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Cumulative improvements of thermostability and pH-activity profile of Aspergillus niger PhyA phytase by site-directed mutagenesis

  • Applied Genetics and Molecular Biotechnology
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Abstract

Aspergillus niger phytase (PhyA) has been used as a feed supplement to reduce manure phosphorus excretion of swine and poultry but lacks sufficient thermostability for feed pelleting and appropriate pH-activity profile for phytate hydrolysis in the stomach of animals. Previously, a thermostable mutant PhyA18 and two pH-activity profile-improved mutants E228K and K300E were developed. In this study, the mutations were combined to determine if both improvements were cumulative. Four substitutions (S149P, F131L, K112R, and K195R) identified from random mutagenesis were added sequentially to the combined mutants to further improve their thermostability. Mutant E228K shifted the optimum pH of the parent one from 5.5 to 4.0 and increased (P < 0.05) its specific activity at pH 3.5, whereas mutant K300E eliminated the activity dip at pH 3.5 shown in the wild type. Mutant S149P further improved thermostability over PhyA18. Our results illustrate the feasibility and structural basis to improve thermostability and pH-activity profile of PhyA phytase by assembling mutations derived from rational design and random mutagenesis.

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Acknowledgments

This research was supported in part by a Cornell Biotechnology Program grant (to XL).

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Authors and Affiliations

  1. Department of Animal Science, Cornell University, 252 Morrison Hall, Ithaca, NY, 14853, USA

    Wanming Zhang & Xin Gen Lei

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  1. Wanming Zhang
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  2. Xin Gen Lei
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Correspondence to Xin Gen Lei.

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Zhang, W., Lei, X.G. Cumulative improvements of thermostability and pH-activity profile of Aspergillus niger PhyA phytase by site-directed mutagenesis. Appl Microbiol Biotechnol 77, 1033–1040 (2008). https://doi.org/10.1007/s00253-007-1239-7

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  • DOI: https://doi.org/10.1007/s00253-007-1239-7

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