The Catabolic System of Acetovanillone and Acetosyringone in Sphingobium sp. Strain SYK-6 Useful for Upgrading Aromatic Compounds Obtained through Chemical Lignin Depolymerization.

Acetovanillone is a major aromatic monomer produced in oxidative/base-catalyzed lignin depolymerization. However, the production of chemical products from acetovanillone has not been explored due to the lack of information on the microbial acetovanillone catabolic system. Here, the acvABCDEF genes were identified as specifically induced genes during the growth of Sphingobium sp. strain SYK-6 cells with acetovanillone and these genes were essential for SYK-6 growth on acetovanillone and acetosyringone (a syringyl-type acetophenone derivative). AcvAB and AcvF produced in Escherichia coli phosphorylated acetovanillone/acetosyringone and dephosphorylated the phosphorylated acetovanillone/acetosyringone, respectively. AcvCDE produced in Sphingobium japonicum UT26S carboxylated the reaction products generated from acetovanillone/acetosyringone by AcvAB and AcvF into vanilloyl acetic acid/3-(4-hydroxy-3,5-dimethoxyphenyl)-3-oxopropanoic acid. To demonstrate the feasibility of producing cis,cis-muconic acid from acetovanillone, a metabolic modification on a mutant of Pseudomonas sp. strain NGC7 that accumulates cis,cis-muconic acid from catechol was performed. The resulting strain expressing vceA and vceB required for converting vanilloyl acetic acid to vanillic acid and aroY encoding protocatechuic acid decarboxylase in addition to acvABCDEF successfully converted 1.2 mM acetovanillone to approximately equimolar cis,cis-muconic acid. Our results are expected to help improve the yield and purity of value-added chemical production from lignin through biological funneling. IMPORTANCE In the alkaline oxidation of lignin, aromatic aldehydes (vanillin, syringaldehyde, and p-hydroxybenzaldehyde), aromatic acids (vanillic acid, syringic acid, and p-hydroxybenzoic acid), and acetophenone-related compounds (acetovanillone, acetosyringone, and 4'-hydroxyacetophenone) are produced as major aromatic monomers. Also, base-catalyzed depolymerization of guaiacyl lignin resulted in vanillin, vanillic acid, guaiacol, and acetovanillone as primary aromatic monomers. To date, microbial catabolic systems of vanillin, vanillic acid, and guaiacol have been well characterized, and the production of value-added chemicals from them has also been explored. However, due to the lack of information on the microbial acetovanillone and acetosyringone catabolic system, chemical production from acetovanillone and acetosyringone has not been achieved. This study elucidated the acetovanillone/acetosyringone catabolic system and demonstrates the potential of using these genes for the production of value-added chemicals from these compounds.

Direct catalytic oxidation of rice husk lignin with hydroxide nanorod-modified copper foam and muconate production by engineered Pseudomonas sp. NGC7.

For the direct alkaline oxidation of rice husk lignin, we developed a copper foam-based heterogeneous catalyst that offers advantages in its recovery after the reaction mixture. The depolymerized products were utilized for muconate production by an engineered Pseudomonas sp. NGC7-based strain. A hydroxide nanorod-modified copper foam was prepared by the surface oxidation of copper foam, followed by alkaline oxidation of rice husk lignin over the catalyst. The catalyst was easily separated from the cellulosic residues in the reaction mixture, and the residues were then recovered by filtration. The resulting lignin stream was composed of a variety of aromatic monomers containing p-hydroxyphenyl, guaiacyl, and syringyl compounds. The catabolic activity of Pseudomonas sp. NGC7 was demonstrated to be more suitable for muconate production from a mixture comprising 4-hydroxybenzoate (a typical p-hydroxyphenyl compound), vanillate (a guaiacyl compound), and syringate (a syringyl compound), owing to its natural ability to grow on syringate. Thus, it was applied to produce muconate from a rice husk lignin stream prepared through hydroxide nanorod-modified copper foam-catalyzed alkaline oxidation by conferring the genes responsible for converting the acetophenone derivatives to their corresponding aromatic acids and protocatechuate decarboxylase to an NGC7-based strain deficient in protocatechuate 3,4-dioxygenase and muconate cycloisomerase. As a result, the engineered NGC7-based muconate-producing strain produced muconate selectively from the rice husk lignin stream at 93.7 mol% yield.