Cloning of a Novel Pyrethroid-Hydrolyzing Carboxylesterase Gene from Sphingobium sp. Strain JZ-1 and Characterization of the Gene Product

Abstract

A novel esterase gene, pytH, encoding a pyrethroid-hydrolyzing carboxylesterase was cloned from Sphingobium sp. strain JZ-1. The gene contained an open reading frame of 840 bp. Sequence identity searches revealed that the deduced enzyme shared the highest similarity with many ␣/␤-hydrolase fold proteins (20 to 24% identities). PytH was expressed in Escherichia coli BL21 and purified using Ni-nitrilotriacetic acid affinity chromatography. It was a monomeric structure with a molecular mass of approximately 31 kDa and a pI of 4.85. PytH was able to transform p-nitrophenyl esters of short-chain fatty acids and a wide range of pyrethroid pesticides, and isomer selectivity was not observed. No cofactors were required for enzyme activity.Pyrethroid pesticides are now the major class of insecticides used for insect control in agriculture and households as a replacement for more toxic organophosphorus pesticides, and their usage is continuing to grow (10). Although pyrethroid pesticides generally have lower acute oral mammalian toxicity than organophosphate insecticides, exposure to very high levels of pyrethroid pesticides might cause endocrine disruption, lymph node and spleen damage, and carcinogenesis (6, 12). In addition, most pyrethroid pesticides possess acute toxicity to some nontarget organisms, such as bees, fish, and aquatic invertebrates, often at concentrations of less than 0.5 g/kg (19,22). Great concerns have been raised about the persistence and degradation of pyrethroid pesticides in the environment.In general, pyrethroid pesticides are degraded by both abiotic and biotic pathways, including photooxidation, chemical oxidation, and biodegradation. Microorganisms play the most important role in degradation of pyrethroids in soils and sediments. Many pyrethroid-degrading microorganisms have been isolated from soils (13,16,24,27).The major routes of pyrethroid metabolism in pyrethroidresistant insects and pyrethroid-degrading microorganisms include oxidation by cytochrome P450s and ester hydrolysis by carboxylesterases (9). Carboxylesterases are a family of enzymes that are important in the hydrolysis of a large number of endogenous and xenobiotic ester-containing compounds, such as carbamates, organophosphorus pesticides, and pyrethroids. Carboxylesterases from Bacillus cereus SM3 (17), Aspergillus niger ZD11 (13), Nephotettix cincticeps (2), and mouse liver microsomes (23) hydrolyzing the carboxyl ester linkage of the pyrethroids were purified to homogeneity and characterized. Genes encoding the pyrethroid-hydrolyzing carboxylesterases from mouse liver microsomes and Klebsiella sp. strain ZD112 were cloned and functionally expressed (23,27).Pyrethroids differ from many other pesticides in that they contain one to three chiral centers; the chirality may arise from the acid moiety, the alcohol moiety, or both (Fig. 1). A pyrethroid compound therefore consists of two to eight isomers. Isomers of a chiral compound often differ from each other in biological properties. Isomer selectivity has been widely observe...

A Tetrahydrofolate-Dependent Methyltransferase Catalyzing the Demethylation of Dicamba in Sphingomonas sp. Strain Ndbn-20

“…The two recombinant vectors were then transformed into E. coli BL21(DE3). The two genes were induced, and the C-terminal His 6 -tagged proteins were purified using the method described by Wang et al (29). The molecular weights were determined by SDS-PAGE and Coomassie blue staining, and the protein concentrations were quantified by the Bradford method using bovine serum albumin as the standard (30,31).…”

Molecular Mechanism and Genetic Determinants of Buprofezin Degradation

“…In 2009, bifenthrin was listed in toxicity class II (moderately hazardous) by the World Health Organization (WHO). The previously reported biodegradation pathway of bifenthrin is initiated by the breaking of the carboxylesterase linkage (40), which is catalyzed by a pyrethroid-hydrolyzing carboxylesterase (41,42). By contrast, we showed that strain YL-1 degraded bifenthrin in a different way, implying the potential use of strain YL-1 in the dissipation of bifenthrin residue in the environment.…”

PbaR, an IclR Family Transcriptional Activator for the Regulation of the 3-Phenoxybenzoate 1′,2′-Dioxygenase Gene Cluster in Sphingobium wenxiniae JZ-1 T

“…Strain JZ-1 T can utilize pyrethroids, such as cypermethrin, cyhalothrin, deltamethrin, fenpropathrin, fenvalerate, permethrin, and bifenthrin, and their metabolite 3-PBA as the sole carbon and energy sources for growth (5,9). The esterase gene pytH, which is responsible for the first step of hydrolyzing pyrethroids, is constitutively expressed (5), while the expression of the 3-PBA catabolic gene cluster pbaA1A2BC is induced by its substrate 3-PBA (9).…”

“…The esterase gene pytH, which is responsible for the first step of hydrolyzing pyrethroids, is constitutively expressed (5), while the expression of the 3-PBA catabolic gene cluster pbaA1A2BC is induced by its substrate 3-PBA (9). PbaA1 and PbaA2 are the ␣ and ␤ subunits of the dioxygenase, respectively, PbaB is a ferredoxin component, and PbaC is a glutathione reductase (GR)-type reductase (9).…”