Promotion of Activity and Thermal Stability of Chloroperoxidase by Trace Amount of Metal Ions (M²⁺/M ³⁺)

The effect of M²⁺ (Zn²⁺, Cu²⁺, Cd²⁺, Mn²⁺, Pb²⁺) and M³⁺ (Cr³⁺, La³⁺, Fe³⁺, Ce³⁺, Y³⁺, Al³⁺) metal ions on the activity and thermal stability of chloroperoxidase (CPO) was investigated in this work. It was found that the lower concentration of metal ions was favorable to CPO activity whereas the higher concentration reversed the results. CPO activity could be increased to 116.4–127.1 % in the presence of a trace amount of these M²⁺/M³⁺ metal ions at a concentration range of 0–25 μmol L⁻¹ after 2 h of incubation at 25 °C. The activating effect of M³⁺ is better than that of M²⁺, and Cr³⁺ was mostly efficient. The thermal stability of the enzyme was also improved significantly. Only 30.3 % of CPO activity was retained at 50 °C whereas 82.6 % of CPO activity was maintained in the presence of Cr³⁺ after 2 h of incubation at the same temperature. The activation of CPO by metal ions at their low concentration was studied through intrinsic fluorescence, circular dichroism (CD), and UV–Vis spectra assay. A favorable environment around the active site was achieved in the presence of metal ions. Intrinsic fluorescence and CD spectra indicated that the α-helix structure of CPO was strengthened in metal ion-contained media. More exposure of the heme ring was achieved for easy access of the substrate, which was suggested by UV–Vis spectrum analysis. This strategy for enhancing CPO activity is very simple and useful. It will be favorable to the practical application of this enzyme.