IDENTIFICATION OF A DOMAIN OF TRYPANOSOMA-CRUZI METACYCLIC TRYPOMASTIGOTE SURFACE-MOLECULE GP82 REQUIRED FOR ATTACHMENT AND INVASION OF MAMMALIAN-CELLS

Recombinant proteins and synthetic peptides representing various seque nces of gp82, a surface glycoprotein of Trypanosoma cruzi metacyclic t rypomastigotes implicated in mammalian cell invasion, were used in thi s study aiming at the identification of the domain(s) of this molecule required for interaction with target cells. Invasion of cultured HeLa cells by metacyclic trypomastigotes was inhibited by about 80% in the presence of native gp82 or the corresponding recombinant construct J1 8. Inhibition by recombinant proteins J18a and J18b, containing respec tively the N-terminal and the C-terminal portions of gp82, was on the order of 30% and 65%. As compared to J18b (amino acids 224-516), the t runcated gp82 fragments J18b1 (amino acids 303-516) and J18b2 (amino a cids 357-516) displayed lower inhibitory effect (similar to 40% and si milar to 15%, respectively). Compatible with these observations, we fo und that the recombinant protein J18b, but not J18a or J18b2, binds to HeLa cells in a dose-dependent and saturable fashion. Experiments wit h ten overlapping synthetic peptides, representing the gp82 portion sp anning amino acids 224-333, showed that peptides 4 (amino acids 254-27 3) and 8 (amino acids 294-313) have significant inhibitory activity on HeLa cell invasion by metacyclic forms. All these results indicate th at the portion of gp82 required for mammalian cell attachment and inva sion is located in the central domain of the molecule.