STRUCTURAL CHARACTERIZATION OF THE MAJOR GLYCOSYLPHOSPHATIDYLINOSITOLMEMBRANE-ANCHORED GLYCOPROTEIN FROM EPIMASTIGOTE FORMS OF TRYPANOSOMA-CRUZI Y-STRAIN

We have investigated the structure of the glycosylphosphatidylinositol (GPI) anchor and the O-linked glycan chains of the 40/45-kDa glycopro tein from the cell surface of the protozoan parasite Trypanosoma cruzi , This glycoconjugate is the major acceptor for sialic acid transferre d by trans sialidase of T. cruzi Y-strain, epimastigote form. The GPI anchor was liberated by treatment with hot alkali, and the phosphoinos itol-oligosaccharide moiety was characterized and shown to have the fo llowing structure. [GRAPHICS] STRUCTURE 1 Unusually the glucosamine wa s 6-O-substituted with 2-aminoethylphosphonate, and 2-aminoethylphosph onate was also present on the third mannose residue distal to glucosam ine, partially replacing the ethanolamine phosphate, The beta-eliminat ed reduced oligosaccharide chains showed that two novel classes of O-l inked N-acetylglucosamine oligosaccharide were present. The first seri es had the structures Galp beta 1-3GlcNAc-ol; Galp beta-6(Galp beta 1- 3)GlcNAc-ol; and Galp beta 1-2Galp beta 1-6(Galp beta 1-3)GlcNAc-ol, w hereas the other series had a 1-4 linkage to N-acetylglucosaminitol an d had structures Galp beta 1-4GlcNAc-ol, Galp beta 1-6(Galp beta 1-4)G lcNAc-ol, and Galp beta 1-2Galp beta 1-6(Galp beta 1-4)GlcNAc-ol. We h ave also investigated the kinetics of in vitro sialylation of these O- linked oligosaccharides by the T. cruzi trans-sialidase and have shown that incorporation of one molecule of sialic acid hinders entry of a second molecule when two potential acceptor sites are present.