A Key -demethylase in the Degradation of Guaiacol by PD630

Overview

Journal Appl Environ Microbiol

Specialties Environmental Health
Microbiology

Date 2023 Oct 6

PMID 37800939

Authors

Le Xue

Yiquan Zhao

Ling Li

Xinran Rao

Xinjie Chen

Fuying Ma

Hongbo Yu

Shangxian Xie

Affiliations

Soon will be listed here.

Abstract

PD630 is a high oil-producing strain with the ability to convert lignin-derived aromatics to high values, but limited research has been done to elucidate its conversion pathway, especially the upper pathways. In this study, we focused on the upper pathways and demethylation mechanism of lignin-derived aromatics metabolism by PD630. The results of the aromatic carbon resource utilization screening showed that PD630 had a strong degradation capacity to the lignin-derived methoxy-containing aromatics, such as guaiacol, 3,4-veratric acid, anisic acid, isovanillic acid, and vanillic acid. The gene of , which encodes cytochrome P450, showed significant up-regulation when PD630 grew on diverse aromatics. Deletion mutants of and its partner protein resulted in the strain losing the ability to grow on guaiacol, but no significant difference to the other aromatics. Only co-complementation alone of and restored the strain's ability to utilize guaiacol, demonstrating that both genes were equally important in the utilization of guaiacol. assays further revealed that GcoA could convert guaiacol and anisole to catechol and phenol, respectively, with the production of formaldehyde as a by-product. The study provided robust evidence to reveal the molecular mechanism of PD630 on guaiacol metabolism and offered a promising study model for dissecting the demethylation process of lignin-derived aromatics in microbes.IMPORTANCEAryl--demethylation is believed to be the key rate-limiting step in the catabolism of heterogeneous lignin-derived aromatics in both native and engineered microbes. However, the mechanisms of -demethylation in lignin-derived aromatic catabolism remain unclear. Notably, guaiacol, the primary component unit of lignin, lacks demonstration and illustration of the molecular mechanism of guaiacol -demethylation in lignin-degrading bacteria. This is the first study to illustrate the mechanism of guaiacol metabolism by PD630 as well as characterize the purified key -demethylase . This study provided further insight into the lignin metabolic pathway of PD630 and could guide the design of an efficient biocatalytic system for lignin valorization.

Citing Articles

Lignin valorization to bioplastics with an aromatic hub metabolite-based autoregulation system.

Zhao Y, Xue L, Huang Z, Lei Z, Xie S, Cai Z Nat Commun. 2024; 15(1):9288.

PMID: 39468081 PMC: 11519575. DOI: 10.1038/s41467-024-53609-3.

References

Fetherolf M, Levy-Booth D, Navas L, Liu J, Grigg J, Wilson A . Characterization of alkylguaiacol-degrading cytochromes P450 for the biocatalytic valorization of lignin. Proc Natl Acad Sci U S A. 2020; 117(41):25771-25778. PMC: 7568313. DOI: 10.1073/pnas.1916349117. View

Chen Y, Ding Y, Yang L, Yu J, Liu G, Wang X . Integrated omics study delineates the dynamics of lipid droplets in Rhodococcus opacus PD630. Nucleic Acids Res. 2013; 42(2):1052-64. PMC: 3902926. DOI: 10.1093/nar/gkt932. View

Lubbers R, Dilokpimol A, Visser J, Makela M, Hilden K, de Vries R . A comparison between the homocyclic aromatic metabolic pathways from plant-derived compounds by bacteria and fungi. Biotechnol Adv. 2019; 37(7):107396. DOI: 10.1016/j.biotechadv.2019.05.002. View

Ragauskas A, Beckham G, Biddy M, Chandra R, Chen F, Davis M . Lignin valorization: improving lignin processing in the biorefinery. Science. 2014; 344(6185):1246843. DOI: 10.1126/science.1246843. View

Cramer P . AlphaFold2 and the future of structural biology. Nat Struct Mol Biol. 2021; 28(9):704-705. DOI: 10.1038/s41594-021-00650-1. View

