THERMOSTABILIZATION OF KLUYVEROMYCES-MARXIANUS BETA-GALACTOSIDASE BY HISTIDINE - PHYSICAL STUDIES

The beta-galactosidase from Kluyveromyces marxianus was purified from a commercial source (LactAid) to electrophoretic homogeneity. It was r apidly inactivated at 45 degrees C, but was stabilized against activit y loss by histidine at low concentrations, Stabilization by histidine was enhanced in the presence of lactose. Differential scanning calorim etry of enzyme solutions showed two enthalpic peaks: a primary melting transition (T-m) of approximate to 51 degrees C and a secondary trans ition, at a higher temperature, whose existence and melting temperatur e were concentration-dependent and may be related to aggregation, Lact ose, galactose and glucose increased the value of T-m, but histidine h ad no effect. Unfolding of the enzyme under isothermal conditions at 4 7.5 degrees C was followed by aggregation, Histidine delayed unfolding at temperatures from 43 degrees C to 46 degrees C, but not at 47.5 de grees C, Binding of histidine to the enzyme could not be detected by e quilibrium dialysis or gel filtration, It was concluded that histidine stabilized the enzyme by delaying unfolding at temperatures below the melting transition.