title: Two promising alkaline β-glucosidases isolated by functional metagenomics from agricultural soil, including one showing high tolerance towards harsh detergents, oxidants and glucose.
abstract: New β-glucosidase activities were identified by screening metagenomic libraries constructed with DNA isolated from the topsoil of a winter wheat field. Two of the corresponding proteins, displaying an unusual preference for alkaline conditions, were selected for purification by Ni-NTA chromatography. AS-Esc6, a 762-amino-acid enzyme belonging to glycoside hydrolase family 3, proved to be a mesophilic aryl-β-glucosidase with maximal activity around pH 8 and 40 °C. A similar pH optimum was found for AS-Esc10, a 475-amino-acid GH1-family enzyme, but this enzyme remained significantly active across a wider pH range and was also markedly more stable than AS-Esc6 at pH greater than 10. AS-Esc10 was found to degrade cellobiose and diverse aryl glycosides, with an optimal temperature of 60 °C and good stability up to 50 °C. Unlike AS-Esc6, which showed a classically low inhibitory constant for glucose (14 mM), AS-Esc10 showed enhanced activity in the presence of molar concentrations of glucose. AS-Esc10 was highly tolerant to hydrogen peroxide and also to sodium dodecyl sulfate, this being indicative of kinetic stability. This unique combination of properties makes AS-Esc10 a particularly promising candidate whose potential in biotechnological applications is worth exploring further.
authors.affiliation: Microbiology and Genomics Unit, Gembloux Agro-Bio Tech, University of Liège, Avenue Maréchal Juin 6, 5030, Gembloux, Belgium, Sophie.Biver@ulg.ac.be.
authors.forename: Sophie
authors.initials: S
authors.lastname: Biver
authors.forename: Aurore
authors.initials: A
authors.lastname: Stroobants
authors.forename: Daniel
authors.initials: D
authors.lastname: Portetelle
authors.forename: Micheline
authors.initials: M
authors.lastname: Vandenbol
mesh_terms: D003902:Detergents / Q000378:metabolism; D005947:Glucose / Q000378:metabolism; D006863:Hydrogen-Ion Concentration; D007700:Kinetics; D056186:Metagenomics; D008969:Molecular Sequence Data; D011994:Recombinant Proteins / Q000737:chemistry / Q000235:genetics / Q000378:metabolism; D012987:Soil / Q000737:chemistry*; D013379:Substrate Specificity; D013696:Temperature; D001617:beta-Glucosidase / Q000737:chemistry / Q000235:genetics* / Q000302:isolation & purification* / Q000378:metabolism
chemical_list: D003902:Detergents; D011994:Recombinant Proteins; D012987:Soil; D001617:beta-Glucosidase; D005947:Glucose