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Cluis C.P.,Concordia University at Montreal | Ekins A.,Concordia University at Montreal | Narcross L.,Concordia University at Montreal | Jiang H.,Concordia University at Montreal | And 3 more authors.
Metabolic Engineering | Year: 2011

In this work, Escherichia coli was engineered to produce a medically valuable cofactor, coenzyme Q 10 (CoQ 10), by removing the endogenous octaprenyl diphosphate synthase gene and functionally replacing it with a decaprenyl diphosphate synthase gene from Sphingomonas baekryungensis. In addition, by over-expressing genes coding for rate-limiting enzymes of the aromatic pathway, biosynthesis of the CoQ 10 precursor para-hydroxybenzoate (PHB) was increased. The production of isoprenoid precursors of CoQ 10 was also improved by the heterologous expression of a synthetic mevalonate operon, which permits the conversion of exogenously supplied mevalonate to farnesyl diphosphate. The over-expression of these precursors in the CoQ 10-producing E. coli strain resulted in an increase in CoQ 10 content, as well as in the accumulation of an intermediate of the ubiquinone pathway, decaprenylphenol (10P-Ph). In addition, the over-expression of a PHB decaprenyl transferase (UbiA) encoded by a gene from Erythrobacter sp. NAP1 was introduced to direct the flux of DPP and PHB towards the ubiquinone pathway. This further increased CoQ 10 content in engineered E. coli, but decreased the accumulation of 10P-Ph. Finally, we report that the combined over-production of isoprenoid precursors and over-expression of UbiA results in the decaprenylation of para-aminobenzoate, a biosynthetic precursor of folate, which is structurally similar to PHB. © 2011 Elsevier Inc.

Scott S.D.,Fermentation and Metabolic Engineering Group | Armenta R.E.,Fermentation and Metabolic Engineering Group | Berryman K.T.,Fermentation and Metabolic Engineering Group | Norman A.W.,Fermentation and Metabolic Engineering Group
Enzyme and Microbial Technology | Year: 2011

Glucose is the typical carbon source for producing microbial polyunsaturated fatty acids (PUFA) with single cell microorganisms such as thraustochytrids. We assessed the use of a fish oil derived glycerol by-product (raw glycerol), produced by a fish oil processing plant, as a carbon source to produce single cell oil rich in polyunsaturated fatty acids (PUFA), notably docosahexaenoic acid (DHA). These results were compared to those obtained when using analytical grade glycerol, and glucose. The thraustochytrid strain tested produced similar amounts of oil and PUFA when grown with both types of glycerol, and results were also similar to those obtained using glucose. After 6 days of fermentation, approximately 320. mg/g of oil, and 145. mg/g of PUFA were produced with all carbon sources tested. All oils produced by our strain were 99.95% in the triacylglycerol form. To date, this is the first report of using raw glycerol derived from fish oil for producing microbial triglyceride oil rich in PUFA. © 2010 Elsevier Inc.

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