摘要
目的:探讨利用点突变方法改善EGV热稳定性的可能性和有效性。方法:对来源于Melanocarpus albomyces endoglucanase的耐热性纤维素酶maEG进行同源建模和序列比较,删除49位脯氨酸49P(del)进行定点突变,并将得到的突变体在毕氏酵母X33中表达,对表达产物进行酶活性和热稳定性检测。结果:突变酶49P(del)在70℃处理120min,热稳定性比EGV提高了21.6%,且突变酶其他性质与野生型酶基本相似。结论:通过对中性纤维素内切酶EGV的定点突变,提高了该酶的热稳定性,并为进一步研究其结构和功能提供了材料。结果同时表明利用生物信息学和分子模拟技术,缩短表面环区对于酶的热稳定性有一定的作用。
Objective： To explore the possibility and effects of improving enzyme thermostability by site-directed mutagenesis. Methods： Homology models and sequence alignment were conducted for Melanocarpus albomyces endoglucanase maEG. Then 49P（del） mutation was introduced into wide type EGV by site-directed mutagenesis. Both EGV and the mutant 49P （del） were expressed in Pichia pastoris. Their enzymatic properties were determined. Results： The result revealed that the thermostability of49P （del） was 34.5% higher than that of EGV at 70℃ for 180 min. The optimum temperature of 49P（del ） was increased by 10℃ in mutant enzymes than that in natural phytase. Additionally, the other enzymatic properties of mutant were similar to EGV. Conclusions： The mutant 49P（del） is a good materials for further research on structure and function of Cellulase EGV.
出处
《现代生物医学进展》
CAS
2009年第14期2634-2636,共3页
Progress in Modern Biomedicine
基金
国家自然科学基金重大研究计划项目(90412015)
教育部中国网格计划生物信息网格平台子项目(B1-137040130)
科技部基础学科平台建设生物计算网络平台项目(2005DKA64001)
