Acceptor specificity and tissue distribution of three human α‐3‐fucosyltransferases

Abstract

Based on the capacity to transfer a-L-fucose onto type-1 and type-2 synthetic blood group H and sialylated acceptors, a comparison of the a-3-fucosyltransferase activities of different human tissues is shown. Three distinct acceptor specificity patterns are described: (I) myeloid a-3-fucosyltransferase pattern, in which leukocytes and brain enzymes transfer fucose actively onto H type-2 acceptor and poorly onto sialylated N-acetyllactosamine; (11) plasma a-3-fucosyltransferase (EC 2.4.1.152), in which plasma and hepatocyte enzymes transfer, in addition, onto the sialylated N-acetyllactosamine; (111) Lewis a-3/4-fucosyltransferase (EC 2.4.1.65), in which gall-bladder, kidney and milk enzymes transfer, in addition, onto type-1 acceptors. The small amount (< 10%) of a-3-fucosyltransferase activity found in the plasma of an a-3-fucosyltransferase-deficient individual had a myeloidtype acceptor pattern, suggesting that this small proportion of the plasma enzyme is derived from leukocytes. In addition to the three acceptor specificity patterns, these enzyme activities can be differentiated by their optimum pH: 8.0-8.7 for the enzymes from myeloid cells and brain, 7.2-8.0 for liver enzymes and 6.0-7.2 for gallbladder enzymes. Milk samples had two a-3-fucosyltransferase activities, the Lewis or a-3/4-fucosyltransferase under control of the Lewis gene and an a-3-fucosyltransferase with plasma acceptor pattern which was independent of the control of the Lewis gene. The apparent affinity for GDP-fucose of the myeloid-like enzyme was weaker than those of the plasma and Lewis-like enzymes. The apparent affinities for H type 2 and sialylated Nacetyllactosamine were stronger for exocrine secretions as compared to the plasma and myeloid enzymes. The plasma type of a-3-fucosyltransferase activity was more sensitive to N-ethylmaleimide and heat inactivation than the samples with myeloid-like a-3-fucosyltransferase activity.The names X, Le", SSEA-1 and CD15 have been used to describe different properties of the same epitope, the terminal trisaccharide 3-fucosyl-N-acetyllactosamine or Galjl-+ 4(Fucal --f 3)GlcNAcP-, which is found in 0-and N-linked glycoproteins and in glycolipids [I].Many monoclonal antibodies, raised against different cells, were shown to react specifically with this epitope [2, 31. The first monoclonal with this specificity [4] was called SSEA-1 (stage-specific embryonic antigen-1) because it reacted with 8-cell-stage mouse embryos, morulae and early blastocysts. However, monoclonal antibodies reacting with normal human granulocytes [5], myeloid cell lines [6], human neuroblastoma, fetal brain [7], human gastric, colon and lung cancers [8] also had the same trisaccharide specificity.The epitope at the cell surface of leukocytes has been termed CD15 (cluster of differentiation 15) because a group of monoclonal antibodies with this specificity was identified

Significance of Individual Point Mutations, T202C and C314T, in the Human Lewis (FUT3) Gene for Expression of Lewis Antigens by the Human α(1,3/1,4)-Fucosyltransferase, Fuc-TIII

“…These acceptors were: (i) the disaccharide precursor of type 1, which can incorporate fucose in ␣1,2-linkage on the terminal Gal and in ␣1,4-linkage on the internal GlcNAc and (ii) the unambiguous trisaccharide H type 1, which can only incorporate fucose in the ␣1,4 position on the internal GlcNAc. This last trisaccharide was the better acceptor and it was more than twice as efficient as the disaccharide, confirming previous results (35,36). The 314 mutated construct gave results similar to the wild type FUT3 construct and no enzyme activity could be detected with either of the 1067 mutated or the double mutated 202-314 constructs, in good accordance with the fluorescence results (Table I).…”

“…Transfected cells were harvested after a 48-h growth period. (36,37). Enzyme kinetics were determined at initial velocity (15 min for the 314 mutated and the wild type FUT3 constructs and 14 h for the 202 mutated construct) for GDP-fucose and H type 1.…”

Pax6 Controls the Expression of Lewis x Epitope in the Embryonic Forebrain by Regulating α1,3-Fucosyltransferase IX Expression

“…These results suggest that Pax6 may regulate ␣3FucT activity in the E13.5 telencephalon. As the telencephalic cortex showed little synthesis of sialyl Lewis x glycan in contrast with Lewis x glycan (data not shown; described for human brain by Mollicone et al (61)), it is considered that the enzymatic properties of ␣3FucT in the rat cortex are similar to those of FucT-IV and FucT-IX in the other mammalian ␣3FucT family (see Introduction). These data suggest that Pax6 may control the activity of FucT-IV or FucT-IX in the rat embryonic brain, possibly by regulating the gene expression.…”

Relationship of Sialyl-Lewisx/a Underexpression and E-Cadherin Overexpression in the Lymphovascular Embolus of Inflammatory Breast Carcinoma

“…The ␣3-and ␣4-fucosyltransferase assays were performed with different synthetic oligosaccharide acceptors, H-type-2 and H-type-1, respectively, synthesized with a hydrophobic arm (Ϫ(CH 2 ) 8 -COOCH 3 ) (Calbiochem, San Diego, CA) that allows the glycosides to be retained on a Sep-Pak C 18 cartridge (Waters, Milford, MA). 10,11 The reaction mixture was prepared on ice, contained in a volume of 60 l: 30 g tumoral extract, 35 mmol/L Tris/HCl, pH 7.2, 4 mmol/L ATP, 20 mmol/L MnCl 2 , 10 mmol/L ␣-L-fucose, 5 l of a 1m/ml solution of the oligosaccharide acceptors (Htype-2: Fuc␣1-2Gal␤1-4GlcNAc-R); H-type-1: Fuc␣1-2Gal␤1-3GlcNAc-R) and 3.5 mol/L GDP-[ 14 C]-fucose (10 GBq/mmol; Amersham, Piscataway, NJ). The mixture was incubated for 16 hours at 37°C and the reaction was stopped by the addition of 3 ml of cold water and centrifuged.…”