Bacterial Expression and Purification of an Active ω-Atracotoxin-Arl b from Spider Atrax robustus

• Published in: Agricultural sciences in China no. 9; pp. 1292 - 1298
• Main Author: WANG Yu-qin WANG Jue LI Jian LIU Qiang HUANG Shi-di WANG Dun
• Format: Journal Article
• Language: English
• Published: 2010
• Subjects: 大肠杆菌; 硫氧还蛋白; 神经毒素; 融合蛋白; 诱导表达
• ISSN: 1671-2927

The ω-atracotoxin-Arlb toxin （ω-ACTX-Arl b） is one of the arthropod-selective peptide neurotoxins from the venom of Australian funnel-web spider Atrax robustus. The gene of Arlb was synthesized and cloned into pET-32a（＋） vector to allow expression of Arlb as a fusion protein with thioredoxin and the His-tag （rTrx-Arlb） in E. coli BL21 （DE3）. The optimal condition for inducing the expression ofrTrx-Arl b was 1.0 mmol L-1 IPTG for 6 h at 28℃. The fusion protein rTrx- Arlb was expressed in soluble form and was purified effectively by HisTrap HP affinity column and rpHLPC and a final yield of purified rTrx-Arlb was 95 mg from 1 000 mL E. coli culture. The LD50 values for Mythimna separate and Tenebrio molitor were 111.66 and 11.04 ug g-1 determined by injection of the purified rTrx-Arlb. The results indicated that the recombinant Arlb protein was successfully expressed in E. coli and it was high toxicity against tested insects.