Conformational studies on a unique bis-sulfated glycolipid using NMR spectroscopy and molecular dynamics simulations

Abstract

The time-averaged solution conformation of a unique bis-sulfated glycolipid (HSO 3 ) 2 -2,6Mana-2Glca-1-sn-2,3-O-alkylglycerol, was studied in terms of the torsional angles of two glycosidic linkages, f (H1-C1-O-Cx) and c (C1-O-Cx-Hx), derived from heteronuclear three-bond coupling constants ( 3 J C,H ), and inter-residual proton± proton distances from J-HMBC 2D and ROESY experiments, respectively. The dihedral angles of Glca1Gro in glycolipids were determined for the first time. The C1-C4 diagonal line of the a-glucose ring makes an angle of < 120 8 with the glycerol backbone, suggesting that the a-glucose ring is almost parallel to the membrane surface in contrast with the perpendicular orientation of the b-isomer. Furthermore, minimum-energy states around the conformation were estimated by Monte Carlo/stochastic dynamics (MCSD) mixed-mode simulations and the energy minimization with assisted model building and energy refinement (AMBER) force field. The Glca1Gro linkage has a single minimum-energy structure. On the other hand, three conformers were observed for the Mana2Glc linkage. The flexibility of Mana2Glc was further confirmed by the absence of inter-residual hydrogen bonds which were judged from the temperature coefficients of the chemical shifts, dd/dT (210 23 p.p.m.´8C 21), of hydroxy protons. The conformational flexibility may facilitate interaction of extracellular substances with both sulfate groups.