Sequential biosynthesis of sulfated and/or sialylated Lewis x determinantsby transferases of the human bronchial mucosa

The structural determination of sulfated carbohydrate chains from a cystic fibrosis patient respiratory mucins has shown that sulfation may occur eith er on the C-3 of the terminal Gal, or on the C-6 of the GlcNAc residue of a terminal N-acetyllactosamine unit. The two enzymes responsible for the tra nsfer of sulfate from PAPS to the C-3 of Gal or to the C-6 of GlcNAc residu es have been characterized in human respiratory mucosa, These two enzymes, in conjunction with fucosyl- and sialyltransferases, allow the synthesis of different sulfated epitopes such as 3-sulfo Lewis x (with a 3-O-sulfated G al), 6-sulfo Lewis x and 6-sulfo-sialyl Lewis x (with a 6-O-sulfated GlcNAc ), In the present study, the sequential biosynthesis of these epitopes has been investigated using microsomal fractions from human respiratory mucosa incubated with radiolabeled nucleotide-sugars or PAPS, and oligosaccharide accepters, mostly prepared from human respiratory mucins, The structures of the radiolabeled products have been determined by their coelution in HPAEC with known oligosaccharidic standards. In the biosynthesis of 6-O-sulfated carbohydrate chains by the human respiratory mucosa, the 6-O-sulfation of a terminal nonreducing GlcNAc residue precedes beta 1-4-galactosylation, al pha 2-3-sialylation (to generate 6-sulfo-sialyl-N-acetyllactosamine), and a lpha 1-3-fucosylation (to generate the 6-sulfo-sialyl Lewis x determinant). The 3-O-sulfation of a terminal N-acetyllactosamine may occur if this carb ohydrate unit is not substituted. Once an N-acetyllactosamine unit is synth esized, alpha 1-3-fucosylation of the GlcNAc residue to generate a Lewis x structure blocks any further substitution. Therefore, the present study def ines the pathways for the biosynthesis of Lewis x, sialyl Lewis x, sulfo Le wis x, and 6-sulfo-sialyl Lewis x determinants in the human bronchial mucos a.