Biochemical characterization of two novel β-glucosidase genes by metagenome expression cloning.
Two novel β-glucosidase genes designated as bgl1D and bgl1E, which encode 172- and 151-aa peptides, respectively, were cloned by function-based screening of a metagenomic library from uncultured soil microorganisms. Sequence analyses indicated that Bgl1D and Bgl1E exhibited lower similarities with some putative β-glucosidases. Functional characterization through high-performance liquid chromatography demonstrated that purified recombinant Bgl1D and Bgl1E proteins hydrolyzed D-glucosyl-β-(1-4)-D-glucose to glucose. Using p-nitrophenyl-β-D-glucoside as substrate, K(m) was 0.54 and 2.11 mM, and k(cat)/K(m) was 1489 and 787 mM(-1) min(-1) for Bgl1D and Bgl1E, respectively. The optimum pH and temperature for Bgl1D was pH 10.0 and 30°C, while the optimum values for Bgl1E were pH 10.0 and 25°C. Bgl1D exhibited habitat-specific characteristics, including higher activity in lower temperature and at high concentrations of AlCl(3) and LiCl. Bgl1D also displayed remarkable activity across a broad pH range (5.5-10.5), making it a potential candidate for industrial applications.
Jiang
Chengjian
C
Guangxi Key Laboratory of Subtropical Bioresources Conservation and Utilization, College of Life Science and Technology, Guangxi University, Nanning, Guangxi, People's Republic of China.
Li
Shuang-Xi
SX
Luo
Feng-Feng
FF
Jin
Ke
K
Wang
Qin
Q
Hao
Zhen-Yu
ZY
Wu
Lan-Lan
LL
Zhao
Gao-Chao
GC
Ma
Ge-Fei
GF
Shen
Pei-Hong
PH
Tang
Xian-Lai
XL
Wu
Bo
B
D000595:Amino Acid Sequence; D001483:Base Sequence; D003001:Cloning, Molecular / Q000379:methods; D015870:Gene Expression / Q000235:genetics; D054892:Metagenome / Q000235:genetics*; D008969:Molecular Sequence Data; D012988:Soil Microbiology; D001617:beta-Glucosidase / Q000737:chemistry* / Q000502:physiology*
D001617:beta-Glucosidase