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Refolding of partially and fully denatured lysozymes

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Abstract

Lysozyme refolding with high yields sometimes results from incomplete denaturation. Dithiothreitol (DTT) is a reductant commonly used to reduce and unfold disulfide-stabilized lysozymes. Through the use of fluorescence spectroscopy to access the extent of denaturation, we found that the rate and extent of denaturation highly depended on the concentration of DTT. Further, the denaturation exhibited a two-phase transition at a high DTT concentration with DTT at >100 mM and long denaturation time (>24 h) being needed for complete denaturation. A low DTT concentration and a short denaturation time resulted in fast refolding with high activity recovery, while a high DTT concentration and a long denaturation time resulted in slow refolding with low activity recovery. Hence, the renaturation of disulfide-containing lysozyme was highly affected by the extent of denaturation.

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Acknowledgements

This study was financially supported by the National Science Council, Taiwan (NSC94-2214-E-008-002 and NSC93-2214-E-002-034) and the Center-of-Excellence Program on Membrane Technology, the Ministry of Education, Taiwan.

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Authors and Affiliations

  1. Department of Chemical Engineering, National Taiwan University, Taipei, Taiwan

    Jiun-Liang Lin & Hsyue-Jen Hsieh

  2. Department of Chemical and Materials Engineering, National Central University, Jhongli, Taiwan

    Ruoh-Chyu Ruaan

  3. R&D Center of Membrane Technology, Chung Yuan University, Jhongli, Taiwan

    Ruoh-Chyu Ruaan

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  1. Jiun-Liang Lin
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  2. Ruoh-Chyu Ruaan
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  3. Hsyue-Jen Hsieh
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Correspondence to Hsyue-Jen Hsieh.

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Lin, JL., Ruaan, RC. & Hsieh, HJ. Refolding of partially and fully denatured lysozymes. Biotechnol Lett 29, 723–729 (2007). https://doi.org/10.1007/s10529-007-9320-y

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  • DOI: https://doi.org/10.1007/s10529-007-9320-y

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