Role of Acyl-CoA Synthetase (ACS) from Dietzia aurantiaca Strain J3 and Glutathione (GSH) in Fluorotelomer Biotransformation

Despite the environmental persistence of per- and polyfluoroalkyl substances (PFAS), their microbial biotransformation pathways remain poorly characterized due to unidentified metabolites and a lack of direct enzymatic validation. This study examined the roles of acyl-CoA synthetase (ACS) and glutathione S-transferase (GST) from Dietzia aurantiaca strain J3, as well as glutathione (GSH), in the biotransformation of fluorotelomer compounds. Heterologously expressed and purified ACS and GST from strain J3 were evaluated for their catalytic activities toward selected fluorinated and nonfluorinated carboxylates. Results showed that J3 ACS catalyzed the formation of CoA adducts with long-chain fluorinated compounds, including 2H-perfluoro-2-octenoic acid (6:2 FTUCA) and 5:3 fluorotelomer unsaturated carboxylate (5:3 FTUCA), providing the first experimental evidence of CoA adduct formation for eight-carbon PFAS. Notably, no enzymatic activity was observed toward saturated analogues (5:3 FTCA and 6:2 FTCA) or PFOA, with kinetic analysis suggesting that unsaturation and chain length are critical determinants of enzyme reactivity. While GST exhibited no activity toward the tested fluorinated substrates, GSH adducts were observed with 5:3 FTUCA and 6:2 FTUCA via spontaneous, nonenzymatic thiol conjugation. Collectively, these findings support the hypothesis that β-oxidation-like pathways, initiated by CoA conjugation, and GSH-dependent conjugation reactions are both involved in the biotransformation of long-chain polyfluorinated carboxylates.