Evidence for N-linked glycosylation in Toxoplasma gondii.
In this paper we report experiments demonstrating the presence of N-linked oligosaccharide structures in Toxoplasma gondii tachyzoites, providing the first direct biochemical evidence that this sporozoan parasite is capable of synthesizing N-linked glycans. The tachyzoite surface glycoprotein gp23 was metabolically labelled with [3H]glucosamine and [3H]mannose. Gel-filtration chromatography on Bio-Gel P4 columns produced four radiolabelled N-linked glycopeptides which were sensitive to peptidase-N-glycanase F, but resistant to endoglycosidases H and F. Using chemical analysis and exoglycosidase digestions followed by Dionex-high-pH anion-exchange chromatography and size fractionation on Bio-Gel P4 we show that gp23 has N-linked glycans in the hybrid- or complex-type structure composed of N-acetylgalactosamine, N-acetylglucosamine and mannose and devoid of sialic acid and fucose residues. In addition, the sensitivity of glycopeptides from glycoprotein extracts to endoglycosidases H and F revealed the in vivo synthesis of oligomannose-type structures by T. gondii tachyzoites. We have extended these findings by demonstrating the ability of T. gondii microsomes to synthesize in vitro a glucosylated lipid-bound high-mannose structure (Glc3Man9GlcNAc2) that is assumed to be identical with the common precursor for N-glycosylation in eukaryotes.
Odenthal-Schnittler
M
M
Zentum für Hygiene und Medizinische Mikrobiologie, Philipps-Universität Marburg, Germany.
Tomavo
S
S
Becker
D
D
Dubremetz
J F
JF
Schwarz
R T
RT
D000581:Amidohydrolases / Q000378:metabolism; D000818:Animals; D002236:Carbohydrate Conformation; D002462:Cell Membrane / Q000737:chemistry / Q000378:metabolism; D002845:Chromatography; D005690:Galactose / Q000378:metabolism; D005944:Glucosamine / Q000378:metabolism; D006023:Glycoproteins / Q000032:analysis / Q000737:chemistry / Q000378:metabolism*; D006026:Glycoside Hydrolases / Q000378:metabolism; D006031:Glycosylation; D008358:Mannose / Q000378:metabolism; D009844:Oligosaccharides / Q000032:analysis / Q000378:metabolism*; D043524:Peptide-N4-(N-acetyl-beta-glucosaminyl) Asparagine Amidase; D011498:Protein Precursors / Q000378:metabolism; D014122:Toxoplasma / Q000737:chemistry / Q000378:metabolism*; D014709:Vero Cells
D006023:Glycoproteins; D009844:Oligosaccharides; D011498:Protein Precursors; D006026:Glycoside Hydrolases; D000581:Amidohydrolases; D043524:Peptide-N4-(N-acetyl-beta-glucosaminyl) Asparagine Amidase; D005944:Glucosamine; D008358:Mannose; D005690:Galactose