title: Molecular cloning, purification and biochemical characterization of a novel pyrethroid-hydrolyzing carboxylesterase gene from Ochrobactrum anthropi YZ-1.
abstract: 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.
authors.affiliation: Laboratory of Biology, Graduate School, Chinese Academy of Agriculture Science, Beijing 100081, China.
authors.forename: Yi
authors.initials: Y
authors.lastname: Zhai
authors.forename: Kang
authors.initials: K
authors.lastname: Li
authors.forename: Jinlong
authors.initials: J
authors.lastname: Song
authors.forename: Yanhua
authors.initials: Y
authors.lastname: Shi
authors.forename: Yanchun
authors.initials: Y
authors.lastname: Yan
mesh_terms: D000595:Amino Acid Sequence; D001483:Base Sequence; D043182:Carboxylesterase / Q000737:chemistry / Q000235:genetics*; D002849:Chromatography, Gas; D003001:Cloning, Molecular; D017931:DNA Primers; D004926:Escherichia coli / Q000235:genetics; D005798:Genes, Bacterial; D006868:Hydrolysis; D008969:Molecular Sequence Data; D020621:Ochrobactrum anthropi / Q000201:enzymology / Q000235:genetics / Q000302:isolation & purification*; D016133:Polymerase Chain Reaction; D011722:Pyrethrins / Q000378:metabolism*; D017386:Sequence Homology, Amino Acid; D013379:Substrate Specificity
chemical_list: D017931:DNA Primers; D011722:Pyrethrins; D043182:Carboxylesterase