Open AccessThe use of human milk fucosyltransferase in the synthesis of tumor‐associated trimeric X determinants
Author(s) -
VRIES Theodora,
NORBERG Thomas,
LÖNN Hans,
EIJNDEN Dirk H.
Publication year - 1993
Publication title -
european journal of biochemistry
Language(s) - English
Resource type - Journals
eISSN - 1432-1033
pISSN - 0014-2956
DOI - 10.1111/j.1432-1033.1993.tb18197.x
Subject(s) - fucosylation , fucosyltransferase , fucose , chemistry , oligosaccharide , trimer , residue (chemistry) , yield (engineering) , stereochemistry , biochemistry , high performance liquid chromatography , carbohydrate conformation , chromatography , dimer , nuclear magnetic resonance spectroscopy , enzyme , glycoprotein , organic chemistry , materials science , metallurgy
We have studied the fucosylaticn of a chemically synthesized trimer of N ‐acetyllactosamine [(LacNAc) 3 ‐EtPhNHCOCF 3 ] with a fucosyltransferase preparation from normal human milk, which utilizes both type‐1 and type‐2 structures, whether sialylated or not. When fucose residues were added enzymically to the (LacNAc) 3 ‐EtPhNHCOCF 3 hexasaccharide, mono‐, di‐, or trifucosylated oligosaccharide species were formed, containing the Lewis x determinant (Galβ1→4[Fucα1→3]GlcNAcβ1→3). With excess GDP‐fucose and prolonged reaction times, the trifucosylated product was formed in almost quantitative yield. Kinetic analysis of the fucosylation reaction indicated that there is a significant difference in the rate of transfer of the first, second and third fucose residues onto the acceptor molecule. The location of the fucose residues in the monofucosylated and difucosylated intermediate products was assessed by analyzing the digests obtained after endo‐β‐galactosidase treatment by HPLC and reverse‐phase chromatography. In addition, the fucosylated (LacNAc) 3 ‐EtPhNHCOCF 3 , structures were characterized by HPLC and were identified by 400‐MHz 1 H‐NMR spectroscopy. There is a highly preferred order in which the fucosyl residues are attached to (LacNAc) 3 ‐EtPhNHCOCF 3 . In the major pathway, the first two fucose residues are transferred with equal preference to the medial (GN3) and proximal (GN1) GlcNAc residues, whereas the third fucose is attached to the distal (GN5) GlcNAc residue. These results are of relevance in understanding the role of α‐3‐fucosyltransferase in the biosynthesis of Lewis x ‐related cell‐surface carbohydrate structures, that function as ligands for selectin‐type cell‐adhesion molecules and may play a role in the invasion and metastasis of several carcinoma.