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Molecular cloning, characterization, and engineering of xylitol dehydrogenase from Debaryomyces hansenii

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Abstract

Because of its natural ability to utilize both xylose and arabinose, the halotolerant and osmotolerant yeast Debaryomyces hansenii is considered as a potential microbial platform for exploiting lignocellulosic biomass. To gain better understanding of the xylose metabolism in D. hansenii, we have cloned and characterized a xylitol dehydrogenase gene (DhXDH). The cloned gene appeared to be essential for xylose metabolism in D. hansenii as the deletion of this gene abolished the growth of the cells on xylose. The expression of DhXDH was strongly upregulated in the presence of xylose. Recombinant DhXdhp was expressed and purified from Escherichia coli. DhXdhp was highly active against xylitol and sorbitol as substrate. Our results showed that DhXdhp was thermo-sensitive, and except this, its biochemical properties were quite comparable with XDH from other yeast species. Furthermore, to make this enzyme suitable for metabolic engineering of D. hansenii, we have improved its thermotolerance and modified cofactor requirement through modelling and mutagenesis approach.

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Acknowledgments

The authors thank Mr. Deepak Bhat and Mr. Randeep Sharma for technical assistance. The authors acknowledge the help of Ms. Connie Bobroff for correcting grammatical errors. D. B. is a recipient of Senior Research Fellowship from the Council of Scientific and Industrial Research, New Delhi, India.

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  1. Monika Aggarwal

    Present address: Laboratory of Molecular Gerontology, National Institute on Aging NIH, Baltimore, MD, USA

Authors and Affiliations

  1. Institute of Microbial Technology, Sector 39A, Chandigarh, 160 036, India

    Dipanwita Biswas, Manish Datt, Monika Aggarwal & Alok K. Mondal

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  1. Dipanwita Biswas
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  2. Manish Datt
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  3. Monika Aggarwal
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  4. Alok K. Mondal
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Correspondence to Alok K. Mondal.

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Biswas, D., Datt, M., Aggarwal, M. et al. Molecular cloning, characterization, and engineering of xylitol dehydrogenase from Debaryomyces hansenii . Appl Microbiol Biotechnol 97, 1613–1623 (2013). https://doi.org/10.1007/s00253-012-4020-5

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