title: A protein engineering of Bacillus thuringiensis δ-endotoxin by conjugating with 4"-O-succinoyl abamectin.
abstract: Conjugation of Bacillus thuringiensis δ-endotoxin (Bt toxin) with other toxins for insect pest control has been proposed as a new efficient strategy with increasing insecticidal toxicity and target range and delay the onset of insect resistance. A modified method was investigated by conjugating Bt toxin with 4"-O-succinoyl abamectin to form a new biocide which was named as BtA. 'Zero-length' cross-linker EDC in combination with NHS activated 4"-O-succinoyl abamectin and extended half-life period of active intermediate for binding to Bt toxin. The dissociation constant for 4"-O-succinoyl abamectin binding to Bt toxin was 6.44 μM by fluorescence quenching analysis. BtA showed a higher insecticidal toxicity against Plutella xylostella, while the relative-toxicity multiple of BtA to Bt toxin was calculated as 5.6. The interaction between Bt toxins with their receptors played a key role in toxicity of Bt toxins. The binding analysis showed the dissociation rate for the binding of BtA to its receptors (7.495 × 10(-3) S(-1)) was twice slower than that of Bt toxin (1.695 × 10(-2) S(-1)). The relative dissociation constant of BtA to Bt toxin was only 29% for the binding to the receptors. These results demonstrated that BtA bound to the receptor in BBMV with significantly higher affinity compared with Bt toxin.
authors.affiliation: Agricultural Bio-Resources Institute, Fujian Academy of Agricultural Sciences, Fuzhou 350003, China.
authors.forename: Zhi-Zhen
authors.initials: ZZ
authors.lastname: Pan
authors.forename: Yu-Jing
authors.initials: YJ
authors.lastname: Zhu
authors.forename: Zheng
authors.initials: Z
authors.lastname: Chen
authors.forename: Chuan-Qing
authors.initials: CQ
authors.lastname: Ruan
authors.forename: Lian
authors.initials: L
authors.lastname: Xu
authors.forename: Qing-Xi
authors.initials: QX
authors.lastname: Chen
authors.forename: Bo
authors.initials: B
authors.lastname: Liu
mesh_terms: D000818:Animals; D001413:Bacillus thuringiensis; D001681:Biological Assay; D015195:Drug Design; D004731:Endotoxins / Q000737:chemistry* / Q000378:metabolism; D007306:Insecticides / Q000737:chemistry* / Q000378:metabolism; D007559:Ivermectin / Q000031:analogs & derivatives* / Q000737:chemistry; D007915:Lepidoptera / Q000166:cytology; D008871:Microvilli / Q000378:metabolism; D011956:Receptors, Cell Surface / Q000378:metabolism
chemical_list: D004731:Endotoxins; D007306:Insecticides; D011956:Receptors, Cell Surface; C048324:abamectin; D007559:Ivermectin
keywords: 1-ethyl-3-(3-dimethylaminopropyl) carbodiimide hydrochloride; BBMV; Bacillus thuringiensis; Bt; BtA; DMF; EDC; EDTA; EGTA; FQA; Fluorescence quenching analysis (FQA); Forte-Bio Octet Red System (Forte-Bio ORS); K(d); LB; Luria-Bertani; N-hydroxysuccinimide; NHS; ORS; Octet Red System; brush border membrane vesicles; dimethylformamide; dissociation rate; ethylene diamine tetraacetic acid; ethylene glycol-bis(2-aminoethylether)-N,N,N′,N′-tetraacetic acid; fluorescence quenching analysis