Characterization of Two Acanthoscelides obtectus α-Amylases and Their Inactivation by Wheat Inhibitors
- 12 February 2005
- journal article
- research article
- Published by American Chemical Society (ACS) in Journal of Agricultural and Food Chemistry
- Vol. 53 (5) , 1585-1590
Abstract
Wheat α-amylase inhibitors represent an important tool in engineering crop plants against bean bruchids. Because Acanthoscelides obtectus is a devastating storage bean insect-pest, we attempted to purify and characterize its gut α-amylases, to study their interaction with active proteinaceous inhibitors. Two digestives α-amylases (AoA1 and AoA2) were purified from gut larvae, showing molecular masses of 30 and 45 kDa for each one, respectively. The stoichiometry interaction between these 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 rich ammonium 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 the highest concentration (1.0%) reduced the larval weight by 80%. Similar data were observed when 0.53 inhibitor was incorporated at 0.5%. When the concentration of purified 0.53 was enhanced to 1.0%, no larvae or adult emergence were observed. Our data suggest that these α-amylase inhibitors present great potential for use in Phaseolus genetic improvement programs. Keywords: α-Amylase inhibitors; wheat; bruchids; Acanthoscelides obtectus; weevilsKeywords
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