T. Devries et al., THE USE OF HUMAN-MILK FUCOSYL-TRANSFERASE IN THE SYNTHESIS OF TUMOR-ASSOCIATED TRIMERIC-X DETERMINANTS, European journal of biochemistry, 216(3), 1993, pp. 769-777
We have studied the fucosylation of a chemically synthesized trimer of
N-acetyllactosamine [(LacNAc)3-EtPhNHCOCF3] with a fucosyltransferase
preparation from normal human milk, which utilizes both type-1 and ty
pe-2 structures, whether sialylated or not. When fucose residues were
added enzymically to the (LacNAc)3-EtPhNHCOCF, hexasaccharide, mono-,
di-, or trifucosylated oligosaccharide species were formed, containing
the Lewis(x) determinant (Galbeta1-->4[Fucalpha1-->3]GlcNAcbeta1-->3)
. With excess GDP-fucose and prolonged reaction times, the trifucosyla
ted product was formed in almost quantitative yield. Kinetic analysis
of the fucosylation reaction indicated that there is a significant dif
ference in the rate of transfer of the first, second and third fucose
residues onto the acceptor molecule. The location of the fucose residu
es in the monofucosylated and difucosylated intermediate products was
assessed by analyzing the digests obtained after endo-beta-galactosida
se treatment by HPLC and reverse-phase chromatography. In addition, th
e fucosylated (LacNAc)3-EtPhNHCOCF, structures were characterized by H
PLC and were identified by 400-MHz H-1-NMR spectroscopy. There is a hi
ghly preferred order in which the fucosyl residues are attached to (La
cN-Ac)3-EtPhNHCOCF3. In the major pathway, the first two fucose residu
es are transferred with equal preference to the medial (GN3) and proxi
mal (GN1) GlcNAc residues, whereas the third fucose is attached to the
distal (GN5) GlcNAc residue. These results are of relevance in unders
tanding the role of alpha-3-fucosyltransferase in the biosynthesis of
Lewis(X)-related cell-surface carbohydrate structures, that function a
s ligands for selectin-type cell-adhesion molecules and may play a rol
e in the invasion and metastasis of several carcinoma.