Characterization of a novel carboxylesterase with catalytic activity toward di(2-ethylhexyl) phthalate from a soil metagenomic library.

A novel carboxylesterase gene estyz5 was isolated from a soil metagenomic library. The recombinant enzyme EstYZ5 is 298 amino acids in length with a predicted molecular weight of 32 kDa. Sequence alignment and phylogenetic analysis revealed that EstYZ5 belongs to the hormone-sensitive lipase (HSL) family with a deduced catalytic triad of Ser144-Glu238-His268. EstYZ5 contains two conserved motifs, a pentapeptide motif GDSAG and a HGGG motif, which are typically found in members of the HSL family. Esterolytic activity of the recombinant enzyme was optimal at 30 °C and pH 8.0, and the kcat/Km value of the enzyme for the optimum substrate p-nitrophenyl butyrate was as high as 1272 mM·s. Importantly, EstYZ5 showed activity toward di(2-ethylhexyl) phthalate with complex side chains, which is rare for HSLs. Molecular docking simulations revealed that the catalytic triad and an oxyanion hole likely play vital roles in enzymatic activity and specificity. The phthalate-degrading activity of EstYZ5, combined with its high levels of esterolytic activity, render this new enzyme a candidate for biotechnological applications.