Inhibitory activity of proteinaceous α-amylase inhibitors from Triticale seeds against Eurygaster integriceps salivary α-amylases: Interaction of the inhibitors and the insect digestive enzymes

Abstract

In many countries, the Sunn pest, Eurygaster integriceps Puton (Hemiptera: Scutelleridae) is one of the most important insect pests of wheat. Chemical control of E. integriceps is not efficient making it essential to find other control methods. Plant α-amylase inhibitors expressed as transgenes have emerged as a promising way of controlling insect pests and several crop species expressing transgenic amylase inhibitors have been shown to be resistant to certain insect pest. Here we report inhibition of salivary α-amylases of E. integriceps by semi-purified proteinaceous α-amylase inhibitors from Triticale (TAI). A dose dependent trend of inhibition of the enzyme was observed using TAI with an I50 value of 0.79μg of TAI. At the highest concentration of the inhibitor used here, we found 87% inhibition of amylase activity. The inhibitory activity was maximal at pH at 5.0, which is in accordance with the pH optimum of the salivary gland enzymes. Kinetic studies of enzyme-inhibitor interaction revealed a kind of partial mixed inhibition with an apparent inhibitory constant value (Ki[app]) of 1.1μg of TAI against salivary α-amylase in the assay medium. In gel inhibition assays showed that all isoforms of salivary glands α-amylase were sensitive to the TAI, although to different degrees. Moreover, the effectiveness of TAI was retained in different stages of pre-oral and oral digestion of E. integriceps, although some inhibitory activity was lost, possibly by the action of the insect proteases. These findings indicate that TAI shows promise for use in the management of this pest. To the best of our knowledge, this is the first study of inhibitor-insect digestive enzymes interaction, based on the insect feeding strategy and digestion process.