Phenanthrene degradation by a flavoprotein monooxygenase from Phanerodontia chrysosporium

ABSTRACT Phenanthrene (PHEN), a polycyclic aromatic hydrocarbon (PAH), is degraded by white-rot fungi like Phanerochaete chrysosporium (the fungus has been renamed as Phanerodontia chrysosporium ). PHEN is metabolized by P. chrysosporium and transformed into various monohydroxylated and dihydroxylated products. These intermediates are further degraded by cleavage of the aromatic ring. However, the enzymes involved in PHEN conversion in P. chrysosporium remain largely unidentified. We aimed to identify and characterize the P. chrysosporium enzymes involved in the degradation of PHEN and its intermediates. Recombinant P. chrysosporium flavoprotein monooxygenase 11 (FPMO11), a homolog of the salicylate 1-monooxygenase from the naphthalene-degrading bacterium Pseudomonas putida G7, was overexpressed in Escherichia coli . FPMO11 catalyzes the oxidative decarboxylation of 1-hydroxy-2-naphthoate (1H2N) and 2-hydroxy-1-naphthoate (2H1N) to 1,2-dihydroxynaphthalene (1,2DHN). To the best of our knowledge, this is the first study to identify and characterize enzymes with 1H2N and 2H1N monooxygenase activities in members of the FPMO superfamily. Additionally, our search for a dioxygenase with the ability to catalyze the aromatic ring cleavage of 1,2DHN led to the identification of intradiol dioxygenase (IDD) 1 and IDD2 from P. chrysosporium , which catalyzes the ring cleavage of 1,2DHN. Thus, this study also identified, for the first time, intradiol 1,2DHN dioxygenase activity in members of the IDD superfamily. The findings highlight the unique substrate spectra of FPMO11 and IDDs, rendering them attractive candidates for biotechnological applications, especially mitigation of environmental and health risks associated with PAH pollution. IMPORTANCE Phenanthrene (PHEN), a polycyclic aromatic hydrocarbon (PAH), is a widely studied pollutant in environmental science and toxicology due to its presence in fossil fuels, tobacco smoke, and as a byproduct of incomplete combustion processes. White-rot fungi like P. chrysosporium can degrade PHEN through the production of extracellular oxidative enzymes. We investigated the properties of PHEN-degrading enzymes in P. chrysosporium , specifically one flavoprotein monooxygenase (FPMO11) and two intradiol dioxygenases (IDD1 and IDD2). Our findings indicate that the enzymes catalyze the aromatic ring cleavage of PHEN, using the intermediates as substrates, transforming them into less harmful and more biodegradable compounds. This could help reduce environmental pollution and mitigate health risks associated with PAH exposure. The potential of these enzymes for biotechnological applications is also highlighted, emphasizing their critical role in understanding PAH degradation by white-rot fungi.