Caffeic acid is a plant secondary metabolite and its biological synthesis has attracted increased attention due to its beneficial effects on human health. In this study, Escherichia coli was engineered for the production of caffeic acid using tyrosine as the initial precursor of the pathway. The pathway design included tyrosine ammonia lyase (TAL) from Rhodotorula glutinis to convert tyrosine to p-coumaric acid and 4-coumarate 3-hydroxylase (C3H) from Saccharothrix espanaensis or cytochrome P450 CYP199A2 from Rhodopseudomonas palustris to convert p-coumaric acid to caffeic acid. The genes were codon-optimized and different combinations of plasmids were used to improve the titer of caffeic acid. TAL was able to efficiently convert 3mM of tyrosine to p-coumaric acid with the highest production obtained being 2.62mM (472mg/L). CYP199A2 exhibited higher catalytic activity towards p-coumaric acid than C3H. The highest caffeic acid production obtained using TAL and CYP199A2 and TAL and C3H was 1.56mM (280mg/L) and 1mM (180mg/L), respectively. This is the first study that shows caffeic acid production using CYP199A2 and tyrosine as the initial precursor. This study suggests the possibility of further producing more complex plant secondary metabolites like flavonoids and curcuminoids.
Keywords: Biosynthesis; Caffeic acid; E. coli; Synthetic biology; Tyrosine; p-Coumaric acid.
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