Engineering Escherichia coli for High-Yielding Hydroxytyrosol Synthesis from Biobased l-Tyrosine

Abstract

Hydroxytyrosol (HT) is a natural antioxidant with many associated health benefits. In this study, we established efficient enzymatic cascades for the synthesis of HT from biobased L-tyrosine. First, a dopamine-mediated route for HT production was investigated. The combination of native hydroxylase (HpaBC) from Escherichia coli and L-DOPA decarboxylase (DODC) from Pseudomonas putida could efficiently convert 5 mM L-tyrosine into dopamine with conversion above 90%. However, further incorporation of monoamine oxidase (MAO) from Micrococcus luteus and phenylacetaldehyde reductase (PAR) from Solanum lycopersicum only resulted in 3.47 mM HT with 69.4% conversion. Therefore, a second enzyme cascade that comprises HpaBC from E. coli, L-amino acid deaminase (LAAD) from Proteus mirabilis, α-keto acid decarboxylase (ARO10) from Saccharomyces cerevisiae, and PAR from S. lycopersicum was designed. This enzymatic route showed higher catalytic activity and efficiently synthesized HT. The 24.27 mM HT was obtained from 25 mM L-tyrosine with a high conversion of 97.1%, and 32.35 mM HT was produced using 50 mM L-tyrosine, which represents the highest HT titer using L-tyrosine as a substrate reported to date. In summary, we have developed a green and sustainable platform for efficient HT enzymatic synthesis.

Engineering a Synthetic Pathway for Tyrosol Synthesis in Escherichia coli

“…To enable the tyrosol production from lignin-derived p -CA, we further introduced FDC1 from S. cerevisiae , to convert p -CA into p -hydroxystyrene (Figure ). To boost tyrosol production, we also included an aldehyde reductase from S. lycopersicum with its function being experimentally validated in E. coli . For bioconverting p -CA to tyrosol, the overall enzyme cascade contains two plasmids for expressing four enzymes (FDC1-SMO-SOI-PAR) (Figure a).…”

“…To boost tyrosol production, we also included an aldehyde reductase from S. lycopersicum with its function being experimentally validated in E. coli. 30 For bioconverting p-CA to tyrosol, the overall enzyme cascade contains two plasmids for expressing four enzymes (FDC1-SMO-SOI-PAR) (Figure 2a).…”

“…Considering p -CA as a component of lignocellulose is relatively abundant in gramineous plants, we developed a novel biosynthetic pathway for the synthesis of tyrosol from p -CA. By introducing ferulic acid decarboxylase (FDC1) from S. cerevisiae , , SMO and SOI from P. putida , and phenylacetaldehyde reductase (PAR) from S. lycopersicum into E. coli , p -CA was efficiently converted to tyrosol, with the conversion rates ranging from 90% ∼ 99%. Therefore, our approach would offer a potential economic route for synthesis of tyrosol from renewable lignin-based biomass.…”

“…As p -hydroxystyrene is readily available from lignin derived p -coumaric acid ( p -CA), this synthetic route will offer alternative means for sustainable tyrosol production from agricultural wastes. In this study, ferulic acid decarboxylase (FDC1) from S. cerevisiae , , SMO and SOI from P. putida , and phenylacetaldehyde reductase (PAR) from Solanum lycopersicum were chosen for constructing the tyrosol biosynthetic pathway from p -CA, an abundant product from lignin-derived biomass. − To the best of our knowledge, our work represents the first-time to investigate SMO and SOI from P. putida for tyrosol synthesis from p -CA (a potential lignin-derived biomass), glucose, and glycerol.…”

De Novo Production of Hydroxytyrosol by Metabolic Engineering of Saccharomyces cerevisiae

“…Additionally, Zeng et al developed efficient enzymatic cascades in E. coli using 4-hydroxyphenylacetate 3-monooxygenase (EcH-paBC) from E. coli, L-amino acid deaminase from Proteus mirabilis, α-keto acid decarboxylase from Saccharomyces cerevisiae, and phenylacetaldehyde reductase from S. lycopersicum, culminating in up to 97.1% conversion rate of hydroxytyrosol from L-tyrosine. 13 Although progress has been made in the green production of hydroxytyrosol via bioconversion approaches, dependence on high-cost substrates still hampers the industrialization of these schemes. Nevertheless, synthetic biology continues to pave the 14 Since hydroxytyrosol can be produced either via the monophenol route (decarboxylation−deamination of tyrosine) or the diphenol route (hydroxylation of tyrosine to L-3,4-dihydroxyphenylalanine (DOPA)), Trantas et al combined both biosynthetic pathways in their chassis design and obtained significantly improved production efficiency of hydroxytyrosol.…”