Membrane glycoprotein receptor and hole-forming properties of a cytolytic protein toxin

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Abstract
Aerolysin, a cytolytic bacterial exotoxin, was radioiodinated by using the Iodogen reagent. Binding of the labeled toxin to rat erythrocytes was inhibited by the native protein and by anti-aerolysin antibody. Toxin, once bound, was not removed by the addition of a large excess of free aerolysin. Binding of the radioactive toxin to erythrocytes of different species paralleled the hemolytic specificity of the unlabeled toxin. Pretreatment of the rat erythrocytes with trypsin, which removed a major membrane glycoprotein, resulted in a dramatic decrease in binding, whereas chymotrypsin treatment had no effect. Binding was inhibited by a glycoprotein fraction isolated from these cells but not by a total rat erythrocyte glycolipid preparation. Aerolysin caused the formation of holes in erythrocytes which were sized by measuring the release of labeled MW markers. Glucagon (MW 3550) and smaller molecules entrapped in human or rat erythrocytes were released by treatment with aerolysin, whereas methoxyinulin (MW 5500) and larger molecules were not. Aerolysin also caused the release of glucose from large unilamellar lipid vesicles. Thus, a specific glycoprotein receptor evidently facilitates the interaction of aerolysin with erythrocyte membranes. Binding is followed by the formation of discrete holes or pores; this results in cell rupture by a colloid-osmotic process.