title: Cloning, expression and structural stability of a cold-adapted β-galactosidase from Rahnella sp. R3.
abstract: A novel gene was isolated for the first time from a psychrophilic gram-negative bacterium Rahnella sp. R3. The gene encoded a cold-adapted β-galactosidase (R-β-Gal). Recombinant R-β-Gal was expressed in Escherichia coli BL21 (DE3), purified and characterized. R-β-gal belongs to the glycosyl hydrolase family 42. Circular dichroism spectrometry of the structural stability of R-β-Gal with respect to temperature indicated that the secondary structures of the enzyme were stable to 45°C. In solution, the enzyme was a homo-trimer and was active at temperatures as low as 4°C. The enzyme did not require the presence of metal ions to be active, but Mg(2+), Mn(2+), and Ca(2+) enhanced its activity slightly, whereas Fe(3+), Zn(2+) and Al(3+) appeared to inactive it. The purified enzyme displayed K(m) values of 6.5 mM for ONPG and 2.2mM for lactose at 4°C. These values were lower than the corresponding K(m)s reported for other cold-adapted β-Gals.
authors.affiliation: State Key Laboratory of Food Science and Technology, Jiangnan University, 214122 Wuxi, China; School of Food Science and Technology, Jiangnan University, 214122 Wuxi, China. Electronic address: yilin58299@163.com.
authors.forename: Yuting
authors.initials: Y
authors.lastname: Fan
authors.affiliation: State Key Laboratory of Food Science and Technology, Jiangnan University, 214122 Wuxi, China. Electronic address: huaxiao@jiangnan.edu.cn.
authors.forename: Xiao
authors.initials: X
authors.lastname: Hua
authors.affiliation: U.S. Department of Agriculture, Agriculture Research Service, Pacific West Area, Western Regional Research Center, Albany, CA 94710, United States.
authors.forename: Yuzhu
authors.initials: Y
authors.lastname: Zhang
authors.affiliation: School of Food Science and Technology, Jiangnan University, 214122 Wuxi, China.
authors.forename: Yinghui
authors.initials: Y
authors.lastname: Feng
authors.affiliation: School of Food Science and Technology, Jiangnan University, 214122 Wuxi, China.
authors.forename: Qiuyun
authors.initials: Q
authors.lastname: Shen
authors.affiliation: State Key Laboratory of Food Science and Technology, Jiangnan University, 214122 Wuxi, China.
authors.forename: Juan
authors.initials: J
authors.lastname: Dong
authors.affiliation: School of Food Science and Technology, Jiangnan University, 214122 Wuxi, China.
authors.forename: Wei
authors.initials: W
authors.lastname: Zhao
authors.affiliation: School of Food Science and Technology, Jiangnan University, 214122 Wuxi, China.
authors.forename: Wenbin
authors.initials: W
authors.lastname: Zhang
authors.affiliation: State Key Laboratory of Food Science and Technology, Jiangnan University, 214122 Wuxi, China.
authors.forename: Zhengyu
authors.initials: Z
authors.lastname: Jin
authors.affiliation: State Key Laboratory of Food Science and Technology, Jiangnan University, 214122 Wuxi, China; School of Food Science and Technology, Jiangnan University, 214122 Wuxi, China. Electronic address: yrj@jiangnan.edu.cn.
authors.forename: Ruijin
authors.initials: R
authors.lastname: Yang
mesh_terms: D003001:Cloning, Molecular; D003080:Cold Temperature; D004795:Enzyme Stability; D004926:Escherichia coli / Q000235:genetics; D006863:Hydrogen-Ion Concentration; D007700:Kinetics; D019216:Metals, Heavy; D020638:Rahnella / Q000201:enzymology* / Q000235:genetics; D011994:Recombinant Proteins / Q000737:chemistry* / Q000235:genetics / Q000302:isolation & purification / Q000378:metabolism*; D001616:beta-Galactosidase / Q000737:chemistry* / Q000235:genetics / Q000302:isolation & purification / Q000378:metabolism*
chemical_list: D019216:Metals, Heavy; D011994:Recombinant Proteins; D001616:beta-Galactosidase
keywords: Cold-adapted enzyme; Enzyme structure stability; Lactose hydrolysis; β-Galactosidase