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Engineering polyhydroxyalkanoate content and monomer composition in the oleaginous yeast Yarrowia lipolytica by modifying the ß-oxidation multifunctional protein

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Abstract

Recombinant strains of the oleaginous yeast Yarrowia lipolytica expressing the PHA synthase gene (PhaC) from Pseudomonas aeruginosa in the peroxisome were found able to produce polyhydroxyalkanoates (PHA). PHA production yield, but not the monomer composition, was dependent on POX genotype (POX genes encoding acyl-CoA oxidases) (Haddouche et al. FEMS Yeast Res 10:917–927, 2010). In this study of variants of the Y. lipolytica β-oxidation multifunctional enzyme, with deletions or inactivations of the R-3-hydroxyacyl-CoA dehydrogenase domain, we were able to produce hetero-polymers (functional MFE enzyme) or homo-polymers (with no 3-hydroxyacyl-CoA dehydrogenase activity) of PHA consisting principally of 3-hydroxyacid monomers (>80%) of the same length as the external fatty acid used for growth. The redirection of fatty acid flux towards β-oxidation, by deletion of the neutral lipid synthesis pathway (mutant strain Q4 devoid of the acyltransferases encoded by the LRO1, DGA1, DGA2 and ARE1 genes), in combination with variant expressing only the enoyl-CoA hydratase 2 domain, led to a significant increase in PHA levels, to 7.3% of cell dry weight. Finally, the presence of shorter monomers (up to 20% of the monomers) in a mutant strain lacking the peroxisomal 3-hydroxyacyl-CoA dehydrogenase domain provided evidence for the occurrence of partial mitochondrial β-oxidation in Y. lipolytica.

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Acknowledgments

We acknowledge the research grants provided by the European Commission in the framework of the LipoYeasts project (contract number 213068). This work was also partly funded by a grant from the “Fond National Suisse de la Recherche Scientifique” (grant number 3100A0-122493) and the University of Lausanne to Y.P. We thank Julie Sappa of Alex Edelman and Associates for her excellent help in correcting the English version of the manuscript.

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Authors and Affiliations

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

    Ramdane Haddouche, Julia Sabirova, Yves Pagot & Cécile Neuvéglise

  2. CNRS, Micalis, 78352, Jouy-en-Josas, France

    Jean-Marc Nicaud

  3. Laboratory of Industrial Microbiology and Biocatalysis, Department of Biochemical and Microbial Technology, Faculty of Bioscience Engineering, Ghent University, Ghent, Belgium

    Julia Sabirova

  4. Departement de Biologie Moléculaire Végétale, Laboratoire de Biotechnologie Végétale, Biophore, Université de Lausanne, 1015, Lausanne, Switzerland

    Yves Poirier & Syndie Delessert

  5. INRA, UMR1319, Micalis, Centre de Biotechnologie Agro-Industrielle, AgroParisTech, 78850, Thiverval-Grignon, France

    Jean-Marc Nicaud

Authors
  1. Ramdane Haddouche
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Correspondence to Jean-Marc Nicaud.

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Haddouche, R., Poirier, Y., Delessert, S. et al. Engineering polyhydroxyalkanoate content and monomer composition in the oleaginous yeast Yarrowia lipolytica by modifying the ß-oxidation multifunctional protein. Appl Microbiol Biotechnol 91, 1327–1340 (2011). https://doi.org/10.1007/s00253-011-3331-2

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