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Molecular cloning, purification and biochemical characterization of a novel pyrethroid-hydrolyzing carboxylesterase gene from Ochrobactrum anthropi YZ-1
- Zhai, Yi, Li, Kang, Song, Jinlong, Shi, Yanhua, Yan, Yanchun
- Journal of hazardous materials 2012 v.221-222 pp. 206-212
- Escherichia coli, Ochrobactrum anthropi, activated sludge, amino acid sequences, carboxylesterase, enzyme activity, genes, genomic libraries, molecular cloning, open reading frames, pH, pesticides, phylogeny, pyrethrins, sequence homology, substrate specificity, temperature
- Strain YZ-1 was isolated from activated sludge and identified as Ochrobactrum anthropi. This strain was capable of degrading pyrethroids pesticides, suggesting the presence of degrading enzymes. In the present study, a novel esterase gene pytZ was cloned from the genomic library of YZ-1 successfully. The pytZ contained an open reading frame of 606bp encoding a pyrethroid-hydrolyzing carboxylesterase. Deduced amino acid sequence showed moderate identities (39–59%) with most homologous carboxylesterase, except a putative carboxylesterase from O. anthropi ATCC 49188 with the highest identity of 85%. Phylogenetic analysis revealed that PytZ belonged to esterase VI family. The gene pytZ showed no any sequence similarity with reported pyrethroid-hydrolyzing genes and was a new pyrethroid-degrading gene. PytZ was expressed in Escherichia coli BL21 (DE3) and purified using Ni-NTA Fast Start. PytZ was able to degrade various pyrethroids. The optimal temperature and pH were 35°C and 7.5. This enzyme was very stable over a wide range of temperature and pH. No cofactors were required for enzyme activity. Broad substrate specificity, high enzyme activity, and the favorable stability make the PytZ a potential candidate for the detoxification of pyrethroid residues in biotechnological application.