| Abstract
| - Wheat α-amylase inhibitors represent an important tool in engineering crop plants against beanbruchids. Because Acanthoscelides obtectus is a devastating storage bean insect-pest, we attemptedto purify and characterize its gut α-amylases, to study their interaction with active proteinaceousinhibitors. Two digestives α-amylases (AoA1 and AoA2) were purified from gut larvae, showingmolecular masses of 30 and 45 kDa for each one, respectively. The stoichiometry interaction betweenthese α-amylases with two wheat inhibitors (0.19 and 0.53) showed a binding complex of 1:1 enzyme:inhibitor. In vivo activities of these inhibitors against A. obtectus were also evaluated using a richammonium sulfate inhibitor fraction (F20-40) and purified inhibitors after reversed phase high-performance liquid chromatography columns. Incorporation of three different inhibitor concentrations(0.25, 0.5, and 1.0% w/w) into artificial seeds showed that addition of the purified 0.19 inhibitor at thehighest concentration (1.0%) reduced the larval weight by 80%. Similar data were observed when0.53 inhibitor was incorporated at 0.5%. When the concentration of purified 0.53 was enhanced to1.0%, no larvae or adult emergence were observed. Our data suggest that these α-amylase inhibitorspresent great potential for use in Phaseolus genetic improvement programs. Keywords: α-Amylase inhibitors; wheat; bruchids; Acanthoscelides obtectus; weevils
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