Evaluation of a dye-decolorizing peroxidase from Comamonas serinivorans for lignin valorization potentials.

Although dye-decolourising peroxidases (DyPs) are well-known for lignin degradation, a comprehensive understanding of their mechanism remains unclear. Therefore, studying the mechanism of lignin degradation by DyPs is necessary for industrial applications and enzyme engineering. In this study, a dye-decolourising peroxidase (CsDyP) gene from C. serinivorans was heterologously expressed and studied for its lignin degradation potential. Molecular docking analysis predicted the binding of 2, 2-azino-bis (3-ethylbenzothiazoline-6-sulfonic acid) (ABTS), veratryl alcohol (VA), 2, 6-dimethylphenol (2, 6- DMP), guaiacol (GUA), and lignin to the substrate-binding pocket of CsDyP. Evaluation of the enzymatic properties showed that CsDyP requires pH 4.0 and 30 °C for optimal activity and has a high affinity for ABTS. In addition, CsDyP is stable over a wide range of temperatures and pH and can tolerate 5.0 mM organic solvents. Low NaCl concentrations promoted CsDyP activity. Further, CsDyP significantly reduced the chemical oxygen demand decolourised alkali lignin (AL) and milled wood lignin (MWL). CsDyP targets the ß-O-4, CO, and CC bonds linking lignin's G, S, and H units to depolymerize and produce aromatic compounds. Overall, this study delivers valuable insights into the lignin degradation mechanism of CsDyP, which can benefit its industrial applications and lignin valorization.

Genomics and biochemistry investigation on the metabolic pathway of milled wood and alkali lignin-derived aromatic metabolites of Comamonas serinivorans SP-35.

BACKGROUND: The efficient depolymerization and utilization of lignin are one of the most important goals for the renewable use of lignocelluloses. The degradation and complete mineralization of lignin by bacteria represent a key step for carbon recycling in land ecosystems as well. However, many aspects of this process remain unclear, for example, the complex network of metabolic pathways involved in the degradation of lignin and the catabolic pathway of intermediate aromatic metabolites. To address these subjects, we characterized the deconstruction and mineralization of lignin with milled wood lignin (MWL, the most representative molecule of lignin in its native state) and alkali lignin (AL), and elucidated metabolic pathways of their intermediate metabolites by a bacterium named Comamonas serinivorans SP-35. RESULTS: The degradation rate of MWL reached 30.9%, and its particle size range was decreased from 6 to 30 µm to 2-4 µm-when cultured with C. serinivorans SP35 over 7 days. FTIR analysis showed that the C-C and C-O-C bonds between the phenyl propane structures of lignin were oxidized and cleaved and the side chain structure was modified. More than twenty intermediate aromatic metabolites were identified in the MWL and AL cultures based on GC-MS analysis. Through genome sequencing and annotation, and from GC-MS analysis, 93 genes encoding 33 enzymes and 5 regulatory factors that may be involved in lignin degradation were identified and more than nine metabolic pathways of lignin and its intermediates were predicted. Of particular note is that the metabolic pathway to form the powerful antioxidant 3,4-dihydroxyphenylglycol is described for the first time in bacteria. CONCLUSION: Elucidation of the ß-aryl ether cleavage pathway in the strain SP-35 indicates that the ß-aryl ether catabolic system is not only present in the family of Sphingomonadaceae, but also other species of bacteria kingdom. These newly elucidated catabolic pathways of lignin in strain SP-35 and the enzymes responsible for them provide exciting biotechnological opportunities for lignin valorization in future.