Characterization of the phosphotriesterase capable of hydrolyzing aryl-organophosphate flame retardants

Abstract

A related group of phosphotriesters known as organophosphate flame retardants (OPFRs) has become emerging contaminants due to its worldwide use. The lack of an easily hydrolysable bond renders OPFRs inert to the well-known phosphotriesterases capable of hydrolyzing the neurotoxic organophosphates. An OPFRs phosphotriesterase gene stpte was cloned from plasmid pStJH of strain Sphingopyxis terrae subsp. terrae YC-JH3 and was heterologously expressed in Escherichia coli. The recombinant protein St-PTE was purified and analyzed. St-PTE showed the highest catalytic activity at pH 8.5 and 35 °C. The optimal substrate for St-PTE is triphenyl phosphate, with k_cat/K_m of 5.03 × 10⁶ M^-1 s^-1, two orders of magnitude higher than those of tricresyl phosphate (4.17 × 10⁴ M^-1 s^-1), 2-ethylhexyl diphenyl phosphate (2.03 × 10⁴ M^-1 s^-1) and resorcinol bis(diphenyl phosphate) (6.30 × 10⁴ M^-1 s^-1). St-PTE could break the P-O bond of tri-esters and convert aryl-OPFRs into their corresponding di-ester metabolites, including polymers of resorcinol bis(diphenyl phosphate). Mediated by transposase, the gene of OPFRs phosphotriesterase could be transferred horizontally among closely related strains of Sphingomonas, Sphingobium and Sphingopyxis. KEY POINTS: • St-PTE from Sphingopyxis terrae subsp. terrae YC-JH3 could hydrolyze aryl-OPFRs. • Metabolites of RBDPP hydrolyzed by phosphotriesterase were identified. • St-PTE could hydrolyze the P-O cleavage of dimer and trimer of RBDPP. • Phosphotriesterase genes transfer among Sphingomonadaceae mediated by transposase.

Keywords: 2-ethylhexyl diphenyl phosphate; Horizontal gene transfer; Organophosphate flame retardants; Phosphotriesterase; Resorcinol bis(diphenyl phosphate); Sphingopyxis.