guanosine-diphosphate and carbene

N-Acetyllactosamine derivative 4, which has a methylene amide tether between C-6 and C-2', was enzymatically glycosylated using rat liver alpha-2,6-sialyltransferase (ST6GalI) or recombinant human fucosyltransferase V (FucT-V) to give conformationally constrained trisaccharides 5 and 6, respectively. The methylene amide linker of 4 was installed by a two-step procedure, which involved acylation of a C-6 amino function of a LacNAc derivative with chloroacetic anhydride followed by macrocyclization by nucleophilic displacement of the chloride by a C-2' hydroxyl. The conformational properties of 4 were determined by a combination of NOE and trans-glycosidic heteronuclear coupling constant measurements and molecular mechanics simulations and these studies established that the glycosidic linkage of 4 is conformationally constrained and resides in only one of the several energy minima accessible to LacNAc. The apparent kinetic parameters of transfer to LacNAc and conformationally constrained saccharides 3 and 4 indicates that fucosyltransferase V recognize LacNAc in its A-conformer whereas alpha-2,6-sialyltransferase recognizes the B-conformer of LacNAc.

Topics: Amides; Amino Sugars; Animals; beta-D-Galactoside alpha 2-6-Sialyltransferase; Carbohydrate Conformation; Carbohydrate Sequence; Fucose; Fucosyltransferases; Guanosine Diphosphate; Hydrocarbons; Liver; Magnetic Resonance Spectroscopy; Methane; Models, Chemical; Models, Molecular; Models, Theoretical; Molecular Sequence Data; Oligosaccharides; Protein Conformation; Rats; Sialyltransferases; Trisaccharides