Studies on the three-dimensional behaviour of the selectin ligands Lewisa and sulphated Lewisa using NMR spectroscopy and molecular dynamics simulations
Sulphated blood group Lewisa/Lewisx (Lea/Lex) type sequences, with sulphate at the 3-position of galactose, have emerged as potent ligands for the endothelial adhesion molecule E-selectin and the leukocyte adhesion molecule L-selectin. As a first step in elucidating the molecular basis of the strong interactions with the selectins, we have performed conformational studies of the sulphated Lea in comparison with the non-sulphated analogue which is less strongly bound by E-selectin and not at all by L-selectin. Experimental NMR parameters [nuclear Overhauser effects (NOE) and interglycosidic 3JC, H] and theoretical values back-calculated from the minimum energy structures are in excellent agreement for both molecules. Molecular dynamics calculations for SuLea depict only minor torsional fluctuations around the glycosidic linkages over the time course of the 500 ps simulations, leading to the conclusion that the conformation of SuLea approximates to a singlerigid structure, as does the previously investigated Lea molecule. Comparison of experimentally and theoretically obtained parameters for SuLea with those for the nonsulphated Lea molecule indicate that no detectable changes occur in the three-dimensional structure of the trisaccharide upon sulphation. Thus, the enhanced selectin binding to the sulphated Lea is most likely due to favourable electrostatic interactions between the charged sulphate group and corresponding charged groups on the selectin protein.