Abstract
ABSTRACTThe genusPseudomonasis a prolific source of specialized metabolites with significant biological activities, including siderophores, antibiotics, and plant hormones. These molecules play pivotal roles in environmental interactions, influencing pathogenicity, inhibiting microorganisms, responding to nutrient limitation and abiotic challenges, and regulating plant growth. These properties mean that pseudomonads are candidates as biological control agents against plant pathogens. Multiple transposon-based screens have identified aPseudomonasbiosynthetic gene cluster (BGC) associated with potent antibacterial and antifungal activity that produces 7-hydroxytropolone (7-HT). In this study, we show that this BGC also makes 3,7-dihydroxytropolone (3,7-dHT), which has strong antimicrobial activity towardsStreptomyces scabies, a potato pathogen. Both molecules exhibit broad biological activities, suggesting roles in competitive soil and plant microbial communities. Through metabolomics and reporter assays, we unveil the involvement of cluster-situated genes in generating phenylacetyl-coenzyme A, a key precursor for tropolone biosynthesis via the phenylacetic acid catabolon. The clustering of these phenylacetic acid genes within tropolone BGCs is unusual in other Gram-negative bacteria. Our findings support the interception of phenylacetic acid catabolism via an enoyl-CoA dehydratase encoded in the BGC, as well as highlighting an essential biosynthetic role for a conserved thioesterase. Biochemical assays were used to show that this thioesterase functions after a dehydrogenation-epoxidation step catalysed by a flavoprotein. We use this information to identify diverse uncharacterised BGCs that encode proteins with homology to flavoproteins and thioesterases involved in tropolone biosynthesis. This study provides insights into tropolone biosynthesis inPseudomonas, laying the foundation for further investigations into the ecological role of tropolone production.
Publisher
Cold Spring Harbor Laboratory