Abstract
Background4-Hydroxycinnamyl alcohols are a class of natural plant secondary metabolites that include p-coumaryl alcohol, caffeyl alcohol, coniferyl alcohol and sinapyl alcohol, and have physiological, ecological and biomedical significance. While it is necessary to investigate the biological pathways and economic value of these alcohols, research is hindered because of their limited availability and high cost. Traditionally, these alcohols are obtained by chemical synthesis and plant extraction. However, synthesis by biotransformation with immobilized microorganisms is of great interest because it is environmentally friendly and offers high stability and regenerable cofactors. Therefore, we produced 4-hydroxycinnamyl alcohols using immobilized whole cells of engineered Escherichia coli as the biocatalyst.ResultsIn this study, we used the recombinant E. coli strain, M15–4CL1–CCR, expressing the fusion protein 4-coumaric acid: coenzyme A ligase and the cinnamoyl coenzyme A reductase and a recombinant E. coli strain, M15–CAD, expressing cinnamyl alcohol dehydrogenase from Populus tomentosa (P. tomentosa). High performance liquid chromatography and mass spectrometry showed that the immobilized whole cells of the two recombinant E. coli strains could effectively convert the phenylpropanoic acids to their corresponding 4-hydroxycinnamyl alcohols. Further, the optimum buffer pH and the reaction temperature were pH 7.0 and 30 °C. Under these conditions, the molar yield of the p-coumaryl alcohol, the caffeyl alcohol and the coniferyl alcohol was around 58, 24 and 60%, respectively. Moreover, the highly sensitive and selective HPLC–PDA–ESI–MSn method used in this study could be applied to the identification and quantification of these aromatic polymers.ConclusionsWe have developed a dual-cell immobilization system for the production of 4-hydroxycinnamyl alcohols from inexpensive phenylpropanoic acids. This biotransformation method is both simple and environmental-friendly, which is promising for the practical and cost effective synthesis of natural products.Graphical abstractBiotransformation process of phenylpropanoic acids by immobilized whole-cells
Highlights
We developed a novel, fast and highly efficient biological technique to convert diverse phenylpropanoic acids to their corresponding 4-hydroxycinnamyl alcohols using immobilized whole cells of recombinant E. coli as the biocatalyst, along with the recombinant E. coli strain M15–4-coumaric acid (4CL1)–cinnamoyl coenzyme A reductase (CCR) and the recombinant E. coli strain M15–cinnamyl alcohol dehydrogenase (CAD) expressing CAD from Populus
The ultraviolet absorption wavelength was set at 340 nm for the acids and aldehydes and 280 nm for the alcohols, because acids, aldehydes and alcohols cannot be completely detected under the same wavelength (Table 1)
Our results indicated that the immobilized whole cells of two recombinant E. coli were sufficiently active to convert diverse phenylpropanoic acids to 4-hydroxycinnamyl alcohols
Summary
The goals of this study were: (1) to establish a rapid HPLC–PDA–ESI–MSn method for the characterization of 4-hydroxycinnamyl alcohols; (2) to explore the feasibility of using immobilized whole cells of two recombinant E. coli to catalyze the conversion; (3) to investigate the optimum buffer pH and reaction temperature to improve the production; and (4) to evaluate the productivity of this novel biosynthesis system. The goal of this study was to produce 4-hydroxycinnamyl alcohols and simplify the production process using multi-strains immobilization to directly convert phenylpropanoic acids to 4-hydroxycinnamyl alcohols
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