title: Glycosylation of phenolic compounds by the site-mutated β-galactosidase from Lactobacillus bulgaricus L3.
abstract: β-Galactosidases can transfer the galactosyl from lactose or galactoside donors to various acceptors and thus are especially useful for the synthesis of important glycosides. However, these enzymes have limitations in the glycosylation of phenolic compounds that have many physiological functions. In this work, the β-galactosidase from Lactobacillus bulgaricus L3 was subjected to site-saturation mutagenesis at the W980 residue. The recombinant pET-21b plasmid carrying the enzyme gene was used as the template for mutation. The mutant plasmids were transformed into Escherichia coli cells for screening. One recombinant mutant, W980F, exhibited increased yield of glycoside when using hydroquinone as the screening acceptor. The enzyme was purified and the effects of the mutation on enzyme properties were determined in detail. It showed improved transglycosylation activity on novel phenolic acceptors besides hydroquinone. The yields of the glycosides produced from phenol, hydroquinone, and catechol were increased by 7.6% to 53.1%. Moreover, it generated 32.3% glycosides from the pyrogallol that could not be glycosylated by the wild-type enzyme. Chemical structures of these glycoside products were further determined by MS and NMR analysis. Thus, a series of novel phenolic galactosides were achieved by β-galactosidase for the first time. This was a breakthrough in the enzymatic galactosylation of the challenging phenolic compounds of great values.
authors: State Key Lab of Microbial Technology and National Glycoengineering Research Center, Shandong University, Jinan 250100, PR China.
authors: Lili
authors: L
authors: Lu
authors: State Key Lab of Microbial Technology and National Glycoengineering Research Center, Shandong University, Jinan 250100, PR China.
authors: Lijuan
authors: L
authors: Xu
authors: State Key Lab of Microbial Technology and National Glycoengineering Research Center, Shandong University, Jinan 250100, PR China.
authors: Yuchuan
authors: Y
authors: Guo
authors: State Key Lab of Microbial Technology and National Glycoengineering Research Center, Shandong University, Jinan 250100, PR China.
authors: Dayu
authors: D
authors: Zhang
authors: State Key Lab of Microbial Technology and National Glycoengineering Research Center, Shandong University, Jinan 250100, PR China.
authors: Tingting
authors: T
authors: Qi
authors: State Key Lab of Microbial Technology and National Glycoengineering Research Center, Shandong University, Jinan 250100, PR China.
authors: Lan
authors: L
authors: Jin
authors: State Key Lab of Microbial Technology and National Glycoengineering Research Center, Shandong University, Jinan 250100, PR China.
authors: Guofeng
authors: G
authors: Gu
authors: State Key Lab of Microbial Technology and National Glycoengineering Research Center, Shandong University, Jinan 250100, PR China.
authors: Li
authors: L
authors: Xu
authors: State Key Lab of Microbial Technology and National Glycoengineering Research Center, Shandong University, Jinan 250100, PR China.
authors: Min
authors: M
authors: Xiao
mesh_terms: D000595:Amino Acid Sequence; D006031:Glycosylation; D007778:Lactobacillus / Q000201:enzymology*; D008958:Models, Molecular; D008969:Molecular Sequence Data; D016297:Mutagenesis, Site-Directed; D009154:Mutation; D010636:Phenols / Q000378:metabolism*; D011487:Protein Conformation; D013379:Substrate Specificity; D001616:beta-Galactosidase / Q000737:chemistry / Q000235:genetics* / Q000378:metabolism*
chemical_list: D010636:Phenols; D001616:beta-Galactosidase