Exploring Recombinant Flavonoid Biosynthesis in Metabolically Engineered Escherichia coli

Authors

  • Kevin T. Watts,

    1. Department of Biochemistry, Molecular Biology and Biophysics, University of Minnesota, 1479 Gortner Avenue, St. Paul, MN 55108, USA, Fax: (+1) 612-625-5780
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  • Pyung Cheon Lee Dr.,

    1. Department of Biochemistry, Molecular Biology and Biophysics, University of Minnesota, 1479 Gortner Avenue, St. Paul, MN 55108, USA, Fax: (+1) 612-625-5780
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  • Claudia Schmidt-Dannert Prof.

    1. Department of Biochemistry, Molecular Biology and Biophysics, University of Minnesota, 1479 Gortner Avenue, St. Paul, MN 55108, USA, Fax: (+1) 612-625-5780
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Abstract

Flavonoids are important plant-specific secondary metabolites synthesized from 4-coumaroyl coenzyme A (CoA), derived from the general phenylpropanoid pathway, and three malonyl-CoAs. The synthesis involves a plant type III polyketide synthase, chalcone synthase. We report the cloning and coexpression in Escherichia coli of phenylalanine ammonia lyase, cinnamate-4-hydroxylase, 4-coumarate:CoA ligase, and chalcone synthase from the model plant Arabidopsis thaliana. Simultaneous expression of all four genes resulted in a blockage after the first enzymatic step caused by the presence of nonfunctional cinnamate-4-hydroxylase. To overcome this problem we fed exogenous 4-coumaric acid to induced cultures. We observed high-level production of the flavanone naringenin as a result. We were also able to produce phloretin by feeding cultures with 3-(4-hydroxyphenyl)propionic acid. Feeding with ferulic or caffeic acid did not yield the corresponding flavanones. We have also cloned and partially characterized a new tyrosine ammonia lyase from Rhodobacter sphaeroides. Tyrosine ammonia lyase was substituted for phenylalanine ammonia lyase and cinnamate-4-hydroxylase in our E. coli clones and three different growth media were tested. After 48 h induction, high-level production (20.8 mg L−1) of naringenin in metabolically engineered E. coli was observed for the first time.

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