The inhibitory effects of the cysteine protease inhibitor, oryzacystatin, on digestive proteases and on larval survival and development of the southern corn rootworm (Diabrotica undecimpunctata howardi)

At least eight proteolytic activities have been identified in the midgut contents of larval Southern corn rootworm (Diabrotica undecimpunctata howardi). Around 70% of protease activity could be arrested by the cysteine protease inhibitors E-64 and chicken egg-white cystatin, while the aspartic acid protease inhibitor pepstatin caused 30% inhibition. The cysteine protease activity was found to be highly sensitive to inhibition by both chicken egg-white cystatin and the rice cystatin, oryzacystatin I. Oryzacystatin I, expressed as a fully functional fusion protein in E. coli, was found to strongly inhibit larval gut protease activity. This recombinant oryzacystatin, incorporated into artificial diet at concentrations of 10 mM and above, caused significant decreases in larval survival and weight gain. E-64 was also shown to cause a significant antimetabolic in vivo effect. These results demonstrate the great potential for cysteine protease inhibitors, such as oryzacystatin, as tools for exploitation in the control of the Southern corn rootworm.