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Assessment of Schwanniomyces occidentalis as a host for protein production using the wide-range Xplor®2 expression platform

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Abstract

The wide-range transformation/expression platform, Xplor®2, was employed for the assessment of Schwanniomyces occidentalis as a potential producer of the recombinant proteins human IFNα2a (IFNα2a) and S. occidentalis fructofuranosidase (SFfase), and its efficiency was compared to that of Arxula adeninivorans. ADE2 and URA3 genes from both yeast species were isolated, characterized and used as selection markers in combination with the IFNα2a and SFfase expression modules, which used the strong constitutive A. adeninivorans-derived TEF1 promoter. Yeast rDNA integrative expression cassettes and yeast integrative expression cassettes equipped with a selection marker and expression modules were transformed into auxotrophic S. occidentalis and A. adeninivorans strains and a quantitative comparison of the expression efficiency was made. Whilst IFNα2a was mainly accumulated extracellularly (>95 %) in A. adeninivorans, extracellular SFfase (>90 %) was detected in both yeast species. The DNA composition of the selection marker modules and expression modules, especially their open reading frame codon usage, affects auxotrophy recovery as well as protein expression. Auxotrophy recovery was only achieved with selection marker modules of the homologous gene donor yeast. The concentration of recombinant IFNα2a was fivefold higher in A. adeninivorans (1 mg L−1), whereas S. occidentalis accumulated 1.5- to 2-fold more SFfase (0.5 Units ml−1). These results demonstrate the extension of the use of the wide-range expression platform Xplor®2 to another yeast species of biotechnological interest.

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Acknowledgments

MAB was founded by a Comunidad de Madrid Fellowship and granted by the Universidad Autónoma de Madrid. We would like to thank I. Schmeling and A. Martín for excellent technical assistance. Part of this study is based upon work funded National project BIO2010-20508-C04-04 and by an institutional grant from the Fundación Ramón Areces to the Centro de Biología Molecular Severo Ochoa.

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Authors and Affiliations

  1. Centro de Biología Molecular Severo Ochoa, Universidad Autónoma de Madrid, C/ Nicolás Cabrera 1, 28049, Madrid, Spain

    Miguel Álvaro-Benito & María Fernández-Lobato

  2. School of Biological Sciences, University of Canterbury, Private Bag 4800, Christchurch, New Zealand

    Keith Baronian

  3. Leibniz-Institut für Pflanzengenetik und Kulturpflanzenforschung (IPK), Corrensstr. 3, 06466, Gatersleben, Germany

    Gotthard Kunze

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  1. Miguel Álvaro-Benito
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  2. María Fernández-Lobato
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  3. Keith Baronian
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  4. Gotthard Kunze
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Correspondence to Gotthard Kunze.

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Álvaro-Benito, M., Fernández-Lobato, M., Baronian, K. et al. Assessment of Schwanniomyces occidentalis as a host for protein production using the wide-range Xplor®2 expression platform. Appl Microbiol Biotechnol 97, 4443–4456 (2013). https://doi.org/10.1007/s00253-012-4527-9

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