Establishment of a yeast platform strain for production of p-coumaric acid through metabolic engineering of aromatic amino acid biosynthesis.
Aromatic amino acids are precursors of numerous plant secondary metabolites with diverse biological functions. Many of these secondary metabolites are already being used as active pharmaceutical or nutraceutical ingredients, and there are numerous exploratory studies of other compounds with promising applications. p-Coumaric acid is derived from aromatic amino acids and, besides being a valuable chemical building block, it serves as precursor for biosynthesis of many secondary metabolites, such as polyphenols, flavonoids, and some polyketides. Here we developed a p-coumaric acid-overproducing Saccharomyces cerevisiae platform strain. First, we reduced by-product formation by knocking out phenylpyruvate decarboxylase ARO10 and pyruvate decarboxylase PDC5. Second, different versions of feedback-resistant DAHP synthase and chorismate mutase were overexpressed. Finally, we identified shikimate kinase as another important flux-controlling step in the aromatic amino acid pathway by overexpressing enzymes from Escherichia coli, homologous to the pentafunctional enzyme Aro1p and to the bifunctional chorismate synthase-flavin reductase Aro2p. The highest titer of p-coumaric acid of 1.93 ± 0.26 g L(-1) was obtained, when overexpressing tyrosine ammonia-lyase TAL from Flavobacterium johnsoniaeu, DAHP synthase ARO4(K229L), chorismate mutase ARO7(G141S) and E. coli shikimate kinase II (aroL) in Δpdc5Δaro10 strain background. To our knowledge this is the highest reported titer of an aromatic compound produced by yeast. The developed S. cerevisiae strain represents an attractive platform host for production of p-coumaric-acid derived secondary metabolites, such as flavonoids, polyphenols, and polyketides.
The Novo Nordisk Foundation Center for Biosustainability, Technical University of Denmark, Kogle allé 6, 2970 Hørsholm, Denmark.
Angelica
A
Rodriguez
The Novo Nordisk Foundation Center for Biosustainability, Technical University of Denmark, Kogle allé 6, 2970 Hørsholm, Denmark.
Kanchana R
KR
Kildegaard
The Novo Nordisk Foundation Center for Biosustainability, Technical University of Denmark, Kogle allé 6, 2970 Hørsholm, Denmark.
Mingji
M
Li
The Novo Nordisk Foundation Center for Biosustainability, Technical University of Denmark, Kogle allé 6, 2970 Hørsholm, Denmark.
Irina
I
Borodina
The Novo Nordisk Foundation Center for Biosustainability, Technical University of Denmark, Kogle allé 6, 2970 Hørsholm, Denmark; Department of Biology and Biological Engineering, Chalmers University of Technology, Kemivägen 10, SE412 96 Gothenburg, Sweden. Electronic address: nielsenj@chalmers.se.
Jens
J
Nielsen
D010723:3-Deoxy-7-Phosphoheptulonate Synthase / Q000037:antagonists & inhibitors; D024322:Amino Acids, Aromatic / Q000096:biosynthesis*; D002826:Chorismate Mutase / Q000037:antagonists & inhibitors; D003373:Coumaric Acids / Q000378:metabolism*; D060847:Metabolic Engineering; D017853:Phosphotransferases (Alcohol Group Acceptor) / Q000502:physiology; D011422:Propionates; D011767:Pyruvate Decarboxylase / Q000235:genetics; D012441:Saccharomyces cerevisiae / Q000235:genetics / Q000378:metabolism*
D024322:Amino Acids, Aromatic; D003373:Coumaric Acids; D011422:Propionates; D010723:3-Deoxy-7-Phosphoheptulonate Synthase; D017853:Phosphotransferases (Alcohol Group Acceptor); C022642:shikimate kinase; D011767:Pyruvate Decarboxylase; D002826:Chorismate Mutase; C495469:p-coumaric acid
Saccharomyces cerevisiae; Shikimate kinase; p-Coumaric acid