Differential biochemical and kinetic properties of α-amylases from Rhyzopertha dominica (F.) progenies reared on wheat varieties differing in α-amylase inhibitory activity

The present study was accomplished to gain insights into the biochemical and kinetic properties of Rhyzopertha dominica's alpha-amylase isoforms (named RdA70, RdA79, and RdA90) expressed in progenies reared in wheat varieties differing in alpha-amylase inhibitory activity. An inverse relationship was observed between the progenies' amylase activity and the wheat inhibitory activity. Wheat samples with a high and low infestation (named HI-Borlaug and LI-Villa Juarez, respectively) were chosen to simplify the study. The progenies amylases were isolated by hydrophobic interaction chromatography, while the wheat samples were analyzed in alpha-amylase inhibitory activity by size exclusion chromatography. The isoforms RdA70 and RdA90 from LI-Villa Juarez progeny showed higher enzyme activities (73.8 and 43.4%, respectively) than those from HI-Borlaug. When the amylase isoforms were tested in susceptibility to inhibition by the inhibitory albumins, those from LI-Villa Juarez were more susceptible than those of the HI-Borlaug. Determination of the kinetic parameters revealed that RdA70 from the HI-Borlaug progeny showed 3.0-fold less starch affinity than that from the LI-Villa Juarez (K-m of 12.3 +/- 1.8 versus 4.0 +/- 0.3). The rest of the alpha-amylases did not show the same pattern as RdA70 in the HI-Bourlag as that their starch affinity was further reduced (RdA79 K-m = 23.1 +/- 4.3, RdA90 K-m = 17.1 +/- 2.9). Estimation of IC50 values confirmed the high sensitivity of the three alpha-amylases of the LI-Villa Juarez progeny to wheat alpha-amylase inhibitors. The inhibitor constant K-i was the lowest for RdA70 in the LI-Villa Juarez progeny indicating the inhibitor' tight binding to that isoenzyme. These results suggest that R. dominica uses RdA70 to bind large amounts of wheat alpha-amylase inhibitors than RdA79 and RdA90 as a physiological defense mechanism. Published by Elsevier Ltd.

Automated summary

The study investigated the biochemical and kinetic properties of alpha-amylase isoforms in Rhyzopertha dominica, expressed in wheat varieties with different alpha-amylase inhibitory activity. Results showed that the isoforms from the LI-Villa Juarez progeny were more sensitive to wheat alpha-amylase inhibitors compared to those from the HI-Borlaug progeny.