苏云金芽孢杆菌cry基因在大肠杆菌中表达产物和生物活性

Purification and bioassay of some products of Bacillus thuringiensis cry genes expressed in Escherichia coli strain

研究了几种Btcry基因于大肠杆菌 (Escherichiacoli)中表达产物在pH 10 0的 5 0mmol/L碳酸钠和 2 0mmol/L乙醇胺溶解液中的溶解性 ,发现同样的Cry蛋白在碳酸钠中的溶解度大于乙醇胺。通过胰蛋白酶消化 ,明确Cry1Ca7、Cry1Ia8酶解产物为 38kD多肽 ;Cry1Ie1、Cry1Cb2、Cry2Ab4酶解产物为 4 1kD多肽 ;Cry1Ac酶解产物为 6 0kD多肽。采用FPLC层析方法对 6种原毒素及其酶解后得到的毒素多肽进行了分离纯化 ,比较了原毒素和毒素的杀虫活性的差异。其结果表明 ,Cry1Ac的原毒素和毒素对棉铃虫初孵幼虫的校正死亡率均为 10 0 % ,Cry2Ab4的原毒素的毒力高于其酶解毒素。

The solubility of several Bt Cry proteins expressed in Escherichia coli strain in 50 mmol/L Na 2CO 3 and 20 mmol/L ethanolamine were compared and the solubility of all the Cry proteins in Na 2CO 3 were better than those in ethanolamine solution. After being digested by trypsin, a 38kD peptide was obtained from toxin Cry1Ca7 and Cry1Ia8, and a 41kD peptide was detected from Cry1Ie1, Cry1Cb2 and Cry2Ab4, but a 60kD fragment was found from Cry1Ac. Furthermore, the bioassay of all the protoxins and their activated-toxins digested by trypsin were performed against cotton boll worm larvae. No difference of toxicity could be found in protoxins and toxins activated by trypsin of Cry1Ac, but the toxicity of Cry2Ab4 was reduced in comparison with its protoxin.