Open AccessThe glycosylphosphatidylinositol anchor of the trypomastigote‐specific Tc‐85 glycoprotein from Trypanosoma cruzi
Author(s) -
COUTO Alicia S.,
LEDERKREMER Rosa M.,
COLLI Walter,
ALVES M. Julia M.
Publication year - 1993
Publication title -
european journal of biochemistry
Language(s) - English
Resource type - Journals
eISSN - 1432-1033
pISSN - 0014-2956
DOI - 10.1111/j.1432-1033.1993.tb18282.x
Subject(s) - glycoprotein , trypanosoma cruzi , biochemistry , wheat germ agglutinin , trypanosoma brucei , phospholipase c , biology , glycan , affinity chromatography , exoglycosidase , monoclonal antibody , pronase , immunoprecipitation , microbiology and biotechnology , lectin , chemistry , trypsin , enzyme , antibody , parasite hosting , immunology , world wide web , computer science , gene
The Tc‐85 glycoprotein, specific for the infective stage of Trypanosoma cruzi , is anchored via glycosylphosphatidylinositol. The protein was purified from parasites, labeled metabolically with palmitic acid, by immunoprecipitation with the H 1 A 10 monoclonal antibody or by affinity column chromatography on wheat germ agglutinin. Antisera to the soluble form of the variant surface glycoprotein of Trypanosoma brucei brucei cross‐reacted with Tc‐85 when the immunoprecipitate was analysed by Western blotting. The reaction was intensified upon previous incubation of the glycoprotein with phosphatidylinositol‐specific phospholipase C. Such recognition was abolished when the cyclic phosphate was opened by mild acid treatment. The lipid cleaved by phospholipase C digestion, was identified as 1‐ O ‐hexadecylglycerol by reverse‐phase thin‐layer chromatography. The glycan core was deaminated and chemically labeled by reduction with NaB 3 H 4 . The labeled glycoprotein was exhaustively treated with pronase and dephosphorylated with 50% HF. Although microheterogeneity of the oligosaccharide moiety was apparent, by thin layer chromatography, a main spot coincident with Man(α1–2) Man(α1–6) Man(α1–4) anhydromannitol was shown, consistent with the conserved core structure of all glycosylphosphatidylinositol anchors analysed to date.