Lytic anti-alpha-galactosyl antibodies from patients with chronic Chagas' disease recognize novel O-linked oligosaccharides on mucin-like glycosyl-phosphatidylinositol-anchored glycoproteins of Trypanosoma cruzi.
Sera of patients with chronic Chagas' disease (American trypanosomiasis) contain elevated levels of anti-alpha-galactosyl antibodies that are lytic to Trypanosoma cruzi. The T. cruzi trypomastigote F2/3 antigen complex recognized by these antibodies runs as a broad smear on SDS/PAGE [Almeida, Krautz, Krettli and Travassos (1993) J. Clin. Lab. Anal. 7, 307-316]. Treatment of T. cruzi trypomastigote cells with bacterial phosphatidylinositol-specific phospholipase C (PI-PLC) abolished most of their reactivity to chronic Chagas'-disease ((Chagasic, Ch) anti-alpha-galactosyl antibodies (anti-Gal). The F2/3 antigen complex, purified by solvent extraction and hydrophobic-interaction chromatography, contained 60% carbohydrate by weight and substantial amounts of Thr, Ser, Glx, Asx, Gly, Ala and Pro, but relatively few hydrophobic amino acids. The presence of myoinositol, ethanolamine and 1-O-hexadecylglycerol suggested the presence of glycosyl-phosphatidylinositol membrane anchors. This was confirmed by PI-PLC treatment, which rendered the F2/3 molecules hydrophilic and reactive to anti-(cross-reacting determinant) antibodies. The majority of the GlcNAc content of the F2/3 antigens was found at the reducing termini of oligosaccharides in O-glycosidic linkage to Thr residues. These O-linked oligosaccharides could be released by beta-elimination and by mild hydrazinolysis. The smallest released oligosaccharitol that was reactive with the Ch anti-Gal was Gal alpha 1-3Gal beta 1-4GlcNAcol (where GlcNAcol is N-acetyl-glucosaminitol). Several other Gal-containing oligosaccharitols were observed, most of which were branched and contained 4,6-di-O-substituted GlcNAcol at their reducing termini. About half of the total released oligosaccharitols could bind to immobilized Ch anti-Gal, but none of them bound to the anti-Gal isolated from normal human sera. These data suggest that the specificities of the Ch anti-Gal are quite different from the natural anti-Gal isolated from normal human sera. Therefore, these novel T. cruzi O-linked oligosaccharides are highly immunogenic under the conditions of natural infection and are the targets for lytic Ch anti-Gal.
Almeida
I C
IC
Escola Paulista de Medicina, São Paulo, Brazil.
Ferguson
M A
MA
Schenkman
S
S
Travassos
L R
LR
D000818:Animals; D000913:Antibodies, Protozoan / Q000276:immunology* / Q000378:metabolism; D000918:Antibody Specificity; D001894:Borohydrides / Q000494:pharmacology; D002240:Carbohydrate Sequence; D014355:Chagas Disease / Q000276:immunology*; D005690:Galactose / Q000276:immunology*; D005692:Galactose Oxidase / Q000378:metabolism; D006023:Glycoproteins / Q000276:immunology* / Q000302:isolation & purification / Q000378:metabolism; D017261:Glycosylphosphatidylinositols / Q000276:immunology* / Q000378:metabolism; D006801:Humans; D006834:Hydrazines / Q000494:pharmacology; D008969:Molecular Sequence Data; D009077:Mucins / Q000276:immunology* / Q000378:metabolism; D009844:Oligosaccharides / Q000276:immunology* / Q000378:metabolism; D015800:Protozoan Proteins / Q000276:immunology* / Q000378:metabolism; D014349:Trypanosoma cruzi / Q000276:immunology*
D000913:Antibodies, Protozoan; D001894:Borohydrides; D006023:Glycoproteins; D017261:Glycosylphosphatidylinositols; D006834:Hydrazines; D009077:Mucins; D009844:Oligosaccharides; D015800:Protozoan Proteins; D005692:Galactose Oxidase; D005690:Galactose