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A kidney bean trypsin inhibitor with an insecticidal potential against Helicoverpa armigera and Spodoptera litura

Mittal, Anuradha, Kansal, Rekha, Kalia, Vinay, Tripathi, Monika, Gupta, Vijay Kumar
Acta physiologiae plantarum 2014 v.36 no.2 pp. 525-539
Helicoverpa armigera, Phaseolus vulgaris, Spodoptera litura, adults, artificial diets, bioassays, bovine serum albumin, calcium chloride, casein, cattle, dissociation, insecticidal properties, insects, instars, kidney beans, larvae, larval development, molecular weight, mortality, pH, polyacrylamide gel electrophoresis, pupation, sucrose, temperature, thermal stability, trypsin
In the present study, trypsin inhibitor extracts of ten kidney bean seed (Phaseolus vulgaris) varieties exhibiting trypsin and gut trypsin-like protease inhibitor activity were tested on Helicoverpa armigera and Spodoptera litura. Trypsin inhibitor protein was isolated and purified using multi-step strategy with a recovery of similar to 15 % and purification fold by similar to 39.4. SDS-PAGE revealed a single band corresponding to molecular mass of similar to 15 kDa and inhibitory activity was confirmed by reverse zymogram analyses. The inhibitor retained its inhibitory activity over a broad range of pH (3-11), temperature (40-60 A degrees C) and thermostability was promoted by casein, CaCl2, BSA and sucrose. The purified inhibitor inhibited bovine trypsin in 1:1 molar ratio. Kinetic studies showed that the protein is a competitive inhibitor with an equilibrium dissociation constant of 1.85 mu M. The purified trypsin inhibitor protein was further incorporated in the artificial diet and fed to second instar larvae. A maximum of 91.7 % inhibition was obtained in H. armigera, while it was moderate in S. litura (29 %) with slight varietal differences. The insect bioassay showed 40 and 22 % decrease in larval growth followed by 3 and 2 days delay in pupation of H. armigera and S. litura, respectively. Some of the adults emerged were deformed and not fully formed. Trypsin inhibitor protein was more effective against H. armigera as it showed 46.7 % mortality during larval growth period compared to S. litura (13.3 %).