High-Yielding Protocatechuic Acid Synthesis from L-Tyrosine in Escherichia coli

Protocatechuic acid (PCA) is an important drug intermediate with antibacterial and antioxidant properties. In this study, the whole-cell bioconversion of L-tyrosine into PCA was explored using artificial enzymatic cascades engineered in Escherichia coli (E. coli). In particular, the first biocatalytic route comprises L-amino acid deaminase (LAAD) from Proteus mirabilis, hydroxymandelate synthase (HmaS) from Amycolatopsis orientalis, two-component flavin-dependent monooxygenase (HpaBC) from E. coli, hydroxymandelate oxidase (HMO) from Streptomyces coelicolor, benzoylformate decarboxylase (BFD) from Pseudomonas putida (P. putida), and aldehyde dehydrogenase (ALDH) from Saccharomyces cerevisiae. Combining LAAD-HmaS-HpaBC resulted in efficient synthesis of 3,4-dihydroxymandelate (DHMA), which could be further converted to PCA by HMO-BFD-ALDH to a final conversion of 64.4%. The second route utilizes 4-hydroxybenzoate (HBA) hydroxylase (PobA from P. putida) to convert HBA into PCA. As the recombinant E. coli produced >99% conversion of L-tyrosine into HBA within 12 h, further incorporation of PobA resulted in complete conversion of HBA into PCA, reaching >99% conversion. In summary, the developed biocatalytic pathway has great potential to produce various high-valued fine chemicals such as 4-hydroxymandelate (HMA), DHMA, HBA, and PCA.