title: A novel family VIII carboxylesterase hydrolysing third- and fourth-generation cephalosporins.
abstract: A metagenomic library was constructed from a soil sample of spindle tree-rhizosphere. From this library, one clone with esterase activity was selected. The sequence analysis revealed an open reading frame (EstSTR1) encoded protein of 390 amino acids. EstSTR1 is a family VIII carboxylesterase and retains the S-X-X-K motif conserved in both family VIII carboxylesterases and class C β-lactamases. The estSTR1 gene was overexpressed in E. coli and the recombinant protein was purified by purified by metal chelating affinity chromatography and size-exclusion chromatography. EstSTR1 hydrolysed p-nitrophenyl esters, exhibited the highest activity toward p-nitrophenyl butyrate. Furthermore, EstSTR1 could hydrolyse third- and fourth-generation cephalosporins (cefotaxime and cefepime) as well as first-generation cephalosporin (cephalothin). Site-directed mutagenesis studies revealed that a catalytic residue, Ser71, of EstSTR1 plays an essential role in hydrolysing both antibiotics and p-nitrophenyl esters. We demonstrate that a metagenome-derived carboxylesterase displays β-lactam-hydrolysing activities toward third- and fourth-generation cephalosporins.
authors.affiliation: Marine Biotechnology Research Division, Korea Institute of Ocean Science and Technology, Ansan, 15627 Republic of Korea ; National Leading Research Laboratory of Drug Resistance Proteomics, Department of Biological Sciences, Myongji University, 116 Myongjiro, Yongin, Gyeonggido, 17058 Republic of Korea.
authors.forename: Jeong Ho
authors.initials: JH
authors.lastname: Jeon
authors.affiliation: Marine Biotechnology Research Division, Korea Institute of Ocean Science and Technology, Ansan, 15627 Republic of Korea ; Department of Marine Biotechnology, University of Science and Technology, Daejeon, 34113 Republic of Korea.
authors.forename: Hyun Sook
authors.initials: HS
authors.lastname: Lee
authors.affiliation: National Leading Research Laboratory of Drug Resistance Proteomics, Department of Biological Sciences, Myongji University, 116 Myongjiro, Yongin, Gyeonggido, 17058 Republic of Korea.
authors.forename: Jung Hun
authors.initials: JH
authors.lastname: Lee
authors.affiliation: Department of Agricultural Biotechnology, National Academy of Agricultural Science, RDA, Jeonju, 54875 Republic of Korea.
authors.forename: Bon-Sung
authors.initials: BS
authors.lastname: Koo
authors.affiliation: Department of Agricultural Biotechnology, National Academy of Agricultural Science, RDA, Jeonju, 54875 Republic of Korea.
authors.forename: Chang-Muk
authors.initials: CM
authors.lastname: Lee
authors.affiliation: National Leading Research Laboratory of Drug Resistance Proteomics, Department of Biological Sciences, Myongji University, 116 Myongjiro, Yongin, Gyeonggido, 17058 Republic of Korea.
authors.forename: Sang Hee
authors.initials: SH
authors.lastname: Lee
authors.affiliation: Marine Biotechnology Research Division, Korea Institute of Ocean Science and Technology, Ansan, 15627 Republic of Korea ; Department of Marine Biotechnology, University of Science and Technology, Daejeon, 34113 Republic of Korea.
authors.forename: Sung Gyun
authors.initials: SG
authors.lastname: Kang
authors.affiliation: Marine Biotechnology Research Division, Korea Institute of Ocean Science and Technology, Ansan, 15627 Republic of Korea ; Department of Marine Biotechnology, University of Science and Technology, Daejeon, 34113 Republic of Korea.
authors.forename: Jung-Hyun
authors.initials: JH
authors.lastname: Lee
keywords: Carboxylesterase; Extended-spectrum cephalosporins; Metagenome; β-Lactamase