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

The time-averaged solution conformation of a unique bis-sulfated glycolipid (HSO3)(2)-2,6Man alpha -2Glc alpha -1-sn-2,3-O-alkylglycerol, was studied in terms of the torsional angles of two glycosidic linkages, phi (H1-C1-O-C x) and psi (C1-O-Cx-Hx), derived from heteronuclear three-bond coupling con stants ((3)J(C,H)), and inter-residual proton-proton distances from J-HMBC 2D and ROESY experiments, respectively. The dihedral angles of Glc alpha 1G ro in glycolipids were determined for the first time. The C1-C4 diagonal li ne of the alpha -glucose ring makes an angle of approximate to 120 degrees with the glycerol backbone, suggesting that the alpha -glucose ring is almo st parallel to the membrane surface in contrast with the perpendicular orie ntation of the beta -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 Glc alpha 1Gro linkage has a single minimum-energy structure. On the other hand, three conformers were observed for the Man alpha 2Glc linkage. The flexibility of Man alpha 2Glc was further confirmed by the absence of inter-residual hydrogen bonds which were judged from the temperature coefficients of the chemical shifts, d de lta /dT (-10(-3) p.p.m..degreesC(-1)), of hydroxy protons. The conformation al flexibility may facilitate interaction of extracellular substances with both sulfate groups.