Immobilized β-galactosidase BgaC from Bifidobacterium adolescentis retains stability and activity during repeated cycles of use

Abstract

β-Galactosidase enzymes catalyze the hydrolysis of terminal non-reducing β-D-galactose residues in β-galactosides. These enzymes are important in producing lactose-free dairy products, reducing the lactose content of whey in dairy products, and for production of galactooligosaccharides (GOS) as prebiotic additives to infant formula. To use β-galactosidases in industrial settings, enzyme immobilization procedures are used to enhance their activity and stability and to minimize enzyme quantities and cost. In this study, recombinant Bifidobacterium adolescentis β-galactosidase BgaC was immobilized in calcium alginate and gelatin cross-linked with glutaraldehyde. The kinetic parameters and stability properties of immobilized BgaC were characterized in comparison with free soluble enzyme. The K_M for immobilized BgaC using ortho-nitrophenyl-β-galactoside (ONPG) was 810 ± 220 μM and the K_M of free BgaC was 2500 ± 3 μM. The k_cat and k_cat/K_M of immobilized BgaC were 802 s⁻¹ and 990 s⁻¹ mM⁻¹, respectively, compared to k_cat and k_cat/K_M values of 209 s⁻¹ and 84 s⁻¹ mM⁻¹, respectively, for free BgaC. Immobilized BgaC β-galactosidase was active at all tested pH (pH 4–10), while the free enzyme had decreased activity at pH < 5.5 and > 8.0. The immobilized enzyme had optimum activity at 40 °C, while the free enzyme was most active at 37 °C. In addition, immobilization enhanced acidic pH and temperature stability compared to the free enzyme. Reutilization of the BgaC beads was assessed and the enzyme maintained 69% activity after 12 rounds of reutilization. Therefore, the enhanced performance properties of immobilized BgaC make it a promising candidate for industrial applications.

• Bifidobacterium adolescentis β-galactosidase BgaC was successfully immobilized

• Immobilized BgaC has enhanced enzymatic activity and stability and allows recycling

• Sustained activity of immobilized BgaC is advantageous for industrial applications