title: Influence of mutagenesis of Bacillus thuringiensis Cry1Aa toxin on larvicidal activity.
abstract: Domain III of Bacillus thuringiensis Cry δ-endotoxins are considered to be related to the stability of the structure and avoidance of overdigestion by proteases. In this study, some residues of potential chymotrypsin and trypsin sites in Domain III of B. thuringiensis Cry1Aa were replaced individually with alanine by site-directed mutagenesis, in order to investigate their functional roles. Except F574A, all mutants F536A, R543A, F550A, F565A, R566A, F570A, F576A, F583A, and F590A were highly expressed the 130 kD protoxins at levels comparable to the wild-type tested by SDS-PAGE. In bioassays, F536A, R566A, and F590A increased toxicity against Spodoptera exigua Hüner larve by 20, 40, and 40%, respectively, as compared to the wild-type. F536A and F565A showed an increase of 6 and 10% in toxicity against Heliothis armigera Hubner than the wild-type. Toxicities of some mutants were altered greatly, and the same mutants were shown to have different toxicities against those two insects. Structural analyses showed that mutants R543A, F574A, F576A-affecting insecticidal activity might be relational to structural stability of toxin or decreased affinity for receptor binding. These results indicated that those residues were involved in the larvicidal activity of the Cry1Aa toxin.
authors.affiliation: Key Lab of Microbial Molecular Biology of Hunan Province, College of Life Science, Hunan Normal University, Changsha, 410081, People's Republic of China.
authors.forename: Chunyan
authors.initials: C
authors.lastname: Zhang
authors.forename: Liqiu
authors.initials: L
authors.lastname: Xia
authors.forename: Xuezhi
authors.initials: X
authors.lastname: Ding
authors.forename: Fan
authors.initials: F
authors.lastname: Huang
authors.forename: Huanfa
authors.initials: H
authors.lastname: Li
authors.forename: Yunjun
authors.initials: Y
authors.lastname: Sun
authors.forename: Jia
authors.initials: J
authors.lastname: Yin
mesh_terms: D019943:Amino Acid Substitution / Q000235:genetics; D000818:Animals; D000083722:Bacillus thuringiensis Toxins; D001426:Bacterial Proteins / Q000235:genetics* / Q000633:toxicity*; D004591:Electrophoresis, Polyacrylamide Gel; D004731:Endotoxins / Q000235:genetics* / Q000633:toxicity*; D015870:Gene Expression; D006460:Hemolysin Proteins / Q000235:genetics* / Q000633:toxicity*; D007306:Insecticides / Q000633:toxicity*; D007814:Larva / Q000187:drug effects; D007915:Lepidoptera / Q000187:drug effects*; D016297:Mutagenesis, Site-Directed; D020125:Mutation, Missense; D016019:Survival Analysis
chemical_list: D000083722:Bacillus thuringiensis Toxins; D001426:Bacterial Proteins; D004731:Endotoxins; D006460:Hemolysin Proteins; D007306:Insecticides; C030956:insecticidal crystal protein, Bacillus Thuringiensis