The abundance of additional N-acetyllactosamine units in N-linked tetraantennary oligosaccharides of human Tamm-Horsfall glycoprotein is a donor-specific feature

Previously, treatment of Tamm-Horsfall glycoprotein (THp) from different donors with endo-β-galactosidase has been shown to liberate a tetra- and a Sda-active pentasaccharide, concluding the presence of N-linked carbohydrate chains containing additional N-acetyllactosamine units. These type of oligosaccharides were not found in a detailed structure elucidation of the carbohydrate moiety of THp of one male donor, suggesting a donor-specific feature for these type of structures. Therefore, THp was isolated from four healthy male donors and each subjected to endo-β-galactosidase treatment in order to release these tetra- and Sda-active pentasaccharide. Differences were observed in the total amount of released tetra- and Sda-active pentasaccharide of the used donors (42, 470, 478, 718 μg/100 mg THp), indicating that the presence of repeating N-acetyllactosamine units incorporated into the N-glycan moiety of THp is donor specific. Furthermore, a higher expression of the Sda determinant on antennae which display N-acetyllactosamine elongation was observed, suggesting a better accessibility for the β-N-acetylgalactosaminyltransferase. In order to characterize the N-glycans containing repeating N-acetyllactosamine units, carbohydrate chains were enzymatically released from THp and isolated. The tetraantennary fraction, which accounts for more than 33% of the total carbohydrate moiety of THp, was used to isolate oligosaccharides containing additional N-acetyllactosamine units. Five N-linked tetraantennary oligosaccharides containing a repeating N-acetyllactosamine unit were identified, varying from structures bearing four Sda determinants to structures containing no Sda determinant. One compound was used in order to specify the branch location of the additional N-acetyllactosamine unit, and it appeared that only the Gal-6' and Gal-8' residues were occupied by a repeating N-acetyllactosamine unit.