EFFECTS OF THE IONIC ENVIRONMENT ON THE STABILITY OF KLUYVEROMYCES-LACTIS BETA-GALACTOSIDASE

The effect of several cations on the stability of soluble and whole ce ll Kluyveromyces lactis lactase was studied in buffer solutions as a f unction of cation concentration and temperature. From several divalent cations tested at 45-degrees-C in phosphate buffer (pH 6.6), only Mn2 + (0.1 to 0.2 mmol/L) and to a lesser extent Mg2+ (2.5 to 5.0 mmol/L), protected the enzyme against thermal deactivation, while Zn2+ (10(-3) mmol/L) and Cu2+ (10(-4) mmol/L) deactivated the lactase. Stability w as also dependent on the potassium concentration, but was not affected by sodium ions (20 to 40 mmol/L). Studies in barbital buffer (pH 6.6) showed a strong deactivation effect of Ca2+ (5 to 13 mmol/L). Based o n data of the thermal deactivation kinetics, it could be estimated tha t at low temperatures (<20-degrees-C) the lactases will exhibit high s tability even without the addition of Mg or Mn ions (half life >1 year ). Deactivation by Ca2+, Zn2+ or Cu2+ was markedly dependent on the te mperature and highly susceptible to small changes of the cation concen tration. At 5-degrees-C the deactivation rate was significantly reduce d. Kinetic data of the initial rate of lactose hydrolysis showed that the ionic effect on enzyme stability was not correlated with changes i n the enzymatic activity. The whole cell lactase showed higher stabili ty than the soluble product and was less sensitive to deactivation by cations. Enzyme activity after deactivation by Ca2+ could be almost re stored by washing the cells with phosphates. Activity after deactivati on by Zn+ was partially restored with chelating agents such as EDTA, a nd no significant restoration occurred after deactivation by Cu2+.