Improvement of the catalytic performance of a hyperthermostable GH10 xylanase from Talaromyces leycettanus JCM12802
Improvement of the catalytic performance of a hyperthermostable GH10 xylanase from Talaromyces leycettanus JCM12802
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改进来自 Talaromyces leycettanus JCM12802 的超热稳定 GH10 木聚糖酶的催化性能
DOI：
10.1016/j.biortech.2016.10.003
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发表时间：
2016-12-01
影响因子：
11.4
通讯作者：
Luo, Huiying
中科院分区：
文献类型：
--
作者：
Wang, Xiaoyu;Huang, Huoqing;Luo, Huiying
从Talaromyces leycettanus JCM 12802中克隆了GH 10木聚糖酶基因Tlxyn 10A，并在毕赤酵母中表达。纯化的重组TlXyn 10A是酸性和极端嗜热的，并且在2.0-6.0的pH范围内和在90 ℃下保持稳定。TlXyn 10A的序列分析确定了7个可能参与底物接触的残基。然后通过用来自Bispora sp. MEY-1的高温Xyl 10 C的相应残基取代部分或全部残基来构建三个突变体（TlXyn10A_P、_N和_C）。在亚位点+2至+4处具有突变的TlXyn 10A_P表现出改善的比活性（0.44倍）和pH稳定性（2.0-10.0）。分子动力学模拟分析表明，突变E229 I和F232 E可能减弱了+3 ~+4亚位点的底物亲和力，G149 D可能引入了新的氢键。TlXyn10A_P具有较好的水解小麦秸秆的能力，在工业上具有很好的应用前景。(C)2016由Elsevier Ltd.出版
A xylanase gene of GH 10, Tlxyn10A, was cloned from Talaromyces leycettanus JCM12802 and expressed in Pichia pastoris. Purified recombinant TlXyn10A was acidic and hyperthermophilic, and retained stable over the pH range of 2.0-6.0 and at 90 degrees C. Sequence analysis of TlXyn10A identified seven residues probably involved in substrate contacting. Three mutants (TlXyn10A_P, _N and _C) were then constructed by substituting some or all of the residues with corresponding ones of hyperthermal Xyl10C from Bispora sp. MEY-1. TlXyn10A_P with mutations at subsites +2 to +4 exhibited improved specific activity (by 0.44-fold) and pH stability (2.0-10.0). Molecular dynamics simulation analysis indicated that mutations E229I and F232E probably weaken the substrate affinity at subsites +3 to +4, and G149D may introduce a new hydrogen bond. These modifications altogether account for the improved performance of TlXyn10A_P. Moreover, TlXyn10A_P was able to hydrolyze wheat straw persistently, and has the application potentials in various industries. (C) 2016 Published by Elsevier Ltd.