PRODUCTION OF RECOMBINANT LACTASE FROM Lactobacillus helveticus VA FOR APPLICATION IN ADVANCED MILK PROCESSING TECHNOLOGIES

β-Galactosidases (lactases, EC 3.2.1.23) are widely applied in the production of lactose-free dairy products and whey processing. The aim of the study is to obtain recombinant β-galactosidase and to determine its biochemical and kinetic parameters. In the present work, eight strains were isolated from goat, sheep, mare, and camel milk, as well as cheeses and fermented dairy products from the Akmola and Almaty regions: Lactobacillus plantarum NAR, Lactobacillus plantarum VGM1, Lactobacillus paracasei VK2, Lactococcus lactis ZCM, Lactococcus lactis IS2, Lactobacillus curvatus ICH3, Lactobacillus rhamnosus ICH5, and Lactobacillus helveticus VA. Screening for lactase activity revealed that the cheese-isolated strain Lactobacillus helveticus VA exhibited the highest activity compared to the other strains, reaching 212.63 U/mg. The β-galactosidase gene from Lactobacillus helveticus was cloned into the pET-28c(+) vector, and the recombinant lactase rLacLM was produced via plasmid expression in the Escherichia coli BL21(DE3) strain. Using immobilized metal affinity chromatography (IMAC), rLacLM was purified from the induced recombinant culture. The recombinant β-galactosidase exhibited maximum activity at pH 6.5–7.0 and a temperature of 50°C. The specific activity of the enzyme was 76.1 U/mg, while the Km, max, kcat, and kcat/Km values were 21.76 mM, 1015 µM·min⁻¹, 141.48 s⁻¹, and 6.5 × 10³ M⁻¹·s⁻¹, respectively. Thin-layer chromatography confirmed efficient lactose hydrolysis yielding glucose and galactose. These results indicate that the recombinant β-galactosidase from Lactobacillus helveticus VA possesses biochemical and catalytic characteristics compatible with milk processing conditions, rendering it a promising enzyme for the production of lactose-free dairy products and the further development of domestic enzyme preparations.