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

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• 作者：A. Beopoulos (1)
  J. Verbeke (1)
  F. Bordes (2) (3) (4)
  M. Guicherd (2) (3) (4)
  M. Bressy (2) (3) (4)
  A. Marty (2) (3) (4)
  Jean-Marc Nicaud (5) (6)
  
• 关键词：Ricinoleic acid ; Oleaginous yeast ; Yarrowia lipolytica ; Lipid metabolism ; Metabolic engineering ; Acyltransferase
• 刊名：Applied Microbiology and Biotechnology
• 出版年：2014
• 出版时间：January 2014
• 年：2014
• 卷：98
• 期：1
• 页码：251-262
• 全文大小：853 KB
• 作者单位：A. Beopoulos (1)
  J. Verbeke (1)
  F. Bordes (2) (3) (4)
  M. Guicherd (2) (3) (4)
  M. Bressy (2) (3) (4)
  A. Marty (2) (3) (4)
  Jean-Marc Nicaud (5) (6)
  
  1. INRA, UMR1319, Micalis, 78352, Jouy-en-Josas, France
  2. Université de Toulouse; INSA, UPS, INP, LISBP, 135 Avenue de Rangueil, 31077, Toulouse, France
  3. CNRS, UMR5504, 31400, Toulouse, France
  4. INRA, UMR792 Ingénierie des Systèmes Biologiques et des Procédés, 31400, Toulouse, France
  5. CNRS, Micalis, 78352, Jouy-en-Josas, France
  6. INRA, UMR1319, Micalis, AgroParistech, Centre de Biotechnologie Agro-Industrielle, 78850, Thiverval-Grignon, France
  
• ISSN：1432-0614

文摘

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 ?2 desaturase (Fad2p). We also expressed the Ricinus communis ?2 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.