title: Structural analysis of glycosyl-phosphatidylinositol membrane anchor of the Toxoplasma gondii tachyzoite surface glycoprotein gp23.
abstract: In this study we describe the biochemical features of the Toxoplasma gondii tachyzoite surface glycoprotein, gp23, demonstrating that it is attached to the parasite membrane by a glycosyl-phosphatidyl inositol anchor. Gp23 was metabolically labeled with tritiated palmitate, myristate, ethanolamine, inositol, glucosamine, mannose and galactose, as expected for a GPI-anchor structure. Gp23 was released from the surface of living parasites after treatment with phosphatidylinositol-specific phospholipase C(PI-PLC) and the resulting water-soluble protein was immunoprecipitated with a monoclonal antibody specific for gp23. The GPI-core glycan was generated after aqueous-HF dephosphorylation followed by nitrous acid deamination and its carbohydrate structure was analyzed using selective exo- and endoglycosidase treatments. Finally, the phosphatidylinositol moiety of gp23 was characterized using PI-PLC and phospholipase A2 (PLA2) digestions. Our cumulative data suggest that gp23 of T gondii tachyzoites contains a modified GPI-backbone similar to the mammalian Thy-1 anchor, consisting of a conserved core structure (ethanolamine-PO4-6-Man alpha1-2-Man alpha1-6-Man alpha1-4-GlcN alpha1-6-PI) bearing beta-linked N-acetylgalactosamine residue(s).
authors.affiliation: Zentrum für Hygiene und Medizinische Mikrobiologie, Philipps- Universitaet Marburg, Germany.
authors.forename: S
authors.initials: S
authors.lastname: Tomavo
authors.forename: J F
authors.initials: JF
authors.lastname: Dubremetz
authors.forename: R T
authors.initials: RT
authors.lastname: Schwarz
mesh_terms: D000818:Animals; D000953:Antigens, Protozoan / Q000096:biosynthesis / Q000737:chemistry* / Q000302:isolation & purification; D050260:Carbohydrate Metabolism; D002846:Chromatography, Affinity; D004075:Diglycerides / Q000032:analysis; D019856:Ethanolamine; D004983:Ethanolamines / Q000378:metabolism; D017261:Glycosylphosphatidylinositols / Q000737:chemistry* / Q000302:isolation & purification / Q000378:metabolism; D007294:Inositol / Q000378:metabolism; D008562:Membrane Glycoproteins / Q000096:biosynthesis / Q000737:chemistry* / Q000302:isolation & purification; D019814:Myristic Acid; D009227:Myristic Acids / Q000378:metabolism; D019308:Palmitic Acid; D010169:Palmitic Acids / Q000378:metabolism; D015800:Protozoan Proteins / Q000096:biosynthesis / Q000737:chemistry* / Q000302:isolation & purification; D014122:Toxoplasma / Q000378:metabolism*; D014316:Tritium; D014709:Vero Cells
chemical_list: D000953:Antigens, Protozoan; D004075:Diglycerides; D004983:Ethanolamines; D017261:Glycosylphosphatidylinositols; D008562:Membrane Glycoproteins; D009227:Myristic Acids; D010169:Palmitic Acids; D015800:Protozoan Proteins; D019814:Myristic Acid; D014316:Tritium; D019308:Palmitic Acid; D007294:Inositol; D019856:Ethanolamine