Characterization of a membrane pore-forming protein from Entamoeba histolytica.

Abstract

We describe the partial purification and characterization of a pore-forming material (PEM) from Entamoeba histolytica. The formation of ion channels by PFM was examined in three systems. (a) PFM depolarizes J774 macrophages and mouse spleen lymphocytes as measured by [3H]TPP+ uptake. (b) PFM induces rapid monovalent cation flux across the membrane of phosphatidylcholine-cholesterol vesicles. (c) PFM confers a voltage-dependent conductance to artificial planar bilayers, which is resolved as a summation of opening of individually conducting steps of 67 pS in 0.1 M KCl. Monomers of PFM are functional; however, a preferential aggregation occurs in the planar bilayer. Activity is pronase, trypsin, and heat sensitive and is stable between pH 5-8. PFM is not secreted by unstimulated amoebae but after exposure to the calcium ionophore A23187, concanavalin A, and E. coli lipopolysaccharide, 5-10% of the total cell content of PFM is released into the medium within 5-10 min. High-performance gel filtration results in an approximately 1,000-fold purification of PFM and gives an Mr of 30,000. This protein may play a role in the cytotoxicity mediated by E. histolytica.

Properties of a purified pore-forming protein (perforin 1) isolated from H-2-restricted cytotoxic T cell granules.

“…Pore-forming activity has been described for a number of bacterial toxins and antibiotics (33,34). A PFP/P1 from Entamoeba histolytica (22,35,36), which may be responsible for the contact-dependent cytolysis mediated by that protozoan, has also been extracted. It is interesting that the 70-75 kD protein described here bears close structural and functional homology to the ninth component of complement (C9); C9 polymerizes spontaneously at 37°C to form structural and functional transmembrane tubules similar to the ones described here; moreover, C9 and PFP/P 1 show immunological crossreactivity (Young, J., z. Cohn, and E. Podack, manuscript submitted for publication).…”

“…Planar bilayers were formed in indicated buffers according to previously published protocols (22,23 (25,26). The bilayers were formed in buffer A.…”

Identificación de antígenos de aislamientos colombianos de Giardia duodenalis reconocidos por IgG total y subclases.

“…Aún más, se conoce que E. histolytica puede ser invasivo en el hospedero y G. duodenalis puede estar en contacto con el huésped en el intestino delgado. Por ello, son parásitos intestinales que pueden inducir respuesta humoral en el hospedero (19,20). En cambio E. coli, E. nana, I. butschlii, T. hominis, uncinarias y B. hominis no inducen respuesta inmune y, por ende, ninguna producción de anticuerpos (21).…”

Trophozoites ofEntamoeba histolyticaepigenetically silenced in several genes are virulence-attenuated

“…D iscovered in 1982 (Lynch et al, 1982;Young et al, 1982), the amoebapores are membraneinteracting proteins which display pore-forming activity toward liposomes (Leippe et al, 1991). They have antibacterial activity (Leippe et al, 1994a) and are cytotoxic to human cell lines in vitro (Leippe et al, 1994b).…”