β-Galactosidase from Lactobacillus acidophilus isolated from fermented ragi (Eleusine coracana)

Lactobacillus acidophilus was isolated from fermented millet, Eleusine coracana. It was characterized using several biochemical tests. This strain was found to be homofermentative, slime forming and β-galactosidase producer. Recovery and characterization of β-galactosidase were studied at laboratory scale. Since the enzyme was intracellular various cell lysis methods were studied to achieve maximum enzyme release from cell fragments. Homogenization at a pressure of 2000 psig two passes showed an enzyme release of 2050 U/mL and was found the best method for cell lysis. The cell extract was purified using ultrafiltration and gel permeation chromatography. Specific activity and fold purification of β-galactosidase was found to be 568.61 and 21.2, respectively. Kinetic parameters were determined using ONPG (o-nitrophenyl galactopyranoside) as a substrate. Michaeli’s Menten equation was found to fit the reaction of ONPG using β-galactosidase from L. acidophilus. Vmax and Km were calculated from the Lineweaver Burk plot. Vmax was found to be 4.94 min -1 and Km to be 0.11 mM. Using gel permeation chromatography, the molecular weight was found to be in the range of 450-500 kDa.