Generation of a membrane-bound, oligomerized pre-pore complex is necessary for pore formation by Clostridium septicum alpha toxin

Abstract

Low-temperature inhibition of the cytolytic activity of alpha toxin has facilitated the identification of an important step in the cytolytic mechanism of this toxin. When alpha toxin-dependent haemolysis was measured on erythrocytes at various temperatures it was clear that at temperatures ≤15°C the haemolysis rate was significantly inhibited with little or no haemolysis occurring at 4°C. Alpha toxin appeared to bind to and oligomerize on erythrocyte membranes with similar kinetics at 4°C and 37°C. The slight differences in these two processes at 4°C and 37°C could not account for the loss of cytolytic activity at low temperature. At 4°C alpha toxin neither stimulated potassium release from erythrocytes nor formed pores in planar membranes. In contrast, at temperatures 25°C both processes proceeded rapidly. Pores that were opened in osmotically stabilized erythrocytes could not be closed by low temperature. Therefore, low temperature appeared to prevent the oligomerized complex from forming a pore in the membrane. These data support the hypothesis that alpha toxin oligomerizes into a membrane-bound, pre-pore complex prior to formation of a pore in a lipid bilayer.