Purification of the blood group H gene associated alpha-2-L-fucosyltransferase from human plasma.
The alpha-2-L-fucosyltransferase in human plasma has been freed from alpha-3-L-fucosyltransferase activity and purified approximately 200,000-fold by a series of steps involving ammonium sulphate precipitation, hydrophobic chromatography on Phenyl Sepharose 4B and affinity chromatography first on GDP-adipate-Sepharose and then on GDP-hexanolamine-Sepharose. The purified alpha-2-L-fucosyltransferase had a M(r) on gel filtration HPLC of 158,000 and showed optimal activity in the pH range 6.5-7.0. The enzyme transferred fucose equally well to Type 1 (Gal beta 1-3GlcNAc) and Type 2 (Gal beta 1-4GlcNAc) substrates but Type 3 (Gal beta 1-3GalNAc) structures were less efficient acceptors. Competition experiments indicated that a single enzyme species in the purified preparation was responsible for reactivity with the Type 1 and Type 2 structures. Thus the differences in conformation between the Type 1 and Type 2 disaccharides do not appear to influence the capacities of their terminal non-reducing beta-D-galactosyl residues to function as acceptor substrates for the alpha-2-L-fucosyltransferase expressed by the blood group H gene in haemopoietic tissue.
Kyprianou
P
P
Division of Immunochemical Genetics, MRC Clinical Research Centre, Harrow, Middlesex, U.K.
Betteridge
A
A
Donald
A S
AS
Watkins
W M
WM
D001667:Binding, Competitive; D002240:Carbohydrate Sequence; D002851:Chromatography, High Pressure Liquid; D004591:Electrophoresis, Polyacrylamide Gel; D005647:Fucosyltransferases / Q000097:blood* / Q000737:chemistry / Q000235:genetics / Q000302:isolation & purification; D006801:Humans; D006863:Hydrogen-Ion Concentration; D008969:Molecular Sequence Data; D008970:Molecular Weight; D013379:Substrate Specificity; D000097631:Galactoside 2-alpha-L-fucosyltransferase
D005647:Fucosyltransferases; D000097631:Galactoside 2-alpha-L-fucosyltransferase