Recombinant scorpion insect excitatory toxin BmK IT accelerates the growth of insect Spodoptera frugiperda 9 cells
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Several scorpion insect toxins are selectively active on the lepidopterous and dipterous insects. The gene encoding insect excitatory neurotoxin (BmK IT) from the scorpion Buthus martensii Karsch was expressed in Escherichia coli BL21(DE3) at a high level of 3 mg/0.5 L using the prokaryotic expression system pTWIN1. Colorimetric 3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyl tetrazolium bromide (MTT), whole-cell patch-clamp technique and immunofluorescence assays were used to evaluate the toxicity of rBmK IT to insect Spodoptera frugiperda 9 (Sf9) cells and to analyze the potential mechanism of this toxicity. rBmK IT accelerated the growth of Sf9 cells in a dose-dependent manner. Voltage-gating sodium channel activity could not be detected in Sf9 cells using a whole-cell patch-clamp technique. However, immunofluorescence analysis clearly showed co-localization of tetrodotoxin (TTX) and rBmK IT on the Sf9 cell membrane, which demonstrated that rBmK IT could bind to and act on the voltage-gated sodium channels on the Sf9 cells by the high affinity action power. The findings presented in this study are essential for further study of this peptide.
KeywordsButhus martensii Karsch insect toxin (BmK IT) Toxicity Spodoptera frugiperda 9 cell Sodium channel
We would like to thank Dr. Spyridon Theofilopoulos (Karolinska Institute, Sweden), for critical comments and linguistic styling of the manuscript. This project was supported by grants from “National Natural Science Foundation of China” (No. 30700534 and No. 30901774), “Natural Science Foundation of Shanxi Province” (2008021039), “Medical Science and Technique Foundation of Shanxi Province (200617)” and the “National High Technology Research and Development Program of China (863 Program)”.
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