Caffeic acid production enhancement by engineering a phenylalanine over‐producing Escherichia coli strain

Abstract

Caffeic acid is a plant-specific phenylpropanoic acid with multiple health-improving effects reported, and its therapeutic derivatives have also been studied throughout the last decade. To meet its market need and achieve high-level production, microbial production of caffeic acid approaches have been developed in metabolically engineered Escherichia coli. In our previous work, we have established the first artificial pathway that realized de novo production of caffeic acid using E. coli endogenous 4-hydroxyphenylacetate 3-hydroxylase (4HP3H). In this work, we exploited the catalytic potential of 4HPA3H in the whole-cell bioconversion study and produced 3.82 g/L (461.12 mg/L/OD) caffeic acid from p-coumaric acid, a direct precursor. We further engineered a phenylalanine over-producer into a tyrosine over-producer and then introduced the artificial pathway. After adjusting the expression strategy and optimizing the inoculants timing, de novo production of caffeic acid reached 766.68 mg/L. Both results from the direct precursor and simple carbon sources represent the highest titers of caffeic acid from microbial production so far.

Production of curcuminoids from tyrosine by a metabolically engineered Escherichia coli using caffeic acid as an intermediate

“…This stepwise approach was tested due to the possible toxicity of p-coumaric and ferulic acids to the cells [12,[18][19][20][21][22], and because we found that this strategy improved the production of caffeic acid in our previous work (manuscript submitted). However, in the current study the two-step feeding was shown to be counterproductive, i.e.…”

Heterologous production of caffeic acid from tyrosine in Escherichia coli

“…To meet the 73 phenylpropanoids market needs, it is imperative to look for new green and environmental 74 production methods to replace the traditional extraction one. One way to achieve high 75 levels of caffeic acid is the production via engineered microorganisms, such as Escherichia 76 coli [8][9][10].…”

Heterologous Production of Curcuminoids

“…Some studies related to production of phenylpropanoic acids (140,141,(150)(151)(152) and flavonoids (153) have used tyrosine-or phenylalanine-overproducing strains, and all obtained better results with these strains than with the wild type. All of them used similar approaches, in which they overexpressed aroG fbr and tyrA fbr , and in some cases ppsA and tktA, and deleted tyrR (⌬tyrR).…”

“…In the last years, several studies have used the enzymes involved in the phenylpropanoid pathway to produce compounds other than curcuminoids, such as phenylpropanoic acids (coumaric acid, caffeic acid, and ferulic acid) (140,141,151,152,(154)(155)(156)(157), stilbenoids (155,158), and flavonoids (116,117,150,155,(159)(160)(161)(162). In the design and construction of those pathways, some enzymes from other organisms were used that could also be considered for the optimization of the heterologous production of curcuminoids.…”

“…This production was also obtained by alleviating feedback inhibition and redirecting carbon flux into tyrosine biosynthesis. Huang et al (151) explored the catalytic potential of 4HPA3H and were able to produce 3.82 g/liter caffeic acid from 3.5 g/liter p-coumaric acid in a wild-type E. coli strain. When using a tyrosine-overproducing strain, 767 mg/liter caffeic acid was produced in 72 h using shake flasks.…”