Identification and characterization of a UDP-GalNAc:GlcNAcη-R η→4–N-acetylgalactosaminyltransferase from cercariae of the schistosome Trichobilharzia ocellata. Catalysis of a key step in the synthesis of N,N’-diacetyllactosediamino (lacdiNAc)-type glycans

Abstract

Three different stages of the avian schistosome Trichobilharzia ocellata appeared to contain a novel N-acetylgalactosaminyltransferase activity. To investigate its function in the biosynthesis of schistosome glycoconjugates, the enzyme was partially purified from cercariae, a free-living stage of the parasite, by affinity chromatography on UDP-Sepharose. Acceptor specificity studies showed that the enzyme catalyses the transfer of N-acetylgalactosamine (GalNAc) from UDP-GalNAc to oligosaccharides, glycopeptides and glycoproteins carrying a terminally beta-linked N-acetylglucosamine (GlcNAc) residue, regardless of the underlying structure. Analysis of the products obtained with GlcNAc and a desialylated and degalactosylated diantennary glycopeptide by 400 MHz 1H-NMR spectroscopy revealed that a GalNAc beta 1-->4GlcNAc (N,N'-diacetyllactosediamine,lacdiNAc) unit was formed. The enzyme can therefore be described as a UDP-GalNAc:GlcNAc beta-R beta 1-->4-N-acetylgalactosaminytransferase (beta 4-GalNAcT). Using specific acceptors, the enzyme could be distinguished from all other beta 4-GalNAcTs described to date, including the one from pituitary gland that is involved in the specific glycosylation of pituitary glycohormones. By contrast, the enzymatic properties of the schistosome beta 4-GalNAcT (except for the sugar-donor specificity) strongly resemble those of the beta 4-galactosyltransferase of higher animals, an enzyme which is known to control the synthesis of Gal1-->4GlcNAc (lacNAc)-type oligosaccharide chains. By analogy, the beta 4-GalNAcT is concluded to control the key step in the synthesis of lacdiNAc-type chains. LacdiNAc-type glycans are also common to the mollusc Lymnaea stagnalis, which is the intermediate host of T.ocellata. It is proposed that the schistosome beta 4-GalNAcT functions in the expression of specific carbohydrate structures that contribute to a molecular mimicry, enabling the schistosome to evade the defence system of the snail host.

Molecular Cloning and Enzymatic Characterization of a UDP-GalNAc:GlcNAcβ-R β1,4-N-Acetylgalactosaminyltransferase fromCaenorhabditis elegans

“…The doublet at 5.146 ppm and the signal at ␦ ϭ 2.013 ppm can be assigned to the H-1 and the CH 3 -NAc of GlcNAc␤1-S-pNP by analogy to the resonance po- sitions in GlcNAc␤1-4GlcNAc␤1-S-pNP (38). The doublet at ␦ ϭ 4.540 ppm and the signal at ␦ ϭ 2.077 ppm have shifts that are close to those reported for a ␤4-linked GalNAc residue (44,45). The NMR spectrum therefore confirms that the analyzed product is GalNAc␤1-4GlcNAc␤1-S-pNP.…”

Identification of a Novel UDP‐GalNAc:GlcNAcβ‐Rβ1–4 N‐Acetylgalactosaminyltransferase from the Albumen Gland and Connective Tissue of the Snail Lymnaea stagnalis

“…The function of the p4Gal-NAc-T activity in albumen glands is not known, but it is tempting to speculate that this enzyme controls the synthesis of N,N'-diacetyllactosediamine-containing oligosaccharide chains on albumen gland glycoproteins. Recently, an enzyme activity similar to the snail GalNAc-T activity has been described in cercariae of Trichobilharzia ocellata for which the parasite L. stagnalis is the intermediate host [41].…”

Monosialylated biantennary N-glycoforms containing GalNAc–GlcNAc antennae predominate when human EPO is expressed in goat milk

“…Attachment of GalNAc to GlcNAc has been thought to be controlled by an N-acetylgalactosaminyltransferase acting in competition with a b1,4-galactosyltransferase on terminal GlcNAc-containing oligosaccharides [38,39]. Even though no glycan-dependent function has been reported for the GalNAc-GlcNAc non-reducing terminals present in bovine milk glycoproteins, it has been postulated that the occurrence of such structures in milk is associated with high a-lactalbumin concentrations during the lactation.…”