Synthesis,Expression and Purification of a Type of Chlorotoxin-like Peptide from the Scorpion, <Emphasis Type="Italic">Buthus martensii</Emphasis> Karsch,and its Acute Toxicity Analysis |
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Authors: | Yue-jun?Fu Li-tian?Yin Wei?Wang Bao-feng?Chai Email author" target="_blank">Ai-hua?LiangEmail author |
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Institution: | (1) Key Laboratory of Chemical Biology and Molecular Engineering of Ministry of Education, Institute of Biotechnology, Shanxi University, 030006 Taiyuan, People’s Republic of China |
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Abstract: | A gene, rBmK Cta, encoding a chlorotoxin-like peptide from the scorpion, Buthus martensii Karsch, was synthesized according to the sequence optimized for codon usage in Escherichia coli and was expressed in E.␣coli BL21 (DE3) using a pExSecI expression system in which the IgG-binding domain-ZZ of protein A is fused to the N-terminal of rBmK CTa. The fusion protein, ZZ-rBmK CTa, was expressed in soluble form (7.8 mg l−1) and was purified to give a single band on SDS-PAGE. The domain-ZZ of fusion protein ZZ-rBmK CTa was removed by cleavage
of an Asn–Gly peptide bond with hydroxylamine. The rBmK CTa was separated from the IgG-binding moiety by a second passage
through the IgG affinity column. Western blot analysis demonstrated that this protein was rBmK CTa. Acute toxicity assay in
mice demonstrated that the rBmK CTa had an LD50 value of 4.3 mg kg−1. |
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Keywords: | acute toxicity analysis Buthus martensii chlorotoxin-like peptide prokaryotic secretive expression |
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