Processing of delta-endotoxin of Bacillus thuringiensis subsp. kurstaki HD-1 in Heliothis armigera midgut juice and the effects of protease inhibitors.
Bombyx mori was found to be more sensitive to the protoxins of HD-1 than Heliothis armigera. SDS-PAGE analysis showed that a large amount of activated toxin was yielded from protoxin by B. mori gut juice while little was yielded by H. armigera. Further degradation of activated toxin was observed in H. armigera midgut juice detected by SDS-PAGE. pH influenced the proteolytic activity of the midgut juice significantly, but there was no obvious effect of pH on the degradation of activated toxin. Specific inhibitor study revealed the presence of trypsin, chymotrypsin, and elastase in the midgut juice. TLCK, TPCK, elastatinal and some general serine protease inhibitors successfully prevented the excessive degradation of protoxin in H. armigera midgut juice. Chymotrypsin inhibitors showed strong inhibitory effects against the further degradation of activated toxin, indicating that chymotrypsin played a major role in the process. It was presumed that the excessive degradation of protoxin in H. armigera midgut juice was responsible for the low sensitivity of the insect to Bt. Further study demonstrated that the excessive degradation in vitro was triggered by SDS treatment. However, all of the tested serine protease inhibitors expressed synergism with protoxin against H. armigera larvae, suggesting that the excessive degradation of protoxin may occur in vivo to some extent and may be triggered by receptor binding of activated toxin.
Shao
Z
Z
Institute of Crop Genetics & Breeding, Shandong Agricultural University, Taian, Shandong, 271018, China.
Cui
Y
Y
Liu
X
X
Yi
H
H
Ji
J
J
Yu
Z
Z
D000818:Animals; D001413:Bacillus thuringiensis / Q000378:metabolism*; D000083722:Bacillus thuringiensis Toxins; D001426:Bacterial Proteins / Q000378:metabolism*; D001427:Bacterial Toxins / Q000378:metabolism; D004064:Digestive System; D004731:Endotoxins / Q000378:metabolism*; D006460:Hemolysin Proteins; D006863:Hydrogen-Ion Concentration; D009036:Moths / Q000378:metabolism* / Q000382:microbiology*; D011480:Protease Inhibitors; D011498:Protein Precursors / Q000378:metabolism; D012967:Sodium Dodecyl Sulfate
D000083722:Bacillus thuringiensis Toxins; C029103:Bacillus thuringiensis protoxin; D001426:Bacterial Proteins; D001427:Bacterial Toxins; D004731:Endotoxins; D006460:Hemolysin Proteins; D011480:Protease Inhibitors; D011498:Protein Precursors; C030956:insecticidal crystal protein, Bacillus Thuringiensis; D012967:Sodium Dodecyl Sulfate