重组α-银环蛇毒素的纯化及活性分析

Purification and Biological Activity of the Recombinant α-bungarotoxin

以表达重组α-银环蛇毒素的高效表达菌株BL21(PDZ04)为材料,研究重组α-银环蛇毒素(α-bungarotoxin,α-BgTx)的分离纯化.采取亲和色谱和离子交换色谱的方法都得到了重组α-银环蛇毒素的纯品.首先从天然的银环蛇毒干粉中分离纯化得到了天然α-银环蛇毒素的纯品.然后以天然α-银环蛇毒素为对照来检测重组α-银环蛇毒素的抗原性和毒性.ELISA结果显示其具有与天然α-银环蛇毒素相似的抗原性,以小鼠为动物模型,纯化的重组α-银环蛇毒素与天然α-银环蛇毒素相比,其腹腔注射的LD50也基本一致,约为0.22μg/g.结果表明利用基因工程的方法生产蛇神经毒素是可行的.

The recombinant plasmid pDZ04 can efficiently express α bungarotoxin in the E coli BL21 This paper was to study the isolation and purification of the recombinant α bungarotoxin from the recombinant stains BL21(pDZ04) contructed by our laboratory laterly Firstly,on the basis of the biochemical properties of α bungarotoxin and the methods reported previously,the natural α bungarotoxin was purified from the crude snake venom Secondly,the recombinant α bungarotoxin was purified by affinity chromatography and ionexchange chromatography,respectively,the purity of the recombinant α bungarotoxin was over 95% Finally,the biological activity of recombinant α bungarotoxin and the fusion protein were studied with the natural α bungarotoxin purified from the snake venom as control.ELISA results showed that they had the similar antigenicity The fusion toxin had no obvious toxicity to mice while the recombinant and natural α bungarotoxin had almost the same LD 50 to mice,around 0 22 μg/g This research shows that it is promising to produce the snake neurotoxin by the genetical technology