Download NMR spectroscopy: an excellent tool to study protein

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 innate
immunity in mammals as well as for defense strategies of plants or fungi. The molecular basis of
protein-mediated fungal defense is largely unexplored. Here, I demonstrate the power of NMR
spectroscopy to study the interaction of a novel fungal defense lectin, CCL2, from the ink cap
mushroom
Coprinopsis
cinerea
with
its
in
vivo
target,
the
trisaccharide
GlcNAcβ1,4[Fucα1,3]GlcNAc. This interaction is essential for toxicity of CCL2 towards
invertebrates. Since this glycoepitope is characteristic for invertebrates and represents an
important allergen, these results suggest that the same glycoepitope is targeted by both fungal
defense and mammalian immune systems.
In addition, I introduce a novel class of N-linked protein glycosylation that is found in
pathogenic bacteria. In contrast to typical N-glycosylation that is achieved by membrane-bound
oligosaccharyltransferase (OST) the novel N-glycosyltransferase (NGT) is active in the
cytoplasm. Glycosylation is crucially important for the maturation of a bacterial adhesin and
ultimatively for adherence to human epithelial cells. Here NMR spectroscopy demonstrated that
NGT is able to add glucose moieties to asparagine residues of peptides and proteins in vitro and
that it is an inverting enzyme. In conclusion, NMR spectroscopy is an excellent technique to
detect, localize and understand biomolecular interactions and modifications awaiting its
utilization in most fields of biology.