title: Expression and characterization of two β-galactosidases from Klebsiella pneumoniae 285 in Escherichia coli and their application in the enzymatic synthesis of lactulose and 1-lactulose.
abstract: The two genes lacZ1 and lacZ2 from Klebsiella pneumoniae 285, encoding β-galactosidase isoenzymes II and III (KpBGase-II and -III), were each cloned downstream of a T7 promoter for expression in Escherichia coli BL21(DE3), and the resulting recombinant enzymes were characterized in detail. The optimum temperature and pH value of KpBGase-II were 40 °C and 7.5, and those of KpBGase-III were 50 °C and 8.0, respectively. KpBGase-III was more stable than KpBGase-II at higher temperature (>60°C). Both β-galactosidases were more active towards o-nitrophenyl-β- D-galactopyranoside as compared to lactose. The enzymatic synthesis of lactulose and 1-lactulose catalyzed by KpBGase-II and KpBGase-III was investigated. Using 400 g/L lactose and 200 g/L fructose as substrates, the resulting lactulose and 1-lactulose yields with KpBGase-II were 6.2 and 42.3 g/L, while those with KpBGase-III were 5.1 and 23.8 g/L, respectively. KpBGase-II has a potential for the production of 1-lactulose from lactose and fructose. Like other β-galactosidases, the two isozymes catalyze the transgalactosylation in the presence of fructose establishing the β-(1→1) linkage.
authors: He
authors: H
authors: Wang
authors: Ruijin
authors: R
authors: Yang
authors: Xiaoyan
authors: X
authors: Jiang
authors: Xiao
authors: X
authors: Hua
authors: Wei
authors: W
authors: Zhao
authors: Wenbin
authors: W
authors: Zhang
authors: Xuan
authors: X
authors: Chen
mesh_terms: D004926:Escherichia coli; D005632:Fructose / Q000737:chemistry; D015971:Gene Expression Regulation, Enzymologic; D007527:Isoenzymes / Q000096:biosynthesis* / Q000235:genetics; D007711:Klebsiella pneumoniae / Q000201:enzymology* / Q000235:genetics; D007785:Lactose / Q000737:chemistry; D007792:Lactulose / Q000096:biosynthesis* / Q000737:chemistry; D009598:Nitrophenylgalactosides / Q000737:chemistry / Q000378:metabolism; D013696:Temperature; D001616:beta-Galactosidase / Q000096:biosynthesis* / Q000235:genetics
chemical_list: D007527:Isoenzymes; C055012:2-nitrophenylgalactoside; D009598:Nitrophenylgalactosides; D005632:Fructose; D007792:Lactulose; D001616:beta-Galactosidase; D007785:Lactose