Expression of N-linked sialyl Lex determinants and O-glycans in the carbohydrate moiety of human amniotic fluid transferrin during pregnancy

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Abstract
Transferrin, a glycoprotein involved in iron transport in body fluids, was isolated from amniotic fluid of a hydramnios patient by sequential anion-exchange chromatography and gel filtration. The N-glycans of human amniotic fluid transferrin (hAFT) were enzymatically liberated by PNGase-F digestion, isolated by gel filtration and fractionated by (high-pH) anion-exchange chromatography. After alkaline borohydride treatment of native hAFT, the released O-glycans were isolated by gel filtration and fractionated by anion-exchange chromatography. Structure elucidation of 14 N- and 2 O-glycans was performed by 500 or 600 MHz 1H-NMR spectroscopy. Besides conventional N-glycans established earlier for human serum transferrin (hST), new (α1–3)-fucosylated N-glycans were found, representing sialyl Lex elements. Furthermore, as compared to hST, a higher degree of (α1–6)-fucosylation and an increase in branching from di- to triantennary compounds has been detected. The presence of O-glycans is demonstrated for the first time in transferrin.