Catabolism of acetosyringone and co-metabolic transformation of 2,4,6-trichlorophenol by a novel FAD-dependent...

Acetosyringone (AS), a prototypical syringyl-type monomer of lignin, functions as a model compound for the study of microbial catabolism of S-lignin-derived aromatics. In this study, we present the discovery of a novel metabolic pathway for AS catabolism, initiated by a previously uncharacterized FAD-dependent oxidoreductase, designated AsdA. In contrast to the sole previously documented AS funneling route, which entails side chain modification and conversion to syringic acid, AsdA catalyzes direct hydroxylation of the aromatic core. This represents a mechanistically distinct entry into central metabolism. The identification of this enzyme was achieved through metagenomic and functional analyses of a bacterial consortium enriched on AS as the sole carbon source. The consortium, predominantly comprising Pseudomonas rhizophila, exhibited co-metabolic transformation of the chlorinated pollutants 2,4,6-trichlorophenol (2,4,6-TCP) and 2,6-dichlorophenol. Subsequent functional assays substantiated the hypothesis that AsdA facilitates the transformation of both AS and 2,4,6-TCP. Induction assays employing a biosensor strain derived from the bacterial isolate Pseudomonas rhizophila AS1 confirmed AS-specific upregulation of the asd gene cluster. A survey of publicly available metagenomes has revealed that asdA is narrowly distributed but enriched in rhizosphere environments, pointing to its ecological significance. In summary, the present study unveils a hitherto unrecognized route for AS transformation and identifies an enzyme that exhibits dual functions in lignin-derived aromatic catabolism and environmental pollutant transformation. While the mechanisms underlying TCP degradation are well-established, the specific enzyme responsible for the conversion to 2,6-dichloro-p-hydroquinone had remained elusive-a knowledge gap that has now been addressed by AsdA.IMPORTANCEThe microbial conversion of lignin monomers is central to the global carbon cycle, yet pathways for syringyl-derived aromatics remain poorly resolved. Here, we identify AsdA, an enzyme initiating a previously unrecognized route for acetosyringone catabolism, providing new insight into how this abundant plant-derived compound is integrated into microbial metabolism. Beyond expanding the mechanistic diversity of lignin degradation, AsdA also catalyzes a key step in the transformation of the chlorinated pollutant 2,4,6-trichlorophenol, linking natural and anthropogenic compounds within a shared metabolic framework. The restricted yet rhizosphere-enriched distribution of asdA underscores its specialized role in plant-microbe interactions. By integrating enzyme function, microbial community context, and metagenomic distribution, we demonstrate how a single catalytic activity connects metabolic pathways and ecosystem processes, illustrating a multi-scale systems biology perspective on aromatic compound turnover.