Production of β-carotene and acetate in recombinant Escherichia coli with or without mevalonate pathway at different culture temperature or pH
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Natural β-carotene has received much attention as consumers have become more health conscious. Its production by various microorganisms including metabolically engineered Escherichia coli or Saccharomyces cerevisiae has been attempted. We successfully created a recombinant E. coli with an engineered whole mevalonate pathway in addition to β-carotene biosynthetic genes and evaluated the engineered cells from the aspects of metabolic balance between central metabolism and β-carotene production by comparison with conventional β-carotene producing recombinant E. coli (control) utilizing a native methylerythritol phosphate (MEP) pathway using bioreactor cultures generated at different temperatures or pHs. Better production of β-carotene was obtained in E. coli cultured at 37°C than at 25°C. A two-fold higher titer and 2.9-fold higher volumetric productivity were obtained in engineered cells compared with control cells. Notably, a marginal amount of acetate was produced in actively growing engineered cells, whereas more than 8 g/L of acetate was produced in control cells with reduced cell growth at 37°C. The data indicated that the artificial operon of the whole mevalonate pathway operated efficiently in redirecting acetyl-CoA into isopentenyl pyrophosphate (IPP), thereby improving production of β-carotene, whereas the native MEP pathway did not convert a sufficient amount of pyruvate into IPP due to endogenous feedback regulation. Engineered cells also produced lycopene with a reduced amount of β-carotene in weak alkaline cultures, consistent with the inhibition of lycopene cyclase.
Keywordsrecombinant Escherichia coli engineered whole mevalonate pathway bioreactor culture β-carotene acetate
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- 7.Saenge, C., B. Cheirsilp, T. T. Suksaroge, and T. Bourtoom (2011) Efficient concomitant production of lipids and carotenoids by oleaginous red yeast Rhodotorula glutinis cultured in palm oil mill effluent and application of lipids for biodiesel production. Biotechnol. Bioproc. Eng. 16: 23–33.CrossRefGoogle Scholar
- 8.Verwaal, R., J. Wang, J. -P. Meijnen, H. Visser, G. Sandmann, J. A. V. D. Berg, and A. J. J. V. Ooyen (2007) High-level production of beta-carotene in Saccharomyces cerevisiae by successive transformation with carotenogenic genes from Xanthophyllomyces dendrorhous. Appl. Environ. Microbiol. 73: 4342–4350.CrossRefGoogle Scholar
- 10.Kim, J. H., S. -W. Kim, D. Q. A. Nguyen, H. Li, S. B. Kim, Y. -G. Seo, J. -K. Yang, I. -Y. Chung, D. H. Kim, and C. -J. Kim (2009) Production of β-carotene by recombinant Escherichia coli with engineered whole mevalonate pathway in batch and fed-batch cultures. Biotechnol. Bioproc. Eng. 14: 559–564.CrossRefGoogle Scholar
- 18.Sambrook, J. and D. W. Russel (2010) Molecular Cloning: A Laboratory Manual. 3rd ed. Cold Spring Harbor Laboratory Press, Cold Spring Harbor, NY, USA.Google Scholar
- 19.Schierle, J., B. Pietsch, A. Ceresa, and C. Fizet (2004) Method for the determination of β-carotene in supplements and raw materials by reversed-phase liquid chromatography: Single laboratory validation. J. AOAC Int. 87: 1070–1082.Google Scholar
- 24.Roe, A. J., C. O’Byrns, D. Mclaggan, and I. R. Booth (2002) Inhibition of Escherichia coli growth by acetic acid: A problem with methionine biosynthesis and homocystein toxicity. Microbiol. 148: 2215–2222.Google Scholar
- 33.Zilberstein, D., V. Agmon, S. Schuldiner, and E. Padan (1984) Escherichia coli intracellular pH, membrane potential, and cell growth. J. Bacteriol. 158: 246–252.Google Scholar