial cells. Here NMR spectroscopy demonstrated thatNGT is able to add glucose moieties to asparagine residues of peptides and proteins in vitro andthat it is an inverting enzyme. In conclusion, NMR spectroscopy is an excellent technique todetect, localize and understand biomolecular interactions and modifications awaiting itsutilization in most fields of biology.