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Metabolic engineering for ricinoleic acid production in the oleaginous yeast Yarrowia lipolytica

  • Applied genetics and molecular biotechnology
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Abstract

Although there are numerous oleochemical applications for ricinoleic acid (RA) and its derivatives, their production is limited and subject to various safety legislations. In an effort to produce RA from alternative sources, we constructed a genetically modified strain of the oleaginous yeast Yarrowia lipolytica. This strain is unable to perform β-oxidation and is invalidated for the native triacylglycerol (TAG) acyltransferases (Dga1p, Dga2p, and Lro1p) and the ∆12 desaturase (Fad2p). We also expressed the Ricinus communis ∆12 hydroxylase (RcFAH12) under the control of the TEF constitutive promoter in this strain. However, RA constituted only 7 % of the total lipids produced by this modified strain. By contrast, expression of the Claviceps purpurea hydroxylase CpFAH12 in this background resulted in a strain able to accumulate RA to 29 % of total lipids, and expression of an additional copy of CpFAH12 drove RA accumulation up to 35 % of total lipids. The co-expression of the C. purpurea or R. communis type II diacylglycerol acyltransferase (RcDGAT2 or CpDGAT2) had negative effects on RA accumulation in this yeast, with RA levels dropping to below 14 % of total lipids. Overexpression of the native Y. lipolytica PDAT acyltransferase (Lro1p) restored both TAG accumulation and RA levels. Thus, we describe the consequences of rerouting lipid metabolism in this yeast so as to develop a cell factory for RA production. The engineered strain is capable of accumulating RA to 43 % of its total lipids and over 60 mg/g of cell dry weight; this is the most efficient production of RA described to date.

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Acknowledgments

A. Beopoulos and J. Verbeke were supported by INRA. F. Bordes and M. Bressy were supported by INSA. This work was financially supported by DGCIS, Conseil Régional Midi-Pyrénées, and FEDER funding within the frame of the French national FUI project OLEOVISION.

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Authors and Affiliations

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

    A. Beopoulos & J. Verbeke

  2. Université de Toulouse; INSA, UPS, INP, LISBP, 135 Avenue de Rangueil, 31077, Toulouse, France

    F. Bordes, M. Guicherd, M. Bressy & A. Marty

  3. CNRS, UMR5504, 31400, Toulouse, France

    F. Bordes, M. Guicherd, M. Bressy & A. Marty

  4. INRA, UMR792 Ingénierie des Systèmes Biologiques et des Procédés, 31400, Toulouse, France

    F. Bordes, M. Guicherd, M. Bressy & A. Marty

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

    Jean-Marc Nicaud

  6. INRA, UMR1319, Micalis, AgroParistech, Centre de Biotechnologie Agro-Industrielle, 78850, Thiverval-Grignon, France

    Jean-Marc Nicaud

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  1. A. Beopoulos
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  2. J. Verbeke
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  3. F. Bordes
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  4. M. Guicherd
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  7. Jean-Marc Nicaud
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Correspondence to Jean-Marc Nicaud.

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A. Beopoulos, J. Verbeke, and F. Bordes contributed equally to this work.

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Beopoulos, A., Verbeke, J., Bordes, F. et al. Metabolic engineering for ricinoleic acid production in the oleaginous yeast Yarrowia lipolytica . Appl Microbiol Biotechnol 98, 251–262 (2014). https://doi.org/10.1007/s00253-013-5295-x

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  • DOI: https://doi.org/10.1007/s00253-013-5295-x

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