A method to rationally increase protein stability based on the charge–charge interaction, with application to lipase LipK107

Authors

  • Lujia Zhang,

    1. State Key Laboratory of Bioreactor Engineering, East China University of Science and Technology, Shanghai, People's Republic of China
    2. State Key Laboratory of Materials-Oriented Chemical Engineering, Nanjing University of Technology, Nanjing, People's Republic of China
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  • Xiaomang Tang,

    1. State Key Laboratory of Bioreactor Engineering, East China University of Science and Technology, Shanghai, People's Republic of China
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  • Dongbing Cui,

    1. State Key Laboratory of Bioreactor Engineering, East China University of Science and Technology, Shanghai, People's Republic of China
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  • Zhiqiang Yao,

    1. State Key Laboratory of Bioreactor Engineering, East China University of Science and Technology, Shanghai, People's Republic of China
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  • Bei Gao,

    1. State Key Laboratory of Bioreactor Engineering, East China University of Science and Technology, Shanghai, People's Republic of China
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  • Shuiqin Jiang,

    1. State Key Laboratory of Bioreactor Engineering, East China University of Science and Technology, Shanghai, People's Republic of China
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  • Bo Yin,

    1. State Key Laboratory of Bioreactor Engineering, East China University of Science and Technology, Shanghai, People's Republic of China
    2. National University of Singapore (Suzhou) Research Institute, People's Republic of China
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  • Y. Adam Yuan,

    Corresponding author
    1. National University of Singapore (Suzhou) Research Institute, People's Republic of China
    2. Department of Biological Sciences, Centre for Bioimaging Sciences, National University of Singapore, Singapore
    • Corresponding authors: Dongzhi Wei, 130 Meilong road, Shanghai, People's Republic of China, E-mail: dzhwei@ecust.edu.cn and Y Adam Yuan, 377 Lin Quan Street, Suzhou Industrial Park, Jiangsu 215123, People's Republic of China. E-mail: dbsyya@nus.edu.sg

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  • Dongzhi Wei

    Corresponding author
    1. State Key Laboratory of Bioreactor Engineering, East China University of Science and Technology, Shanghai, People's Republic of China
    • Corresponding authors: Dongzhi Wei, 130 Meilong road, Shanghai, People's Republic of China, E-mail: dzhwei@ecust.edu.cn and Y Adam Yuan, 377 Lin Quan Street, Suzhou Industrial Park, Jiangsu 215123, People's Republic of China. E-mail: dbsyya@nus.edu.sg

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Abstract

We report a suite of enzyme redesign protocol based on the surface charge–charge interaction calculation, which is potentially applied to improve the stability of an enzyme without compromising its catalytic activity. Together with the experimental validation, we have released a suite of enzyme redesign algorithm Enzyme Thermal Stability System, written based on our model, for open access to meet the needs in wet labs. Lipk107, a lipase of a versatile industrial use, was chosen to test our software. Our calculation determined that four residues, D113, D149, D213, and D253, located on the surface of LipK107 were critical to the stability of the enzyme. The model was validated with mutagenesis at these four residues followed by stability and activity tests. LipK107 mutants D113A and D149K were more resistant to thermal inactivation with ∼10°C higher half-inactivation temperature than wild-type LipK107. Moreover, mutant D149K exhibited significant retention in residual activity under constant heat, showing a 14-fold increase in the half-inactivation time at 50°C. Activity tests showed that these mutants retained the equal or higher specific activity, among which noteworthy was the mutant D253A with as much as 20% higher activity. We suggest that our protocol could be used as a general guideline to redesign protein enzymes with increased stabilities and enhanced activities.

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