Inhibitory effect of Ni2+ on the tolaasin-induced hemolysis

Abstract

The bacterial toxin, tolaasin, causes brown blotch disease on the cultivated mushrooms by collapsing fungal and fruiting body structure of mushroom. Cytotoxicity of tolaasin was evaluated by measuring hemolytic activity because tolaasins form membrane pores on the red blood cells and destroy cell structure. While we investigated the inhibitions of hemolytic activity of tolaasin by Zn2+and Cd2+, we found that Ni2+ is another antagonist to block the toxicity of tolaasin. Ni2+ inhibited the tolaasin-induced hemolysis in a dose-dependent manner and its Ki value was ~10 mM, implying that the inhibitory effect of Ni2+ is stronger than that of Cd2+. The hemolytic activity was completely inhibited by Ni2+ at the concentration higher than 50 mM. The effect of Ni2+ was reversible since it was removed by the addition of EDTA. When the tolaasin-induced hemolysis was suppressed by the addition of 20 mM Ni2+, the subsequent addition of EDTA immediately initiated the hemolysis. Although the mechanism of Ni2+-induced inhibition on tolaasin toxicity is not known, Ni2+ could inhibit any of following processes of tolaasin action, membrane binding, molecular multimerization, pore formation, and massive ion transport through the membrane pore. Our results indicate that Ni2+ inhibits the pore activity of tolaasin, the last step of the toxic process.