Scientific Publications by FDA Staff

J Bacteriol 2014 Oct 1;196(19):3503-15

Pleiotropic and Epistatic Behavior of a Ring-Hydroxylating Oxygenase System in the Polycyclic Aromatic Hydrocarbon Metabolic Network from Mycobacterium vanbaalenii PYR-1.

Kweon O, Kim SJ, Kim DW, Kim JM, Kim HL, Ahn Y, Sutherland JB, Cerniglia CE

Despite the considerable knowledge of bacterial high-molecular-weight (HMW) polycyclic aromatic hydrocarbon (PAH) metabolism, the key enzyme(s) and their pleiotropic and epistatic behavior(s) responsible for low-molecular weight (LMW) PAHs in HMW PAH-metabolic networks remain poorly understood. In this study, a phenotype-based strategy, coupled with a spray plate method, selected a Mycobacterium vanbaalenii PYR-1 mutant (6G11) that degrades HMW PAHs but not LMW PAHs. Sequence analysis determined that the mutant was defective in pdoA2, encoding an aromatic ring-hydroxylating oxygenase (RHO). A series of metabolic comparison using high-performance liquid chromatography (HPLC) analysis revealed that the mutant had a lower rate of degradation of fluorene, anthracene, and pyrene. Unlike the wild-type, the mutant did not produce a color change in culture media containing fluorene, phenanthrene, and fluoranthene. An Escherichia coli expression experiment confirmed the ability of the Pdo system to oxidize biphenyl, the LMW PAHs naphthalene, phenanthrene, anthracene, and fluorene, and the HMW PAHs pyrene, fluoranthene, and benzo[a]pyrene, with the highest enzymatic activity toward three-ring PAHs. Structure analysis and PAH substrates docking simulations of the Pdo substrate-binding pocket rationalized the experimentally observed metabolic versatility on a molecular scale. Using information obtained in this study and from previous work, we constructed an RHO-centric functional map, allowing pleiotropic and epistatic enzymatic explanation of PAH metabolism. Taken together, the Pdo system is a RHO system with pleiotropic responsibility of LMW PAH-centric hydroxylation, and its epistatic functional contribution is also a crucial for metabolic quality and quantity of the PAH-MN.