title: Characterization of a novel thermostable β-glucosidase from a metagenomic library of termite gut.
abstract: A novel β-glucosidase-encoding gene, bgl-gs1, which was identified from a positive fosmid clone in a metagenomic library of the gut of Globitermes brachycerastes, [corrected] encodes a 455 amino acid polypeptide that contains a catalytic domain belonging to glycoside hydrolase family 1 (GH1). It was expressed in Escherichia coli BL21 (DE3) and the expression product showed a molecular mass of ∼51.7 kDa by SDS-PAGE. The optimal temperature and pH for the activity of the purified recombinant enzyme Bgl-gs1 with p-nitrophenyl-β-D-glucoside (pNPG) were 90°C and 6.0, respectively. The specific activities of Bgl-gs1 on pNPG and salicin were 110 and 14U/mg of protein, respectively, and its K(m) values were 0.18 and 2.59 mM, respectively. The residual activity of Bgl-gs1 was maintained above 70% after the recombinant enzyme was incubated at 75°C and pH 6.0 for 2h, and its half-life at 90°C was approximately 1h in the presence of 4mM pNPG. Bgl-gs1 showed synergistic effect with either a crude enzyme mixture of the fungal strain Trichoderma reesei Rut-C30 or a fusion protein (TcE1) created from the cellobiohydrolase cbh1 gene of T. reesei and endoglucanase from Acidothermus cellulolyticus; 87 and 137% increases in hydrolytic efficiency were noted on microcrystalline cellulose, respectively. These results suggest that the thermostable β-glucosidase Bgl-gs1 is a likely candidate for industrial applications.
authors.affiliation: Key Laboratory of Synthetic Biology, Institute of Plant Physiology and Ecology, Shanghai Institutes for Biological Sciences, Chinese Academy of Sciences, Shanghai 200032, China.
authors.forename: Qianfu
authors.initials: Q
authors.lastname: Wang
authors.forename: Changli
authors.initials: C
authors.lastname: Qian
authors.forename: Xiao-Zhou
authors.initials: XZ
authors.lastname: Zhang
authors.forename: Ning
authors.initials: N
authors.lastname: Liu
authors.forename: Nin
authors.initials: N
authors.lastname: Liu
authors.forename: Xing
authors.initials: X
authors.lastname: Yan
authors.forename: Zhihua
authors.initials: Z
authors.lastname: Zhou
mesh_terms: D000818:Animals; D001483:Base Sequence; D001709:Biotechnology; D020134:Catalytic Domain; D002482:Cellulose / Q000378:metabolism; D004247:DNA / Q000235:genetics; D004064:Digestive System / Q000201:enzymology; D004795:Enzyme Stability; D015723:Gene Library; D017344:Genes, Insect; D006358:Hot Temperature; D006868:Hydrolysis; D019476:Insect Proteins / Q000737:chemistry* / Q000235:genetics / Q000378:metabolism*; D020049:Isoptera / Q000201:enzymology* / Q000235:genetics; D007700:Kinetics; D054892:Metagenome; D017434:Protein Structure, Tertiary; D011994:Recombinant Proteins / Q000737:chemistry / Q000235:genetics / Q000378:metabolism; D013379:Substrate Specificity; D014242:Trichoderma / Q000201:enzymology; D001617:beta-Glucosidase / Q000737:chemistry* / Q000235:genetics / Q000378:metabolism*
chemical_list: D019476:Insect Proteins; D011994:Recombinant Proteins; D002482:Cellulose; D004247:DNA; D001617:beta-Glucosidase