STUDIES ON THE 3-DIMENSIONAL BEHAVIOR OF THE SELECTIN LIGANDS LEWIS(A) AND SULFATED LEWIS(A) USING NMR-SPECTROSCOPY AND MOLECULAR-DYNAMICS SIMULATIONS

Sulphated blood group Lewis(a)/Lewis(x) (Le(a)/Le(x)) type sequences, with sulphate at the 3-position of galactose, have emerged as potent l igands for the endothelial adhesion molecule E-selectin and the leukoc yte adhesion molecule L-selectin. As a first step in elucidating the m olecular basis of the strong interactions with the selectins, we have performed conformational studies of the sulphated Lea in comparison wi th the non-sulphated analogue which is less strongly bound by E-select in and not at all by L-selectin. Experimental NMR parameters [nuclear Overhauser effects (NOE) and interglycosidic (3)J(C,H)] and theoretica l values back-calculated from the minimum energy structures are in exc ellent agreement for both molecules. Molecular dynamics calculations f or SuLe(a) depict only minor torsional fluctuations around the glycosi dic linkages over the time course of the 500 ps simulations, leading t o the conclusion that the conformation of SuLe(a) approximates to a si ngle-rigid structure, as does the previously investigated Lea molecule . Comparison of experimentally and theoretically obtained parameters f or SuLe(a) with those for the nonsulphated Le(a) molecule indicate tha t no detectable changes occur in the three-dimensional structure of th e trisaccharide upon sulphation. Thus, the enhanced selectin binding t o the sulphated Le(a) is most likely due to favourable electrostatic i nteractions between the charged sulphate group and corresponding charg ed groups on the selectin protein.