疏绵状嗜热丝孢菌外切β-葡聚糖酶的基因克隆与酶学特征  

Gene cloning and biochemical properties of Thermomyces lanuginosus exo-β-glucanase

作　　者：王瑞杰[1] Nokuthula Peace Mchunu 牛丹丹[3] Kugenthiren Permaul Suren Singh 刘晓光[1] 路福平[1] 

机构地区：[1]天津科技大学生物工程学院,天津300457 [2]Department of Biotechnology & Food Technology, Faculty of Applied Sciences, Durban University of Technology [3]福州大学生物科学与工程学院,福建福州350116

出　　处：《微生物学通报》2016年第2期285-291,共7页Microbiology China

基　　金：国家自然基金国际(地区)合作与交流项目(No.31461143026);天津市应用基础与前沿技术研究计划(青年)项目(No.14JCQNJC09200)~~

摘　　要：【目的】疏绵状嗜热丝孢菌是一种嗜热丝状真菌,具有合成与分泌多种耐热酶的能力,从中找寻酶活高、耐热性能优良的β-葡聚糖酶。【方法】对疏绵状嗜热丝孢菌其中一个外切β-葡聚糖酶的编码基因gln B进行克隆,并在毕赤酵母GS115中表达。【结果】在摇瓶水平上重组菌的产酶活为11.5 U/m L,重组酶在SDS-PAGE中的大小约为48 k D。重组Gln B最适作用温度为65°C,最适作用p H为5.0;在低于50°C或p H 3.0-10.0之间具有良好稳定性。重组酶水解海带三糖,首先生成单糖和二糖,延长酶作用时间,可以进一步将其中的二糖部分水解为单糖。【结论】疏绵状嗜热丝孢菌来源的重组酶Gln B为外切β-葡聚糖酶,具有较好的耐热性能和p H稳定性。[Objective] Thermophilic filamentous fungus Thermomyces lanuginosus is capable of secreting a host of thermostable enzymes. We are committed to find a high activity and thermostable β-glucanase gene. [Mehtods] A novel exo-β-glucanase gene（gln B） was cloned from T. lanuginosus and successfully expressed in Pichia pastoris GS115. [Results] SDS-PAGE shows that Gln B has a molecular weight of about 48 k D. Levels of recombinant Gln B reached 11.5 U/m L in the flask fermentation broth of recombinant Pichia cells. Gln B exhibited optimum temperature and p H at 65 ℃ and 5.0, respectively, and was stable below 50 ℃ and over the p H range 3.0-10.0. Hydrolysis of laminarin by the recombinant Gln B produced monosaccharide and disaccharide, and extended incubation resulted in partial hydrolysis of disaccharide. [Conclusion] The recombinant enzyme Gln B from T. lanuginosus is an exo-β-glucanase and has good command of thermo stability and p H stability.

关 键 词：疏绵状嗜热丝孢菌 外切β-葡聚糖酶 酶学特征 

分 类 号：Q936[生物学—微生物学] Q78