title: Identification and characterization of a novel thermostable pyrethroid-hydrolyzing enzyme isolated through metagenomic approach.
abstract: BACKGROUND
  Pyrethroid pesticides are broad-spectrum pest control agents in agricultural production. Both agricultural and residential usage is continuing to grow, leading to the development of insecticide resistance in the pest and toxic effects on a number of nontarget organisms. Thus, it is necessary to hunt suitable enzymes including hydrolases for degrading pesticide residues, which is an efficient "green" solution to biodegrade polluting chemicals. Although many pyrethroid esterases have consistently been purified and characterized from various resources including metagenomes and organisms, the thermostable pyrethroid esterases have not been reported up to the present.
  
  
  RESULTS
  In this study, we identified a novel pyrethroid-hydrolyzing enzyme Sys410 belonging to familyV esterases/lipases with activity-based functional screening from Turban Basin metagenomic library. Sys410 contained 280 amino acids with a predicted molecular mass (Mr) of 30.8 kDa and was overexpressed in Escherichia coli BL21 (DE3) in soluble form. The optimum pH and temperature of the recombinant Sys410 were 6.5 and 55°C, respectively. The enzyme was stable in the pH range of 4.5-8.5 and at temperatures below 50°C. The activity of Sys410 decreased a little when stored at 4°C for 10 weeks, and the residual activity reached 94.1%. Even after incubation at 25°C for 10 weeks, it kept 68.3% of its activity. The recombinant Sys410 could hydrolyze a wide range of ρ-nitrophenyl esters, but its best substrate is ρ-nitrophenyl acetate with the highest activity (772.9 U/mg). The enzyme efficiently degraded cyhalothrin, cypermethrin, sumicidin, and deltamethrin under assay conditions of 37°C for 15 min, with exceeding 95% hydrolysis rate.
  
  
  CONCLUSIONS
  This is the first report to construct metagenomic libraries from Turban Basin to obtain the thermostable pyrethroid-hydrolyzing enzyme. The recombinant Sys410 with broad substrate specificities and high activity was the most thermostable one of the pyrethroid-hydrolyzing esterases studied before, which made it an ideal candidate for the detoxification of pyrethroids.
authors.affiliation: School of life sciences, Sun Yat-sen University, Guangzhou 510275, P R China.
authors.forename: Xinjiong
authors.initials: X
authors.lastname: Fan
authors.forename: Xiaolong
authors.initials: X
authors.lastname: Liu
authors.forename: Rui
authors.initials: R
authors.lastname: Huang
authors.forename: Yuhuan
authors.initials: Y
authors.lastname: Liu
mesh_terms: D000595:Amino Acid Sequence; D003001:Cloning, Molecular; D004926:Escherichia coli / Q000378:metabolism; D004950:Esterases / Q000145:classification / Q000235:genetics / Q000378:metabolism*; D006863:Hydrogen-Ion Concentration; D006868:Hydrolysis; D007306:Insecticides / Q000378:metabolism*; D056186:Metagenomics; D008969:Molecular Sequence Data; D010802:Phylogeny; D055550:Protein Stability; D011722:Pyrethrins / Q000378:metabolism*; D011994:Recombinant Proteins / Q000235:genetics / Q000378:metabolism; D016415:Sequence Alignment; D013379:Substrate Specificity; D013696:Temperature
chemical_list: D007306:Insecticides; D011722:Pyrethrins; D011994:Recombinant Proteins; D004950:Esterases