Characterization of a Novel β-Glucosidase-Like Activity from a Soil Metagenome

摘要

Wereport the cloning of novel β-glucosidase-like gene by function-based screening of metagenomic lib- rary from uncultured soil microorganisms. The gene was named bg11C and has an open reading frame of ,443 base pairs. It encodes 481 amino acid polypeptide with predicted molecular mass of about 57. kDa. The deduced amino acid sequence did not show any homology with known β-glucosidases. The putative β-glucosidase gene was subcloned into the pETBlue- vector and overexpressed in . coli Tuner (DE3) pLac the recombinant protein was purified to homogeneity. Functional characterization with high performance liquid chromatography method demonstrated that the recombinant Bgl1C protein hydrolyzed -gl11cosylβ+(-)--glucose to glucose. The maximum activity for Bgl1C protein occurred at pH . and 42℃ using -nitrophenyl-β--glucoside as the substrate. CaC1_2 concentration of mM was required for optimal activity. The putative β-glucosidase had an apparent K_m value of .19 mM, V_(max).value of .75 /mg and k_(eat) value of 316./min under the optimal reaction conditions. The biochemical characterization of Bgl1C has enlarged our understanding of the novel enzymes that can be isolated from the soil metagenome.