A maize hydrolase with activity against maize insect and fungal pests

Abstract A gene annotated as a chitinase-like protein that occurred in a quantitative trait locus for fungal maize ear rot resistance was cloned from a maize inbred with Fusarium proliferatum/verticillioides resistance. The protein sequence was similar to type III chitinases and homologous proteins without chitinase activity generally referred to as narbonins or chitolectins. The gene was expressed in Pichia pastoris, and the resulting protein was purified and examined for different types of activity. The yeast-produced protein did not hydrolyze chitin substrates, or 4-nitrophenyl sugars, but did hydrolyze 4-nitrophenyl acetate and 4-nitrophenyl palmitate. The protein also significantly retarded the growth of first instar fall armyworms (Spodptera fruigiperda) and significantly enhanced the activity of miconazole nitrate toward F. graminearum. The gene was biolistically introduced into maize BMS callus and the transgene was detectible by PCR. Some transformant lines significantly reduced corn earworm (Helicoverpa zea), fall armyworm, F. graminearum, F. proliferaturm and F. verticillioides growth compared to GUS transformed control callus. When transformed callus protein extracts were separated by native polyacrylamide gel electrophoresis, a band of 4-nitrophenyl acetate activity was found in bioactive callus that was not present in GUS control callus. Although the true target of the hydrolase is not known, it does appear to play a role in resistance to pests of maize and appears to be a previously unreported type of resistance mechanism.