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A molecular approach to optimize hIFN α2b expression and secretion in Yarrowia lipolytica

  • Applied Genetics and Molecular Biotechnology
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Abstract

In this work, we investigated the effect of codon bias and consensus sequence (CACA) at the translation initiation site on the expression level of heterologous proteins in Yarrowia lipolytica; human interferon alpha 2b (hIFN-α2b) was studied as an example. A codon optimized hIFN-α2b gene was synthesized according to the frequency of codon usage in Y. lipolytica. Both wild-type (IFN-wt) and optimized hIFN-α2b (IFN-op) genes were expressed under the control of a strong inducible promoter acyl-co-enzyme A oxidase (POX2). Protein secretion was directed by the targeting sequence of the extracellular lipase (LIP2): pre–proLIP2. Codon optimization increased protein production by 11-fold, whereas the insertion of CACA sequence upstream of the initiation codon of IFN-op construct resulted in 16.5-fold increase of the expression level; this indicates that translational efficiency plays an important part in the increase of hIFN-α2b production level. The replacement of the pre–proLIP2 signal secretion with the LIP2 pre-region sequence followed by the X-Ala/X-Pro stretch but without the pro-region also increased the secretion of the target protein by twofold, suggesting therefore that the LIP2 pro-region is not necessary for extracellular secretion of small heterologous proteins in Yarrowia lipolytica.

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Acknowledgments

Najla Gasmi would like to acknowledge EUGIDE for their financial support in the frame of the CMCU project France Tunisia (grant N°. 07G0913). This work has also been partially funded by Oxyrane Belgium. We would like to thank Ms Mouna Nabli (Pasteur Institute of Tunis) for the construction of PICZα plasmid carrying the wild-type hIFN gene. The authors are very grateful to Dr. Sylvie Blanchin for her help in qPCR technique.

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  1. Franck Fudalej

    Present address: Oxyrane Belgium nv, Technologiepark 3, 9052, Gent-Zwijnaarde, Belgium

Authors and Affiliations

  1. INRA, UMR1319 Micalis, Domaine de Vilvert, 78352, Jouy-en-Josas, France

    Najla Gasmi, Franck Fudalej & Jean-Marc Nicaud

  2. Unité de Biofermentation, Institut Pasteur de Tunis, 13, place Pasteur, BP 74, 1002, Tunis Belvédère, Tunisia

    Najla Gasmi & Héla Kallel

  3. CNRS, Micalis, Domaine de Vilvert, 78352, Jouy-en-Josas, France

    Jean-Marc Nicaud

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  1. Najla Gasmi
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  2. Franck Fudalej
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Correspondence to Héla Kallel.

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Gasmi, N., Fudalej, F., Kallel, H. et al. A molecular approach to optimize hIFN α2b expression and secretion in Yarrowia lipolytica . Appl Microbiol Biotechnol 89, 109–119 (2011). https://doi.org/10.1007/s00253-010-2803-0

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