Glycophosphatidylinositol-anchored proteins in metacyclic trypomastigotes of Trypanosoma cruzi.
We searched for the presence of glycophosphatidylinositol (GPI)-anchored proteins in epimastigotes and metacyclic trypomastigotes of Trypanosoma cruzi, by treatment of parasite lysates with the GPI-specific phospholipase C of Trypanosoma brucei. Upon treatment, several proteins (70-90 kDa) in metacyclics, but none in epimastigotes, reacted with antibodies to the cross-reacting determinant (CRD), an epitope revealed on the variant surface glycoproteins of T. brucei following removal of the diacylglycerol moiety from their GPI-anchor. Since these T. cruzi metacyclic proteins also lost their original amphiphilicity, as judged by Triton X-114 phase separation, it is very likely that they are linked to the membrane by GPI. One of these proteins is the 90 kDa protein, the major surface protein of G and Tulahuen strains, recognized by the monoclonal antibody 1G7. A variable portion of the 90 kDa molecules was resistant to solubilization by T. brucei lipase. The reasons for this are not clear but susceptibility appeared to increase with the age of the T. cruzi culture. Enzymes that solubilize GPI-anchored proteins were detected in epimastigotes and metacyclics, but the enzymatic activity in these forms was smaller than the activity detected in the same cell numbers of trypomastigotes of T. cruzi originated from infected mammalian cells or from T. brucei bloodstream forms. A preliminary characterization of these activities indicates that at least two classes of enzymes, one of them inhibited by o-phenanthroline, are present in epimastigotes and metacyclics. None of the reagents tested fully inhibited the phospholipases.
Schenkman
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Department of Microbiology, Immunology and Parasitology, Escola Paulista de Medicina, São Paulo, Brazil.
Yoshida
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N
Cardoso de Almeida
M L
ML
D000818:Animals; D000911:Antibodies, Monoclonal / Q000276:immunology; D003429:Cross Reactions; D000939:Epitopes / Q000276:immunology; D006017:Glycolipids / Q000378:metabolism*; D017261:Glycosylphosphatidylinositols; D007118:Immunoassay; D008565:Membrane Proteins / Q000032:analysis* / Q000378:metabolism; D010716:Phosphatidylinositols / Q000378:metabolism*; D014346:Trypanosoma brucei brucei / Q000201:enzymology; D014349:Trypanosoma cruzi / Q000032:analysis* / Q000276:immunology; D010738:Type C Phospholipases / Q000378:metabolism
D000911:Antibodies, Monoclonal; D000939:Epitopes; D006017:Glycolipids; D017261:Glycosylphosphatidylinositols; D008565:Membrane Proteins; D010716:Phosphatidylinositols; D010738:Type C Phospholipases