Truncation of a mannanase from Trichoderma harzianum improves its enzymatic properties and expression efficiency in Trichoderma reesei
Truncation of a mannanase from Trichoderma harzianum improves its enzymatic properties and expression efficiency in Trichoderma reesei
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DOI：
10.1007/s10295-013-1359-2
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发表时间：
2013
影响因子：
3.4
通讯作者：
Juan Wang;D. Zeng;Gang Liu;Shaowen Wang;Shaowen Yu
中科院分区：
文献类型：
--
作者：
Juan Wang;D. Zeng;Gang Liu;Shaowen Wang;Shaowen Yu
为获得高效表达的甘露聚糖酶基因ThMan 5A，利用强组成型启动子pdc，在里氏木霉（Trichoderma reeseiQM 9414）中表达了全长基因ThMan 5A和仅含催化结构域的截短基因ThMan 5A △CBM，并纯化至均一。我们发现，基因的截短提高了其表达效率以及编码蛋白的酶性质。表达ThMan 5A △CBM的重组菌株在培养上清中产生2，460 ± 45.1 U/ml的甘露聚糖酶活性，是表达全长ThMan 5A基因的2.3倍。此外，与全长酶相比，截短的甘露聚糖酶具有上级的热稳定性，在60 °C孵育48 h后仍保持100%的活性。我们的结果清楚地表明，截短的ThMan 5A酶在表达效率和其热稳定性方面都表现出改善的特性。这些特性表明ThMan 5A △CBM在食品、饲料、造纸和纸浆工业中具有潜在的应用价值。
To obtain high expression efficiency of a mannanase gene,ThMan5A,cloned fromTrichoderma harzianumMGQ2, both the full-length gene and a truncated gene (ThMan5A△CBM) that contains only the catalytic domain, were expressed inTrichoderma reeseiQM9414 using the strong constitutive promoter of the gene encoding pyruvate decarboxylase (pdc), and purified to homogeneity, respectively. We found that truncation of the gene improved its expression efficiency as well as the enzymatic properties of the encoded protein. The recombinant strain expressingThMan5A△CBM produced 2,460 ± 45.1 U/ml of mannanase activity in the culture supernatant; 2.3-fold higher than when expressing the full-lengthThMan5Agene. In addition, the truncated mannanase had superior thermostability compared with the full-length enzyme and retained 100 % of its activity after incubation at 60 °C for 48 h. Our results clearly show that the truncated ThMan5A enzyme exhibited improved characteristics both in expression efficiency and in its thermal stability. These characteristics suggest that ThMan5A△CBM has potential applications in the food, feed, paper, and pulp industries.