As an important mediator of allergic inflammation, mast cell tryptase is involved in the induction of hypersensitivity, infiltration of inflammatory cells and tissue remodeling in respiratory tract. The effects of try...As an important mediator of allergic inflammation, mast cell tryptase is involved in the induction of hypersensitivity, infiltration of inflammatory cells and tissue remodeling in respiratory tract. The effects of tryptase inhibitors on the actions of tryptase show further the potential of tryptase in the pathogenesis of asthma and its inhibitors in the treatment of asthma.展开更多
文摘苏云金芽胞杆菌(Bacillus thuringiensis,Bt)产生的Cry3Aa毒素在许多鞘翅目害虫的防治中发挥了重要作用。随着昆虫抗性的产生,如何通过大肠杆菌高通量表达筛选cry3Aa基因改造突变蛋白成为了研究的重点。然而,cry3Aa基因在大肠杆菌BL21(DE3)中诱导表达后其蛋白酶活化效果及杀虫活性均受到一定程度影响,对于影响的程度仍知之甚少。为探索cry3Aa基因所表达的融合蛋白GST-Cry3Aa中GST标签对Cry蛋白结构和功能的影响,本研究通过利用丝氨酸类蛋白酶(胰蛋白酶,糜蛋白酶)对纯化后的Cry3Aa融合蛋白进行水解条件研究分析,结果表明:在温度为11~37℃、GST-Cry3Aa与胰蛋白酶的质量比为4:1~30:1时,均可被水解出约55 k D的活性片段,而在同样条件下糜蛋白酶则无法对GST-Cry3Aa进行有效的水解活化。此外,结合I-TASSER和Swiss model对Cry3Aa与GST-Cry3Aa进行三维结构分析比对,发现GST-Cry3Aa在其结构域Ⅰ内有部分改变,推测其可能为两种丝氨酸类蛋白酶对该融合蛋白水解效果产生差异的原因。为后期构建Cry3Aa毒素高通量分子改造平台提供了理论基础。
文摘As an important mediator of allergic inflammation, mast cell tryptase is involved in the induction of hypersensitivity, infiltration of inflammatory cells and tissue remodeling in respiratory tract. The effects of tryptase inhibitors on the actions of tryptase show further the potential of tryptase in the pathogenesis of asthma and its inhibitors in the treatment of asthma.
