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Improvement in the thermostability of a type A feruloyl esterase, AuFaeA, from Aspergillus usamii by iterative saturation mutagenesis

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Abstract

Feruloyl or ferulic acid esterase (Fae, EC 3.1.1.73) catalyzes the hydrolysis of ester bonds between polysaccharides and phenolic acid compounds in xylan side chain. In this study, the thermostability of a type A feruloyl esterase (AuFaeA) from Aspergillus usamii was increased by iterative saturation mutagenesis (ISM). Two amino acids, Ser33 and Asn92, were selected for saturation mutagenesis according to the B-factors analyzed by B-FITTER software and ΔΔG values predicted by PoPMuSiC algorithm. After screening the saturation mutagenesis libraries constructed in Pichia pastoris, 15 promising variants were obtained. The best variant S33E/N92-4 (S33E/N92R) produced a T m value of 44.5 °C, the half-lives (t 1/2) of 35 and 198 min at 55 and 50 °C, respectively, corresponding to a 4.7 °C, 2.33- and 3.96-fold improvement compared to the wild type. Additionally, the best S33 variant S33-6 (S33E) was thermostable at 50 °C with a t 1/2 of 82 min, which was 32 min longer than that of the wild type. All the screened S33E/N92 variants were more thermostable than the best S33 variant S33-6 (S33E). This work would contribute to the further studies on higher thermostability modification of type A feruloyl esterases, especially those from fungi. The thermostable feruloyl esterase variants were expected to be potential candidates for industrial application in prompting the enzymic degradation of plant biomass materials at elevated temperatures.

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Acknowledgments

The authors are grateful to Prof. **anzhang Wu (School of Biotechnology, Jiangnan University, Jiangsu, China) for providing technical assistance.

Funding

This work was financially supported by the Fundamental Research Fund for the Central Universities of China (No. JUDCF13011, JUSRP51412B), the Postgraduate Innovation Training Project of Jiangsu (No. CXZZ13_0757), and the National Training Programs of Innovation and Entrepreneurship for Undergraduates (201410295037).

Conflict of interest

The authors declare that they have no competing interests.

Ethical statement

This article does not contain any studies with human participants or animals performed by any of the authors.

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Authors and Affiliations

  1. Key Laboratory of Carbohydrate Chemistry and Biotechnology, Ministry of Education, School of Biotechnology, Jiangnan University, 1800 Lihu Road, Wuxi, 214122, China

    **n Yin, Die Hu & Qin Wu

  2. State Key Laboratory of Food Science and Technology, School of Food Science and Technology, Jiangnan University, 1800 Lihu Road, Wuxi, 214122, China

    Jian-Fang Li & Chun-Juan Wang

  3. Wuxi Medical School, Jiangnan University, 1800 Lihu Road, Wuxi, 214122, China

    Ying Gu & Min-Chen Wu

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  1. **n Yin
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  2. Jian-Fang Li
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  4. Die Hu
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  5. Qin Wu
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  7. Min-Chen Wu
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Correspondence to Min-Chen Wu.

Additional information

**n Yin and Jian-Fang Li, the two first authors, contributed equally to this work.

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Yin, X., Li, JF., Wang, CJ. et al. Improvement in the thermostability of a type A feruloyl esterase, AuFaeA, from Aspergillus usamii by iterative saturation mutagenesis. Appl Microbiol Biotechnol 99, 10047–10056 (2015). https://doi.org/10.1007/s00253-015-6889-2

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