DELTA-ENDOTOXIN-INDUCED LEAKAGE OF RB-86+-K+ AND H2O FROM PHOSPHOLIPID-VESICLES IS CATALYZED BY RECONSTITUTED MIDGUT MEMBRANE

Brush border membrane from Heliothis virescens catalyzed delta-endotoxin-induced leakage of Rb-86+-K+ and H2O from phospholipid vesicles. Activated delta-endotoxin [CrylA(c)-55 kDa] from Bacillus thuringiensis kurstaki strain EG2244 producing a single CrylA(c) toxin, when incorporated into phospholipid vesicles, made these vesicles more leaky to Rb-86+-K+ than phospholipid vesicles without toxin. This effect was assayed by following the movement of Rb-86+ into the vesicles in response to a KCl gradient. When toxin was added to the outside of phospholipid vesicles, Rb-86+ uptake was impeded. Vesicles prepared with H. virescens brush border membrane catalyzed the association of the toxin with the vesicle, and stimulated KCl gradient-induced Rb-86+ uptake. Toxin did not catalyze the leakage of Cl-36-, suggesting that the toxin created a cation-selective leak. Toxin enhanced the permeability of phospholipid vesicles to H2O, demonstrated by the enhanced rate of vesicle shrinking under increased osmotic pressure. This was analyzed spectrophotometrically by following the rate of vesicle shrinking in response to a 10 mM KCl gradient. In the presence of concentrated phosphatidylcholine vesicles, toxin spontaneously associated with the vesicles so as to enhance the rate of vesicle shrinking in an osmotic gradient. The rate of vesicle shrinking the presence of KCl and toxin was catalyzed by the presence of brush border reconstituted into the vesicles, reducing the effective toxin concentration 1000-fold.