Characterization of a membrane pore-forming protein from Entamoeba histolytica.
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.
D000818:Animals; D001667:Binding, Competitive; D002463:Cell Membrane Permeability / Q000187:drug effects*; D004748:Entamoeba histolytica / Q000032:analysis* / Q000378:metabolism; D006863:Hydrogen-Ion Concentration; D007473:Ion Channels / Q000187:drug effects; D046911:Macromolecular Substances; D008564:Membrane Potentials / Q000187:drug effects; D008565:Membrane Proteins / Q000032:analysis / Q000302:isolation & purification / Q000494:pharmacology*; D008567:Membranes, Artificial; D009861:Onium Compounds / Q000494:pharmacology; D009943:Organophosphorus Compounds; D011402:Pronase / Q000494:pharmacology
D007473:Ion Channels; D046911:Macromolecular Substances; D008565:Membrane Proteins; D008567:Membranes, Artificial; D009861:Onium Compounds; D009943:Organophosphorus Compounds; D011402:Pronase; C013289:tetraphenylphosphonium