earticle

영어

For better understanding the molecular basis of the mechanism underlying oligosaccharides functions, it is quite desirable to gain detail information on their conformational dynamics in solution. However, the conformational analysis of oligosaccharides is still a remaining challenge, because of their branched covalent structures and dynamic conformational multiplicities, which hinder conventional analytical methods. For conformational characterization of flexible oligosaccharides, their structures should not be dealt with as a single well-defined global free energy minimum but as an ensemble of low energy conformers. Hence, we have been developing a methodology for evaluating a dynamic ensemble of oligosaccharide conformations by employing paramagnetic-assisted NMR methods based on paramagnetic effects such as pseudocontact shift (PCS) and paramagnetic relaxation enhancement (PRE) in conjunction with molecular dynamics (MD) simulations [1,2].
We applied this approach to the conformational dynamics analyses of branched GM2 tetrasaccharide, βGalNAc-(1-4)-[αNeu5Ac-(2-3)]-βGal-(1-4)-βGlc, which share the common core structure of gangliosides forming an integral part of cellular membranes. We synthesized the GM2 tetrasaccharide covalently attached to a lanthanide chelating-tag (Fig. 1). Upon complexation with paramagnetic lanthanide ions, the tagged sugar exhibited NMR spectral changes due to PCSs. The observed PCS values were in excellent agreement with those back-calculated from the 3D ensemble models derived from the MD calculations. In addition, we have developed spin-labeling method of gangliosides for their NMR analyses in amphipathic environments. These methodologies open a new prospect for conformational analyses of dynamic structures of gangliosides toward decoding glycocodes from the 3D structural aspects.