Characterization of a novel hyper-thermostable and chlorpyrifos-hydrolyzing carboxylesterase EstC: A representative of the new esterase family XIX.

Carboxylesterases have widely been used in a series of industrial applications, especially, the detoxification of pesticide residues. In the present study, EstC, a novel carboxylesterase from Streptomyces lividans TK24, was successfully heterogeneously expressed, purified and characterized. Phylogenetic analysis showed that EstC can be assigned as the first member of a novel family XIX. Multiple sequence alignment indicated that EstC has highly conserved structural features, including a catalytic triad formed by Ser155, Asp248 and His278, as well as a canonical Gly-His-Ser-Ala-Gly pentapeptide. Biochemical characterization indicated that EstC exhibited maximal activity at pH 9.0 (Tris-HCl buffer) and 55 degrees C. It also showed higher activity towards short-chain substrates, with the highest activity for p-nitrophenyl acetate (pNPA2) (K-m = 0.31 +/- 0.02 mM, k(cat)/K-m = 1923.35 +/- 9.62 s(-1) mM(-1)) compared to other pNP esters used in this experiment. Notably, EstC showed hyper-thermostability and good alkali stability. The activity of EstC had no significant changes when it was incubated under 55 degrees C for 100 h and reached half-life after incubation at 100 degrees C for 8 h. Beyond that, EstC also showed stability at pH ranging from 6.0 to 11.0 and about 90% residual activity still reserved after treatment at pH 8.0 or 9.0 for 26 h, especially. Furthermore, EstC had outstanding potential for bioremediation of chlorpyrifos-contaminated environment. The recombinant enzyme (0.5 U mL(-1)) could hydrolyze 79.89% chlorpyrifos (5 mg L-1) at 37 degrees C within 80 min. These properties will make EstC have a potential application value in various industrial productions and detoxification of chlorpyrifos residues.