嗜热绿色糖单孢菌木聚糖酶XyN10A基因在毕赤酵母中的分泌表达及其酶学性质研究  

Secretory Expression in Pichia pastoris and Characterization Study of Xylanase from Thermophilic Saccharomonospora viridis

作　　者：玉王宁[1] 刘伟娜[1] 郑菲[1] 厚凌宇[1] 王晓宇[1] 梁迪[1] 王伟轩[1] 张朔[1] 柳静言 金一[1] 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

出　　处：《生物技术通报》2014年第6期173-180,共8页Biotechnology Bulletin

基　　金：国家自然科学基金项目(31070635;30870028;J1103516);公益性行业(农业)科研专项(201103027);北京市大学生科学研究与创业行动计划(S1310022019)

摘　　要：木聚糖酶(Xylanase)可降解自然界中大量存在的木聚糖类半纤维素,由于其在饲料、食品、造纸等领域的广泛应用,越来越受到人们的关注。利用分子生物学技术首次成功构建了一株高效表达嗜热绿色糖单孢菌(Saccharomonospora viridis)木聚糖酶SviXyN10A的重组工程菌,建立了SviXyN10A的毕赤酵母表达体系,实现了SviXyN10A在毕赤酵母(Pichia pastoris)中的分泌表达,并对表达产物进行了诱导表达及纯化,以及酶学性质的研究。结果表明,重组木聚糖酶SviXyn10A用甲醇诱导2 d后酶活即达到7.53IU/mL,最适反应温度为70℃,pH7.0。pH作用范围较广(pH5.0-9.0),具有较强的热稳定性和耐碱性,几乎没有纤维素酶活性,故可以应用在很多工业领域,尤其是在纸浆造纸行业具有很大的潜在应用价值。Xylanase has attracted extensive attention because of the application potential in feed, food and paper area, capable of degrading hemicellulose which is prevalent in nature. This research is to construct a recombinant engineered bacteria with efficient secretory expression for thermophilic Saccharomonospora viridis xylanase SviXyN10A using molecular biology techniques, and then study the characterization of the gene product expressed in Pichia pastoris. The results indicate that the gene product of Svixyn10A was successfully expressed in Pichia pastoris, showing the highest enzymatic activity of 7.53 IU/mL with methanol induction for 2 days. Recombinant xylanase SviXyN10A showed good activity at wide range of pH effect（pH5.0-9.0）, and the optimum reaction temperature is 70℃, pH7.0. What is more, the xylanase showed excellent thermo-alkali-stability and almost no cellulase activity. Thus, it is a potential candidate for future use in industrial areas particularly in the pulp and paper industry.

关 键 词：绿色糖单孢菌 木聚糖酶 分泌表达 毕赤酵母 

分 类 号：Q55[生物学—生物化学]