Bt菌株中Cry1Ia和Cry2Ab结构域交换突变体的表达形式

Study on the Expression Pattern of Domain-exchange Mutants of Cry1Ia and Cry2Ab in Bt Strain

前期构建了一系列Cry1Ia和Cry2Ab蛋白的结构域交换突变体,并在Bt菌株中获得了可观的表达。对6种突变体的表达形式及分泌与否进行了分析和检测。结果显示,6种突变体在Bt中以包涵体的形式存在于菌液沉淀中;这些包涵体与正常Cry蛋白形成的晶体不同,不能溶于Na2CO3缓冲液。蛋白印迹法检测结果显示,带有Cry1Ia分泌信号肽的突变体IAA,IIA和IAI蛋白未分泌到胞外,这可能是由于蛋白表达量过快,迅速聚集而形成包涵体,从而无法分泌到胞外。

In the study, we researched the expression pattern of the 6 domain-exchange mutants, constructed previously, in Bacillus thuringiensis. The results showed that all of the 6 mutants expressed in high level as the inclusion bodies. These inclusion bodies could not dissolved in Na2CO3 buffer in which was similar with the pH of caterpillar＇s midgut. Three of these mutants （IAA, IIA and IAI ） contains predicted secretion signal peptides （PSSP）, but none of them could be detected in the supernatant of the cell medium by western blot analysis. This may be due to the rapid accumulation of target protein molecules when expressed in cell, resulting in insoluble inclu- sion bodies, finally.