Directed evolution of a mesophilic fungal xylanase by fusion of a thermophilic bacterial carbohydrate-binding module
Directed evolution of a mesophilic fungal xylanase by fusion of a thermophilic bacterial carbohydrate-binding module
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通过嗜热细菌碳水化合物结合模块的融合定向进化嗜温真菌木聚糖酶
DOI：
10.1016/j.procbio.2010.07.026
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发表时间：
2011-01-01
影响因子：
4.4
通讯作者：
Shen, Jinwen
中科院分区：
文献类型：
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作者：
Liu, Liangwei;Cheng, Jie;Shen, Jinwen
细菌GH10木聚糖酶通常含有与不溶性木聚糖结合的碳水化合物结合模块(CBM)。与GH10木聚糖酶不同，我们从黑曲霉(Aspergilus niger，XYN)中分离到一个含有单一催化结构域的真菌GH11木聚糖酶。为了提高中温XYN的热活性和对不溶性木聚糖的催化效率，我们将其与TmCBM9-12融合，起到了主动抓取木聚糖的作用。融合木聚糖酶XYN-TmCBM9-1_2的最适pH为4.2，最适温度为49℃，比XYN的热活性高2℃。该嵌合木聚糖酶对不溶性燕麦木聚糖的酶活为970.1+/-5.8U/mg，是对可溶性桦木木聚糖的4.2U/mg(228.1+/-1.1U/mg)。相比之下，XYN对不溶性燕麦木聚糖的活力为226.9+/-1.2U/mg，仅为对可溶性桦木木聚糖活力(567.2+/-3.0U/mg)的40%。与TmCBM9-1_2的融合提高了XYN的性质，表明我们可以通过融合细菌和真菌的结构域来指导分子功能的进化。(C)2010爱思唯尔有限公司。保留所有权利。
Bacterial GH10 xylanase usually contains carbohydrate-binding module (CBM) that binds to insoluble xylan. Differing from GH10 xylanase, we isolated a fungal GH11 xylanase containing a single catalytic domain from Aspergilus niger (XYN). The thermophilic CBM from Thermotoga maritima (TmCBM9-12) might increase the mesophilic XYN's thermo-activity and catalytic efficiency on insoluble xylan, we fused it with the TmCBM9-12 behaving as "hand" to grasp xylan actively. The chimeric xylanase XYN-TmCBM9-1_2 exhibited an optimal activity at pH4.2 and 49 degrees C, 2 degrees C higher than the thermo-activity of XYN. The chimeric xylanase's activity was 970.1 +/- 5.8 U/mg on insoluble oat-spelt xylan, 4.2-fold of that on soluble birchwood xylan (228.1 +/- 1.1 U/mg). In contrast, the XYN's activity was 226.9 +/- 1.2 U/mg on insoluble oat-spelt xylan, only 40% of that on soluble birchwood xylan (567.2 +/- 3.0 U/mg). Fusing with the TmCBM9-1_2 increased the XYN's property, indicating that we can direct to evolve a molecule's function through fusing domains of bacteria and fungi. (C) 2010 Elsevier Ltd. All rights reserved.