Biochemical and molecular characterization of delta-endotoxins in Bacillus thuringiensis.
The delta-endotoxins (delta-ETX) of four native strains (RT7, RT19, RT25, and RT25), and one reference strain (4L1) of Bacillus thuringiensis were biochemically and molecularly characterized to determine their potential toxic activity against lepidopteran larvae. Crystals of delta-ETX were purified through a two-phase system to determine their morphology, molar mass, solubility, and resistance to proteinases. Toxic activity and cry gene content were also determined. Crystals from native strains exhibited polyhedral, irregular and cuboidal shapes, while those from 4L1 were bipyramidal. Seven proteins with estimated molar mass approximately 30-134 kDa were detected as the main components of the native delta-ETX. Only crystals from 4L1, RT24, and RT25 underwent complete solubilization at pH >12.0. Crystals from all strains produced trypsin-resistant peptides. None of the cry genes associated with toxicity in lepidopterans (cry1, cry2, cry9) was found in the native strains; however, 4L1 strain harbors cry1 and cry2 genes. Strains RT19 and RT25 caused significant mortality against Trichoplusia ni larvae with partial solubilization at pH 10, strain 4L1 caused 100 % mortality. Toxicity of native strains may come from a novel cry gene.
Laboratorio de Biotecnología Ambiental, Centro de Biotecnología Genómica-Instituto Politécnico Nacional, Reynosa, Tamp., México. nrosas@ipn.mx
N M
NM
Rosas-García
A
A
Sánchez-Varela
J M
JM
Villegas-Mendoza
D000818:Animals; D001413:Bacillus thuringiensis / Q000502:physiology*; D000083722:Bacillus thuringiensis Toxins; D001426:Bacterial Proteins / Q000737:chemistry* / Q000302:isolation & purification / Q000633:toxicity*; D004731:Endotoxins / Q000737:chemistry* / Q000302:isolation & purification / Q000633:toxicity*; D006460:Hemolysin Proteins / Q000737:chemistry* / Q000302:isolation & purification / Q000633:toxicity*; D007814:Larva / Q000187:drug effects; D007915:Lepidoptera / Q000187:drug effects*; D008970:Molecular Weight; D010447:Peptide Hydrolases / Q000378:metabolism; D012995:Solubility; D016019:Survival Analysis
D000083722:Bacillus thuringiensis Toxins; D001426:Bacterial Proteins; D004731:Endotoxins; D006460:Hemolysin Proteins; C030956:insecticidal crystal protein, Bacillus Thuringiensis; D010447:Peptide Hydrolases