In silico identification of Cry1Ac Hot Spot amino acid residues to improve toxicity against mutated cadherin receptor of Bt resistant Helicoverpa armigera

Bacillus thuringiensis Cry proteins are used for biological control of insect pests; however, insect pests are developing resistance against these proteins especially Cry1Ac. We hypothesized that the problem of resistance development can be overcome by mutating hot spot amino acid residues of Cry1Ac protein conferring enhanced toxicity against resistant insects. Cadherin protein region CD7-CD8 is known to act as primary insect midgut receptors involved in Cry1Ac binding. Amino acid mutations in this region may evolve resistance against Bt toxins. Here we have identified three key amino acid residues A1264, H1436 and L1461 of cadherin receptor protein from Helicoverpa armigera genome and in silico protein -protein interaction studies revealed their role in insect resistance against Cry1Ac. Three mutations viz A1264P, H1436L and L1461V showed significantly high increment increment G values as 9.3, 6.0 and 5.9 respectively indicating destabilization of cadherin protein which reduced its binding with Cry1Ac resulting in resistance development. Further, molecular docking of these mutated amino acid residues revealed lack of interaction with amino acid residues of Cry1Ac viz Q509, Y513, W544, N547 and I585 essential for cadherin-Cry1Ac binding in susceptible insects. In second part of our study, we identified two hot spot amino acid residues of Cry1Ac viz S548, I586 whose mutation viz S548H or S548W and I586Y brought about strong interaction with midgut receptors of resistant insects having mutated cadherin. Based on these results, we sugest, reported Cry1Ac hot spot amino acids if mutated can help to overcome resistance mechanism. Here we laid a foundation for further experiments to modify the Cry1Ac hot spot residues which bind with the resistant receptor binding protein (cadherin) more strongly to perform efficient insecticidal activity against resistant strains of Helicoverpa armigera.