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Xylose isomerase from polycentric fungus Orpinomyces: gene sequencing, cloning, and expression in Saccharomyces cerevisiae for bioconversion of xylose to ethanol

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Abstract

The cDNA sequence of the gene for xylose isomerase from the rumen fungus Orpinomyces was elucidated by rapid amplification of cDNA ends. The 1,314-nucleotide gene was cloned and expressed constitutively in Saccharomyces cerevisiae. The deduced polypeptide sequence encoded a protein of 437 amino acids which showed the highest similarity to the family II xylose isomerases. Further, characterization revealed that the recombinant enzyme was a homodimer with a subunit of molecular mass 49 kDa. Cell extract of the recombinant strain exhibited high specific xylose isomerase activity. The pH optimum of the enzyme was 7.5, while the low temperature optimum at 37°C was the property that differed significantly from the majority of the reported thermophilic xylose isomerases. In addition to the xylose isomerase gene, the overexpression of the S. cerevisiae endogenous xylulokinase gene and the Pichia stipitis SUT1 gene for sugar transporter in the recombinant yeast facilitated the efficient production of ethanol from xylose.

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Acknowledgment

The author Anjali Madhavan was supported by the Council of Scientific and Industrial Research, India.

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Authors and Affiliations

  1. Department of Biochemical Engineering and Biotechnology, Indian Institute of Technology Delhi, Hauz Khas, 110016, Delhi, India

    Anjali Madhavan, Aradhana Srivastava & Virendra S. Bisaria

  2. Organization of Advanced Science and Technology, Kobe University, 1-1 Rokkodaicho, Nada-ku, Kobe, 657-8501, Japan

    Sriappareddy Tamalampudi, Satoshi Katahira & Hideki Fukuda

  3. Laboratory of Animal Science, Kyoto Prefectural University, Shimogamo, Kyoto, 606-8522, Japan

    Kazunari Ushida

  4. Research and Development Laboratories for Sustainable Value Creation, Asahi Breweries, Ltd, 1-21, Midori 1-Chome, Moriya-Shi, Ibaraki, 302-0106, Japan

    Daisuke Kanai

  5. Department of Chemical Science and Engineering, Graduate School of Engineering, Kobe University, 1-1 Rokkodaicho, Nada-ku, Kobe, 657-8501, Japan

    Akihiko Kondo

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  1. Anjali Madhavan
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  2. Sriappareddy Tamalampudi
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  3. Kazunari Ushida
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  9. Akihiko Kondo
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Correspondence to Akihiko Kondo.

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Madhavan, A., Tamalampudi, S., Ushida, K. et al. Xylose isomerase from polycentric fungus Orpinomyces: gene sequencing, cloning, and expression in Saccharomyces cerevisiae for bioconversion of xylose to ethanol. Appl Microbiol Biotechnol 82, 1067–1078 (2009). https://doi.org/10.1007/s00253-008-1794-6

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  • DOI: https://doi.org/10.1007/s00253-008-1794-6

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