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. 2021 Dec 1:3:100059.
doi: 10.1016/j.fochms.2021.100059. eCollection 2021 Dec 30.

Microbial synthesis of 4-hydroxybenzoic acid from renewable feedstocks

Yueyang Chen  1 , Yufen Chen  1 , Lijun Liu  1 , Yang Zhang  1 , Jifeng Yuan  1
Affiliations

Microbial synthesis of 4-hydroxybenzoic acid from renewable feedstocks

Yueyang Chen et al. Food Chem (Oxf). .

Abstract

4-Hydroxybenzoic acid (4HBA) and its esterified forms can be used as preservatives in the pharmaceutical and food industries. Here, we reported the establishment of a coenzyme-A (CoA) free multi-enzyme cascade in Escherichia coli to utilize biobased L-tyrosine for efficient synthesis of 4HBA. The multi-enzyme cascade contains L-amino acid deaminase from Proteus mirabilis, hydroxymandelate synthase from Amycolatopsis orientalis, (S)-mandelate dehydrogenase and benzoylformate decarboxylase from Pseudomonas putida, and aldehyde dehydrogenase from Saccharomyces cerevisiae. The whole-cell biocatalysis afforded the synthesis of 128 ± 1 mM of 4HBA (17.7 ± 0.1 g/L) from 150 mM L-tyrosine with > 85% conversion within 96 h. In addition, the artificial enzymatic cascade also allowed the synthesis of benzoic acid from 100 mM L-phenylalanine with a conversion ∼ 90%. In summary, our research offers a sustainable alternative for synthesizing 4HBA and benzoic acid from renewable feedstocks.

Keywords: 4-Hydroxybenzoic acid; Benzoic acid; L-Tyrosine; Multi-enzyme cascade; Whole-cell biocatalyst.

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Conflict of interest statement

The authors declare that they have no known competing financial interests or personal relationships that could have appeared to influence the work reported in this paper.

Figures

Fig. 1
Fig. 1
Schematic diagram of the natural and synthetic 4HBA biosynthetic routes. (A) Chorismate pyruvate-lyase (ubiC) mediated 4HBA in bacteria and the CoA-dependent 4HBA pathway in plants. E4P, D-erythrose-4-phosphate; PEP, phosphoenolpyruvate; DHAP, dihydroxyacetone phosphate; CHOR, chorismite; L-TYR, L-tyrosine; 4HBA, 4-hydroxybenzoic acid. (B) The synthetic CoA-independent 4HBA pathway. L-amino acid deaminase (LAAD) from P. mirabilis; HmaS, hydroxymandelate synthase (HmaS) from A. orientalis; (S)-mandelate dehydrogenase (SMDH) from P. putida; benzoylformate decarboxylase (BFD) from P. putida; aldehyde dehydrogenase (ALDH) from S. cerevisiae.
Fig. 2
Fig. 2
Synthesis of 4HBA via the CoA independent enzymatic route. (A) Schematic diagram of plasmids used for CoA-free synthesis of 4HBA. (B) Time course of whole-cell biotransformation of L-tyrosine (10 mM) to 4HBA. The reactions were conducted in KP buffer (200 mM, pH 8.0) at 30 °C with 10 g cdw/L resting cells of E. coli MG1655 RARE cells. The data are the mean values with standard deviations from triplicate experiments. (C) Representative HPLC result showing the 4HBA production. The retention time for 4HBA is 2.9 min. (D) HPLC result of the negative control. E. coli carrying empty plasmids under the same reaction condition was used as the negative control. The retention time for L-tyrosine is 1.7 min.
Fig. 3
Fig. 3
Optimizing the buffer pH conditions for enhanced production of 4HBA. (A) Dose effect of different L-tyrosine concentrations on 4HBA production at pH 8.0. (B) The effect of pH conditions on 4HBA productions. All the reactions were conducted at 30 °C with 10 g cdw/L resting cells of E. coli MG1655 RARE cells. The data are the mean values with standard deviations from triplicate experiments.
Fig. 4
Fig. 4
Synthesis of benzoic acid via the CoA-independent enzymatic route. Dose effect of different L-phenylalanine concentrations on benzoic acid productions. Time course of benzoic acid synthesis from 50 mM or 100 mM L-phenylalanine with 10 g cdw/L resting cells of E. coli MG1655 RARE cells. All the reactions were conducted in KP buffer (pH 9.0) at 30 °C. The data are the mean values with standard deviations from triplicate experiments.
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References

    1. Barker J.L., Frost J.W. Microbial synthesis of p-hydroxybenzoic acid from glucose. Biotechnology and Bioengineering. 2001;76(4):376–390. doi: 10.1002/bit.10160. - DOI - PubMed
    1. Chen Y., Fu B., Xiao G., Ko L.-Y., Kao T.-Y., Fan C., Yuan J. Bioconversion of lignin-derived feedstocks to muconic acid by whole-cell biocatalysis. ACS Food Science & Technology. 2021;1(3):382–387. doi: 10.1021/acsfoodscitech.1c0002310.1021/acsfoodscitech.1c00023.s001. - DOI
    1. Choi I.S., Cho E.J., Moon J.H., Bae H.J. Onion skin waste as a valorization resource for the by-products quercetin and biosugar. Food Chemistry. 2015;188:537–542. doi: 10.1016/j.foodchem.2015.05.028. - DOI - PubMed
    1. Choi K.Y., Wernick D.G., Tat C.A., Liao J.C. Consolidated conversion of protein waste into biofuels and ammonia using Bacillus subtilis. Metabolic Engineering. 2014;23:53–61. doi: 10.1016/j.ymben.2014.02.007. - DOI - PubMed
    1. Hazan R., Levine A., Abeliovich H. Benzoic acid, a weak organic acid food preservative, exerts specific effects on intracellular membrane trafficking pathways in Saccharomyces cerevisiae. Applied and Environmental Microbiology. 2004;70(8):4449–4457. doi: 10.1128/AEM.70.8.4449-4457.2004. - DOI - PMC - PubMed

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