Insecticidal 6-endotoxin proteins

CjjHsjNjO, , Mr 701.49, [a]g +30.9° (HjO), a thermostable nucleotide toxin from cultures of Bacillus thuringiensis van gelechiae. In contrast to the Thurin-giensis endotoxins (proteins), T. are only formed and excreted by certain serotypes. The insecticidal and cytotoxic activity is based on the inhibition of DNA-de-pendent RNA polymerase. 

Commercial preparations of B.t.. obtained through conventional fermentation techniques, have been used for more than two decades as biological insecticides (I). They exhibit desirable properties such as high insect toxicity and environmental safety. B.t. does not affect non-target insects and is completely nontoxic to vertebrates. Nevertheless its use has been limited due to high production costs, limited stability in field conditions and a too narrow insecticidal spectrum. The insecticidal activity of B.t. relies in the crystalline inclusions which are produced upon sporulation. The crystals contain insecticidal proteins, delta-endotoxins, which affect the midgut epithelium of sensitive insects. The exact mechanism of their toxic activity is still unknown. 

Mode of Action. Despite the wide spectrum of insect host range activities displayed by Bt varieties, the mode of action of the insecticidal delta endotoxins is similar. When susceptible insects ingest Bt protein crystals, the first gross symptom observed is feeding inhibition due to paralysis of the digestive tract, including mouth parts. This usually occurs within one hour of ingestion ( ). 

As mentioned, the 8-endotoxin (Cry protein) that makes up most of the conspicuous crystals is the main insecticidal component of B. thuringiensis. At sporulation, the majority of Bt strains produce crystalline inclusions that contain this insecticidal 8-endotoxin. The crystals account for 20% or more of the total bacterial protein at sporulation and may contain one or several endotoxins, which differ in activity. Many Bt toxin genes and genes for some... 

The 6-endotoxin of B. . israelensis (H-14) is not as clearly understood as that of BTk. The parasporal crystal of BTi contains several proteins with molecular weights ranging from 26,000 to over 100,000d (9). Several authors have reported that the 26,000d protein has insecticidal activity (j , 11, 12). However, other reports suggest that the toxin is 66,000d, (13, 1 ). Because BTi has hemolytic and cytolytic activities in addition to insecticidal activities (15) the possibility exists that multiple toxin proteins exist in the crystal. However, evidence from genetics... 

The 6-endotoxin gene from BTk was first cloned by recombinant WJA techniques in 1961 ( 6). Since then, the toxin genes have been cloned from a variety of B.t. strains and from B. stAiaericus. A list of these cloned genes is in Table I. The isolation of these genes enables both the analysis and manipulation necessary to alter or inprove the insecticidal proteins. 

For example, in the case of Bt-corn, a gene from a naturally occurring soil bacterium. Bacillus thuring-ensis, was inserted into corn to provide resistance to the corn borer. The gene from the bacterium produces a protein, Bt delta endotoxin, which kills the European and southwestern com borer larvae. Bt-com eliminates the need to spray insecticides to control com borers. Although planting these crops reduces the amount of pesticides released into the environment, the long-term effects of Bt-com on human health and the environment are not known. 

Binding of activated toxins to a specific binding site(s) in the brush border membrane of midgut epithelial cells is thought to be a major determinant of the specificity of Cry insecticidal proteins 9-11). In the last few years, several toxin-binding proteins have been purified, and genes encoding them have been cloned. However, in the case of dipteran-specific 8-endotoxins, no receptor protein has been found so far 12, 13). 

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