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Improving the thermostability of a mesophilic family 10 xylanase, AuXyn10A, from Aspergillus usamii by in silico design

  • Biotechnology Methods
  • Published:
Journal of Industrial Microbiology & Biotechnology

Abstract

To improve the thermostability of a mesophilic GH family 10 xylanase, AuXyn10A, from Aspergillus usamii E001, its modification was performed by in silico design. Based on the comparison of B-factor values, a mutant xylanase ATXyn10 was predicted by substituting a segment YP from Tyr25 to Pro34 of AuXyn10A with the corresponding one from Asn24 to Ala32 of TaXyn10, a thermophilic GH family 10 xylanase from Thermoascus aurantiacus. Analysis of a TaXyn10 crystal structure indicated that there is a close interaction between segments YP and FP. For that reason, another mutant xylanase ATXyn10M was designed by mutating Ser286 and His288 of ATXyn10 into the corresponding Gly285 and Phe287 in the FP of TaXyn10. Then, two ATXyn10- and ATXyn10M-encoding genes, ATxyn10 and ATxyn10 M, were expressed in Pichia pas toris GS115. The temperature optimum of recombinant (re) ATXyn10M was 60 °C, 10 °C higher than that of reAuXyn10A. Its thermal inactivation half-life (t 1/2) at 55 °C was 10.4-fold longer than that of reAuXyn10A. As compared with reAuXyn10A, reATXyn10M displayed a slight decrease in K m value and a significant increase in V max value from 6,267 to 8,870 U/mg.

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Acknowledgments

This work was supported by the National Nature Science Foundation of China (No. 31101229), Key Laboratory of Carbohydrate Chemistry and Biotechnology, Ministry of Education, Jiangnan University (No. KLCCB-KF201208), Doctoral Research Funds of Jiangnan University (No. JUDCF11032) and Postgraduate Innovation Training Project of Jiangsu (No. CXZZ12_0758). The authors are grateful to Prof. Xianzhang Wu (School of Biotechnology, Jiangnan University) for providing technical assistance.

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Authors and Affiliations

  1. Key Laboratory of Carbohydrate Chemistry and Biotechnology, Ministry of Education, Jiangnan University, 1800 Lihu Road, Wuxi, 214122, Jiangsu, People’s Republic of China

    Junqing Wang & Minchen Wu

  2. Key Laboratory of Industrial Biotechnology, Ministry of Education, School of Biotechnology, Jiangnan University, 1800 Lihu Road, Wuxi, 214122, Jiangsu, People’s Republic of China

    Junqing Wang & Zhongbiao Tan

  3. State Key Laboratory of Food Science and Technology, School of Food Science and Technology, Jiangnan University, Wuxi, 214122, Jiangsu, China

    Jianfang Li

  4. Wuxi Medical School, Jiangnan University, Wuxi, 214122, Jiangsu, China

    Minchen Wu & Jing Wu

  5. School of Medicine and Pharmaceutics, Jiangnan University, 1800 Lihu Road, Wuxi, 214122, Jiangsu, People’s Republic of China

    Minchen Wu

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  1. Junqing Wang
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  2. Zhongbiao Tan
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  3. Minchen Wu
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  4. Jianfang Li
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  5. Jing Wu
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Correspondence to Minchen Wu.

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J. Wang and Z. Tan, the two first authors, contributed equally to this work.

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Wang, J., Tan, Z., Wu, M. et al. Improving the thermostability of a mesophilic family 10 xylanase, AuXyn10A, from Aspergillus usamii by in silico design. J Ind Microbiol Biotechnol 41, 1217–1225 (2014). https://doi.org/10.1007/s10295-014-1463-y

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  • DOI: https://doi.org/10.1007/s10295-014-1463-y

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