Kurosawa K, Boccazzi P, de Almeida N, Sinskey A . High-cell-density batch fermentation of Rhodococcus opacus PD630 using a high glucose concentration for triacylglycerol production. J Biotechnol. 2010; 147(3-4):212-8. DOI: 10.1016/j.jbiotec.2010.04.003. View

Mallinson S, Machovina M, Silveira R, Garcia-Borras M, Gallup N, Johnson C . A promiscuous cytochrome P450 aromatic O-demethylase for lignin bioconversion. Nat Commun. 2018; 9(1):2487. PMC: 6021390. DOI: 10.1038/s41467-018-04878-2. View

Wells Jr T, Ragauskas A . Biotechnological opportunities with the β-ketoadipate pathway. Trends Biotechnol. 2012; 30(12):627-37. DOI: 10.1016/j.tibtech.2012.09.008. View

Li F, Ma F, Zhao H, Zhang S, Wang L, Zhang X . A Lytic Polysaccharide Monooxygenase from a White-Rot Fungus Drives the Degradation of Lignin by a Versatile Peroxidase. Appl Environ Microbiol. 2019; 85(9). PMC: 6495745. DOI: 10.1128/AEM.02803-18. View

10.

DeLorenzo D, Diao J, Carr R, Hu Y, Moon T . An Improved CRISPR Interference Tool to Engineer . ACS Synth Biol. 2021; 10(4):786-798. DOI: 10.1021/acssynbio.0c00591. View

11.

Chen H, Chow M, Liu C, Lau A, Liu J, Eltis L . Vanillin catabolism in Rhodococcus jostii RHA1. Appl Environ Microbiol. 2011; 78(2):586-8. PMC: 3255756. DOI: 10.1128/AEM.06876-11. View

12.

Harwood C, Parales R . The beta-ketoadipate pathway and the biology of self-identity. Annu Rev Microbiol. 1996; 50:553-90. DOI: 10.1146/annurev.micro.50.1.553. View

13.

Wolf M, Hinchen D, DuBois J, McGeehan J, Eltis L . Cytochromes P450 in the biocatalytic valorization of lignin. Curr Opin Biotechnol. 2021; 73:43-50. DOI: 10.1016/j.copbio.2021.06.022. View

14.

Bugg T, Ahmad M, Hardiman E, Rahmanpour R . Pathways for degradation of lignin in bacteria and fungi. Nat Prod Rep. 2011; 28(12):1883-96. DOI: 10.1039/c1np00042j. View

15.

Ralph J, Lapierre C, Boerjan W . Lignin structure and its engineering. Curr Opin Biotechnol. 2019; 56:240-249. DOI: 10.1016/j.copbio.2019.02.019. View

16.

Liang Y, Wei Y, Jiao S, Yu H . A CRISPR/Cas9-based single-stranded DNA recombineering system for genome editing of PD630. Synth Syst Biotechnol. 2021; 6(3):200-208. PMC: 8365321. DOI: 10.1016/j.synbio.2021.08.001. View

17.

Bugg T, Rahmanpour R . Enzymatic conversion of lignin into renewable chemicals. Curr Opin Chem Biol. 2015; 29:10-7. DOI: 10.1016/j.cbpa.2015.06.009. View

18.

Kweon O, Kim S, Baek S, Chae J, Adjei M, Baek D . A new classification system for bacterial Rieske non-heme iron aromatic ring-hydroxylating oxygenases. BMC Biochem. 2008; 9:11. PMC: 2358900. DOI: 10.1186/1471-2091-9-11. View

19.

Machovina M, Mallinson S, Knott B, Meyers A, Garcia-Borras M, Bu L . Enabling microbial syringol conversion through structure-guided protein engineering. Proc Natl Acad Sci U S A. 2019; 116(28):13970-13976. PMC: 6628648. DOI: 10.1073/pnas.1820001116. View

20.

Zhou H, Xu Z, Cai C, Li J, Jin M . Deciphering the metabolic distribution of vanillin in Rhodococcus opacus during lignin valorization. Bioresour Technol. 2021; 347:126348. DOI: 10.1016/j.biortech.2021.126348. View