Site-directed mutagenesis in Cry proteins of Bacillusthuringiensis to demonstrate the role of domain II and domain III in toxicity enhancement toward Spodopteralitura

Egyptian Journal of Biological Pest Control

Research

Fakhar-un-Nisa Yunus¹  Huma Khurshid¹  Farzana Rashid¹  Farkhanda Manzoor¹  Aasma Abdul Latif¹  Hafsa Zaheer¹ 

[1] Department of Zoology, Lahore College for Women University, 54000-Jail Road, Lahore, Pakistan;

关键词: Bacillus;    Overlap extension PCR;    Site-directed mutagenesis;    Spodoptera;   

DOI  :  10.1186/s41938-023-00731-x

 received in 2023-06-05, accepted in 2023-08-22,  发布年份 2023

来源: Springer

【 摘 要 】

BackgroundBacillusthuringiensis (Bt) is a gram-positive bacterium responsible for the production of a wide variety of insecticidal Cry, Cyt, and Vip proteins with distinct insect specificities. The bioinsecticides derived from Bt Cry proteins account for > 95% of the microbial biopesticide market to combat a variety of pest species belonging to the order Lepidoptera (including Spodoptera spp.), Coleoptera, Diptera, etc. Cry proteins are engineered by using different molecular techniques to control the development of multiple insecticide resistance problems in major insect pests using bio-toxicity assays.Main bodyIt is common knowledge that the Cry proteins domain II and III are involved in pore formation or interaction between several insect larval receptors and the membranes of epithelial cells. In the present research, the PCR site-directed mutagenesis technique was used to introduce a total of four mutations into the cry genes (cry1 and cry2) near key regions of active proteins. The diet overlay bioassay was used to test the efficacy of expressed mutant Cry proteins against Spodopteralitura (Fabricius) (Lepidoptera: Noctuidae), one of the most damaging bollworms to cotton, causing severe output losses.ResultsTwo amino acid replacements in the receptor binding domain of Cry1Ac toxins (S573, L588) showed decrease in activity for Cry1Ac mutants. However, amino acid substitutions in the receptor binding/pore formation domain of Cry2Aa (T325, S445) were to some extent proved more toxic than wild-type recombinant Cry2Aa protein, with an increase in mortality percentage from 3.33 to 6.66% after 24 to 72 h of treatment, respectively, against 2nd instar larvae of S.litura.ConclusionA comparison of activity demonstrated that larvae of S.litura were more susceptible to Cry2Aa toxins than those of Cry1Ac toxins after being treated with Bt toxins. The LC50 values of Cry2Aa mutants were slightly reduced for S.litura larvae than those of Cry1Ac mutants.

【 授权许可】

CC BY   
© Egyptian Society for Biological Control of Pests 2023

【 图 表 】

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