Identification, kinetics and thermodynamic analysis of novel β-galactosidase from Convolvulus arvensis seeds: An...

The β-galactosidase was extracted and purified from 100 g of C. arvensis seeds using a variety of protein purification procedures such as ammonium sulphate fractionation, gel filtration, and finally chromatography on a cationic ion exchanger. The effects of metal ions, kinetics parameters, and glycoprotein nature were determined, as well as the optimal pH and temperature of the purified enzyme. With a high specific activity (72 units/mg), β-galactosidase was isolated to a 24-fold apparent electrophoretic homogeneity. The molecular mass of β-galactosidase was determined as monomeric, which was further confirmed by SDS-PAGE and MALDI-TOF/MS analysis, with a 45 kDa molecular weight. The enzyme has a K_m of 0.33 mM and a V_max of 42 μmol/min Lactose in milk was reduced by 38.5 and 70 % after 4 h of incubation with β-galactosidase from C. arvensis. The β-galactosidase thermal inactivation kinetic parameters ΔH°, ΔS°, and ΔG° were calculated, indicating that the enzyme undergoes significant unfolding events during denaturation. Using β-galactosidase from C. arvensis seeds, lactose hydrolysis in milk up to approx. 50 % was observed. The findings indicate the potential use of C. arvensis seeds for the production of low/delactosed milk for lactose-intolerant population.