Is divalent magnesium cation the best cofactor for bacterial β-galactosidase?

β-Galactosidase is a metal-activated enzyme, which breaks down the glucosidic bond of lactose and produces glucose and galactose. Among several commercial applications, preparation of lactose-free milk has gained special attention. The present objective is to demonstrate the activity kinetics of β-galactosidase purified from a non-pathogenic bacterium Arthrobacter oxydans SB. The enzyme was purified by DEAE-cellulose and Sephadex G-100 column chromatography. The purity of the protein was checked by high-performance liquid chromatography (HPLC). The purified enzyme of molecular weight ~95 kDa exhibited specific activity of 137.7 U mg-1 protein with a purification of 11.22-fold and yield 12.42%. The exact molecular weight (95.7 kDa) of the purified protein was determined by MALDI-TOF. Previously, most of the studies have used Mg+2 as a cofactor of β-galactosidase. In this present investigation, we have checked the kinetic behavior of the purified β-galactosidase in presence of several bivalent metals. Lowest Km with highest substrate (orthonitrophenyl- β-galactoside or ONPG) affinity was measured in presence of Ca2+ (42.45 μM ONPG). However, our results demonstrated that Vmax was maximum in presence of Mn+2 (55.98 μM ONP produced mg-1 protein min-1), followed by Fe=2, Zn+2, Mg+2, Cu+2 and Ca+2. A large number of investigations reported Mg+2 as potential co factor for bgalacosidase. However, β-galactosidase obtained from Arthrobacter oxydans SB has better activity in the presence of Mn+2 or Fe2+.