β-Galactosidase with transgalactosylation activity from Lactobacillus fermentum K4

Abstract

The LacLM β-galactosidase of Lactobacillus fermentum K4 is encoded by 2 consecutive genes, lacL (large subunit) and lacM (small subunit), that share 17 overlapping nucleotides. Phylogenetic analysis revealed that this enzyme was closely related to other Lactobacillus β-galactosidases and provided significant insight into its common and distinct characteristics. We cloned both the lacL and lacM genes of L. fermentum K4 and heterologously expressed each in Escherichia coli, although the recombinant enzyme was only functional when both were expressed on the same plasmid. We evaluated the enzymatic properties of this species-specific LacLM β-galactosidase and discovered that it acts as both a hydrolase, bioconverting lactose into glucose and galactose, and a transgalactosylase, generating prebiotic galacto-oligosaccharides (GOS). The recombinant β-galactosidase showed a broad pH optimum and stability around neutral pH. The optimal temperature and Michaelis constant (K(m)) for the substrates o-nitrophenyl-β-D-galactopyranoside and lactose were, respectively, 40°C and 45 to 50°C and 1.31 mM and 27 mM. The enzyme activity was stimulated by some cations such as Na⁺, K⁺, and Mg²⁺. In addition, activity was also enhanced by ethanol (15%, wt/vol). The transgalactosylation activity of L. fermentum K4 β-galactosidase effectively and rapidly generated GOS, up to 37% of the total sugars from the reaction. Collectively, our results suggested that the β-galactosidase from L. fermentum K4 could be exploited for the formation of GOS.

Multifunctional properties of Lactobacillus plantarum strains WiKim83 and WiKim87 as a starter culture for fermented food

“…In particular, among LAB strains, the β ‐galactosidase activity of lactobacilli including Lactobacillus reuteri , Lactobacillus fermentum , Lactobacillus crispatus , and L . plantarum has been characterized (Kim & Rajagopal, ; Liu, Kong, et al, ; Son et al, ; Splechtna et al, ). The basic function of the starter culture is to convert the milk sugar lactose into an acid product and contribute to the preservation, digestibility, texture, and flavor of the fermented product (Hébert, Raya, Tailliez, & Giori, ; Husain, ).…”

“…β-Galactosidase possesses the important characteristics of hydrolysis and the transglycosylation of lactose (Bras, Fernandes, & Ramos, 2010). LAB strains with β-galactosidase activity can convert lactose into useful short-chain fatty acids and are widely used for lactose intolerance treatment and in the dairy industry (Naidu, Bidlack, & Clemens, 1999 has been characterized (Kim & Rajagopal, 2000;Liu, Kong, et al, 2011;Son et al, 2017;Splechtna et al, 2006). The basic function of the starter culture is to convert the milk sugar lactose into an acid product and contribute to the preservation, digestibility, texture, and flavor of the fermented product (Hébert, Raya, Tailliez, & Giori, 2000;Husain, 2010).…”

MGcV: the microbial genomic context viewer for comparative genome analysis

“…However, in most Lactobacilli the protein is encoded by two neighboring genes (probably the result of gene fission) and the active protein is a heterodimer [59]. It is the LacLM protein that is mostly exploited in biotechnological applications [60,61]. Like E. coli , various Lactobacilli have a second beta-galactosidase encoding gene, lacA .…”

Synthesis of Galactooligosaccharides in Milk and Whey: A Review

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