Bioconversion of volatile fatty acids into lipids by the oleaginous yeast Yarrowia lipolytica

Fontanille, Pierre; Kumar, Vinod; Christophe, Gwendoline; Nouaille, Régis; Larroche, Christian

doi:10.1016/j.biortech.2012.02.091

Cited by 269 publications

(170 citation statements)

References 32 publications

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“…Thus, we included the complementing of leucine and uracil biosynthetic capacity both singly and in tandem as targets for this multiplexing. Integrated expression cassettes were driven by our high-strength synthetic UAS1B 16 -TEF constitutive hybrid promoter 23 . Collectively, the combinatorial multiplexing of enzyme overexpressions, fatty-acid inhibition knockouts and other biosynthetic pathways resulted in 57 distinct genotypes that were analysed for lipogenesis capacity compared with the wild-type strain (Supplementary Table 1).…”

Section: Resultsmentioning

confidence: 99%

“…All Y. lipolytica episomal plasmids were centromeric, replicative vectors derived from plasmid pSl16-Cen1-1(227) 50 after it had been modified to include a multi-cloning site, a hrGFP green fluorescent reporter gene (pIRES-hrGFP, Agilent) driven by the strong UAS1B 16 -TEF promoter 23 and a cyc1 terminator 51 to create plasmid pMCS-UAS1B 16 -TEF-hrGFP. Integrative plasmids were derived from plasmids pUC-S1-UAS1B 16 -Leum or pUC-S1-UAS1B 16 -TEF 25 that contained 5 0 and 3 0 rDNA integrative sequences surrounding the following elements-(from 5 0 to 3 0 ) a uracil section marker surrounded by LoxP sites for marker retrieval, the strong UAS1B 16 -Leum or UAS1B 16 -TEF promoter, AscI and PacI restriction enzyme sites for gene insertion and a XPR2 minimal terminator. These integrative plasmids were also designed to contain two identical NotI restriction enzyme sites directly outside of the rDNA regions so that plasmid linearization would simultaneously remove E. coli pUC19-based DNA.…”

Section: Methodsmentioning

confidence: 99%

“…The following genes were PCR-amplified from Y. lipolytica PO1f gDNA and inserted into vector pMCS-UAS1B 16 -TEF-hrGFP in place of hrGFP with an AscI/PacI digest: AMPD, ACL subunit 1 (ACL1), ACL subunit 2 (ACL2), MAE1, DGA1, and DGA2 with primers JB387/388, JB402/404, JB405/407, AH020/021, JB911/912 and JB913/914, respectively. This formed plasmids pMCS-UAS1B 16 -TEF-AMPD, pMCS-UAS1B 16 -TEF-ACL1, pMCS-UAS1B 16 -TEF-ACL2, pMCS-UAS1B 16 -TEF-MAE1, pMCS-UAS1B 16 -TEF-DGA1 and pMCS-UAS1B 16 -TEF-DGA2.…”

Section: Methodsmentioning

confidence: 99%

“…The following genes were gelextracted from the previously constructed episomal expression vectors and inserted into vector pUC-S1-UAS1B 16 -TEF with an AscI/PacI digest: AMPD, ACL subunit 1 (ACL1), ACL subunit 2 (ACL2), MAE1, DGA1 and DGA2. This formed plasmids pUC-S1-UAS1B 16 -TEF-AMPD, pUC-S1-UAS1B 16 -TEF-ACL1, pUC-S1-UAS1B 16 -TEF-ACL2, pUC-S1-UAS1B 16 -TEF-MAE1, pUC-S1-UAS1B 16 -TEF-DGA1 and pUC-S1-UAS1B 16 -TEF-DGA2.…”

Section: Methodsmentioning

confidence: 99%

“…As an alternative, several groups have explored oleaginous organisms such as the fungus Yarrowia lipolytica, but total oil content and titres are still limited [12][13][14][15][16][17] . Yet, the genetic tractability of Y. lipolytica [18][19][20][21][22][23][24] coupled with its modest, innate de novo lipogenesis (B10-15% lipid content in wild type 13,14,[25][26][27] ) make it a potential candidate as a platform organism for superior lipid production.…”

mentioning

confidence: 99%

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Harnessing Yarrowia lipolytica lipogenesis to create a platform for lipid and biofuel production

et al. 2014

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Economic feasibility of biosynthetic fuel and chemical production hinges upon harnessing metabolism to achieve high titre and yield. Here we report a thorough genotypic and phenotypic optimization of an oleaginous organism to create a strain with significant lipogenesis capability. Specifically, we rewire Yarrowia lipolytica's native metabolism for superior de novo lipogenesis by coupling combinatorial multiplexing of lipogenesis targets with phenotypic induction. We further complete direct conversion of lipid content into biodiesel. Tri-level metabolic control results in saturated cells containing upwards of 90% lipid content and titres exceeding 25 g l À 1 lipids, which represents a 60-fold improvement over parental strain and conditions. Through this rewiring effort, we advance fundamental understanding of lipogenesis, demonstrate non-canonical environmental and intracellular stimuli and uncouple lipogenesis from nitrogen starvation. The high titres and carbon-source independent nature of this lipogenesis in Y. lipolytica highlight the potential of this organism as a platform for efficient oleochemical production.

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Section: Resultsmentioning

confidence: 99%