STUDIES ON THE MODE OF ACTION OF CHOLESTEROL OXIDASE ON INSECT MIDGUTMEMBRANES

Cholesterol oxidase (EC 1.1.3.6.) is an insecticidal protein known to have potent activity against the bell weevil, milder activity against a number of lepidopteran species, and virtually no activity against ot her insects. Several factors that could explain its species-dependent differential activity were examined. We compared cholesterol concentra tions and rates of cholesterol oxidation in the midgut membranes from larvae of bell weevil (Anthonomus grandis grandis Boheman), southern c orn rootworm (Diabrotica undecimpunctata howardi Barber), tobacco budw orm (Heliothis virescens Fabricius), and yellow mealworm (Tenebrio mol itor Linnaeus). Results showed that cholesterol concentration alone co uld not account for the differences in insecticidal activity and that midgut brush-border membranes of all species tested were generally sus ceptible to oxidation by cholesterol oxidase in vitro. We also demonst rated that cholesterol oxidase stability in the midgut environment was similar for the species tested and thus could not account for the dif ferential activity. However, comparison of the pH of the insect midgut fluids with the pH optimum of cholesterol oxidase indicated that the lower sensitivity of lepidopteran larvae to the enzyme may be partiall y due to the alkaline nature of their midgut environments. In some spe cies, oxidation caused significant changes in the activities of brush- border membrane alkaline phosphatase, and these changes did correlate with the susceptibility of the insect to cholesterol oxidase. (C) 1997 Wiley-Liss, Inc.