Identification of an alpha3-fucosyltransferase and a novel alpha2-fucosyltransferase activity in cercariae of the schistosome Trichobilharzia ocellata: biosynthesis of the Fucalpha1-->2Fucalpha1-->3[Gal(NAc)beta1-->4]GlcNAc sequence.
Fucose is a major constituent of the protein- and lipid-linked glycans of the various life-cycle stages of schistosomes. These fucosylated glycans are highly antigenic and seem to play a role in the pathology of schistosomiasis. In this article we describe the identification and characterization of two fucosyltransferases (FucTs) in cercariae of the avian schistosome Trichobilharzia ocellata, a GDP-Fuc:[Galbeta1-->4]GlcNAcbeta-R alpha1-->3-FucT and a novel GDP-Fuc:Fucalpha-R alpha1-->2-FucT. Triton X-100 extracts of cercariae were assayed for FucT activity using a variety of acceptor substrates. Type 1 chain (Galbeta1-->3GlcNAc) based compounds were poor acceptors, whereas those based on a type 2 chain (Galbeta1-->4GlcNAc), whether alpha2'-fucosylated, alpha3'-sialylated, or unsubstituted, and whether present as oligosaccharide or contained in a glycopeptide or glycoprotein, all served as acceptor substrates. In this respect the schistosomal alpha3-FucT resembles human FucT V and VI rather than other known FucTs. N-ethylmaleimide, an inhibitor of several human FucTs, had no effect on the activity of the schistosomal alpha3-FucT, whereas GDP-beta-S was strongly inhibitory. Large scale incubations were carried out with Galbeta1-->4GlcNAc, GalNAcbeta1-->4GlcNAcbeta-O -(CH2)8COOCH3 and Fucalpha1-->3GlcNAcbeta1-->2Man as acceptor substrates and the products of the incubations were isolated using a sequence of chromatographic techniques. By methylation analysis and 2D-TOCSY and ROESY1H-NMR spectroscopy the products formed were shown to be Galbeta1-->4[Fucalpha1-->2Fucalpha1-->3]GlcNAc, GalNAcbeta1-->4[Fucalpha1-->2Fucalpha1-->3]GlcNAcbe ta-O-(CH2)8COOCH3, and Fucalpha1-->2Fucalpha1-->3GlcNAcbeta1-->2Man, respectively. It is concluded that the alpha2-FucT and alpha3-FucT are involved in the biosynthesis of the (oligomeric) Lewisx sequences and the Fucalpha1-->2Fucalpha1-->3GlcNAc structural element that have been described on schistosomal glycoconjugates.
Hokke
C H
CH
Department of Medical Chemistry Vrije Universiteit, Van der Boechorststraat 7, 1081 BT Amsterdam, The Netherlands.
Neeleman
A P
AP
Koeleman
C A
CA
van den Eijnden
D H
DH
D000818:Animals; D002236:Carbohydrate Conformation; D002240:Carbohydrate Sequence; D004372:Ducks; D005647:Fucosyltransferases / Q000378:metabolism*; D008401:Gas Chromatography-Mass Spectrometry; D006001:Glycoconjugates / Q000096:biosynthesis / Q000737:chemistry / Q000276:immunology; D006801:Humans; D007700:Kinetics; D016256:Lewis X Antigen / Q000096:biosynthesis / Q000737:chemistry; D008195:Lymnaea; D008969:Molecular Sequence Data; D009844:Oligosaccharides / Q000096:biosynthesis* / Q000737:chemistry / Q000276:immunology; D012551:Schistosomatidae / Q000201:enzymology* / Q000254:growth & development / Q000276:immunology; D013379:Substrate Specificity; D000097631:Galactoside 2-alpha-L-fucosyltransferase
D005647:Fucosyltransferases; D006001:Glycoconjugates; D016256:Lewis X Antigen; D009844:Oligosaccharides; D000097631:Galactoside 2-alpha-L-fucosyltransferase; C031692:galactoside 3-fucosyltransferase