Title:Construction and Evaluation of Peptide-Linked Lactobacillus brevis β-Galactosidase Heterodimers
Volume: 28
Issue: 2
Author(s): Yuan-Yuan Han, Hai-Yun Yue, Xiao-Yang Zhang, Yong-Mei Lyu, Li Liu* Josef Voglmeir*
Affiliation:
- Glycomics and Glycan Bioengineering Research Center (GGBRC), College of Food Science and Technology, Nanjing Agricultural University, Nanjing,China
- Glycomics and Glycan Bioengineering Research Center (GGBRC), College of Food Science and Technology, Nanjing Agricultural University, Nanjing,China
Keywords:
β-galactosidase, lactobacillus, fusion protein, peptide linkers, lactose hydrolysis, subunit co-expression.
Abstract:
Background: β-galactosidases are enzymes that are utilized to hydrolyze lactose into
galactose and glucose, and are is widely used in the food industry.
Objective: We describe the recombinant expression of an unstudied, heterodimeric β-galactosidase
originating from Lactobacillus brevis ATCC 367 in Escherichia coli. Furthermore, six different
constructs, in which the two protein subunits were fused with different peptide linkers, were also investigated.
Methods: The heterodimeric subunits of the β-galactosidase were cloned in expressed in various expression
constructs, by using either two vectors for the independent expression of each subunit, or
using a single Duet vector for the co-expression of the two subunits.
Results: The co-expression in two independent expression vectors only resulted in low β-galactosidase
activities, whereas the co-expression in a single Duet vector of the independent and fused subunits
increased the β-galactosidase activity significantly. The recombinant β-galactosidase showed
comparable hydrolyzing properties towards lactose, N-acetyllactosamine, and pNP-β-D-galactoside.
Conclusion: The usability of the recombinant L. brevis β-galactosidase was further demonstrated
by the hydrolysis of human, bovine, and goat milk samples. The herein presented fused β-galactosidase
constructs may be of interest for analytical research as well as in food- and biotechnological
applications.