Achaetomium sp.Xz8来源的甘露聚糖酶Man5Xz8热稳定性的研究  

Thermostability Improvement of the Mannanase Man5Xz8 from Achaetomium sp. Xz8

作　　者：刘赟[1,2] 卢海强[2] 石鹏君[2] 谢响明[1] 

机构地区：[1]北京林业大学生物科学与技术学院,北京100083 [2]中国农业科学院饲料研究所,北京100081

出　　处：《中国农业科技导报》2015年第3期49-55,91,共8页Journal of Agricultural Science and Technology

基　　金：国家863计划项目(2012AA022208);中央高校基本科研业务费专项(TD2012-03)资助

摘　　要：嗜热真菌Achaetomium sp.Xz8来源的甘露聚糖酶Man5Xz8和Humicola sp.Y1来源的嗜热甘露聚糖酶Man5A氨基酸序列一致性高达90.0%,然而两者的酶学性质尤其是最适温度和热稳定性却存在着很大的不同。Man5Xz8的最适温度为50℃,仅在30℃下保持稳定,而Man5A最适温度为70℃,且在50℃下具有良好的热稳定性。为研究不同区域关键氨基酸位点对甘露聚糖酶热稳定性的影响,以Man5Xz8为研究材料,通过生物信息学方法分析,分别构建了单点突变体V124I及△SG。实验结果表明,突变体V124I及△SG和原酶Man5Xz8具有相似的最适p H和最适温度,但在热稳定性方面较原酶有很大提高:在50℃下处理10 min后,突变体V124I的相对剩余酶活力较原酶Man5Xz8提高了26.9%,突变体△SG在50℃下处理1 h保持稳定。通过对甘露聚糖酶不同区域氨基酸位点的研究,发现了与酶热稳定性相关的关键结构区域及位点,为以后更好的研究第5家族的甘露聚糖酶提供了一定的理论指导。The consistency of amino acid sequence of two reported GH5 mannase, Man5Xz8 from the thermophilic fungi Achaetomium sp. Xz8 and ManSA from Humicola sp. Y1, was as high as 90.0%. However, their enzymology properties, especially their optimum temperature and thermal stability were different. Man5A had the temperature optima of 70℃ and possessed pretty good thermal stability under 50℃, while ManSXz8 had the temperature optima of 50℃ and only could stay good thermal stability at 30℃. The mutants V124I and ASG were built based on the analysis of bioinformatics methods. The results showed that the mutants had similar optimal pH and temperature with ManSXzS, while their thermal stabilities were much betten than ManSXzS. After treated at 50℃ for 10 rain, the thermal stability of V124I was greatly increased when compared with their wild-type Man5Xz8 and the residual enzyme activity was increased by 26.9%. Moreover, the mutant ASG can stay good thermal stability at 50℃ for 1 h. Through the study of different amino acid sites of ManSXzS, some key factors of the thermal stability were found, which provided certain theoretical guidance for the better research of GH5 famliy mannases.

关 键 词：甘露聚糖酶 嗜热微生物 Achaetomium sp. 热稳定性 生物信息学 

分 类 号：Q814[生物学—生物工程]