Mucin biosynthesis. Enzymic properties of human-tracheal epithelial GDP-L-fucose:beta-D-galactoside alpha-(1----2)-L-fucosyltransferase.
The human-tracheal, epithelial alpha-(1----2)-L-fucosyltransferase that transfers L-fucose from GDP-L-fucose to an acceptor containing a beta-D-galactopyranosyl group at the nonreducing terminal was characterized. Optimal enzyme activity was obtained at pH 6.5. 20-30mM MnCl2 (or CaCl2), and 0.05% Triton X-100 or 0.5% Tween 20. Mg2+ and Ba2+ ions moderately enhanced the enzyme activity, whereas Fe2+, Co2+, Zn2+, and Cd2+ ions were inhibitory. The enzyme activity was inhibited by N-ethylmaleimide and nucleotides of guanine, inosine, xanthine, and uridine. However, ATP and dithiothreitol did not affect the enzyme activity. The apparent Michaelis constant for GDP-L-fucose, freezing point-depressing glycoproteins (expressed as Gal----GalNAc----Thr), and phenyl beta-D-galactopyranoside was 0.29, 5.70, and 25.4mM, respectively. Under alkali-borohydride conditions (0.05M NaOH-M NaBH4, 45 degrees, 20 h), an L-[14C]fucosyltrisaccharide was released from the product obtained by use of freezing point-depressing glycoprotein as the acceptor. The alpha-L anomeric configuration of the fucoside was determined by the release of L-[14C]fucose from the purified trisaccharide by Turbo cornutus alpha-L-fucosidase. The (1----2) linkage of the L-fucosyl group to the D-galactosyl residue was established by methylation technique (m.s.-g.l.c.). The present enzyme has properties similar to those of the human milk alpha-(1----2)-L-fucosyltransferase which is encoded by a secretor gene.
D002413:Cations, Divalent; D004848:Epithelium / Q000201:enzymology; D005647:Fucosyltransferases / Q000378:metabolism*; D006602:Hexosyltransferases / Q000378:metabolism*; D006801:Humans; D006863:Hydrogen-Ion Concentration; D007700:Kinetics; D013058:Mass Spectrometry; D009077:Mucins / Q000096:biosynthesis*; D009844:Oligosaccharides / Q000302:isolation & purification; D014132:Trachea / Q000201:enzymology*; D000097631:Galactoside 2-alpha-L-fucosyltransferase
D002413:Cations, Divalent; D005647:Fucosyltransferases; D006602:Hexosyltransferases; D009077:Mucins; D009844:Oligosaccharides; D000097631:Galactoside 2-alpha-L-fucosyltransferase