Inactivation of an Animal and a Fungal Catalase by Hydrogen Peroxide

Abstract We have quantitatively compared the rates of peroxide-dependent inactivation of bovine liver catalase and Aspergillus niger catalase as class representatives of catalases that contain tightly bound NADPH and those that do not. Inactivation of these catalases in the presence of ethanol has also been quantitated in an effort to assess the importance of compound II, an inactive form of bovine liver catalase, in the inactivation reaction. The values of k 2 , the second-order rate constant for inactivation calculated for the bovine enzyme, in the absence and presence of ethanol, respectively, were 8.9 ± 0.26 and 8.5 ± 0.27 M −1 min −1 . In contrast, the values for the A. niger enzyme were 0.51 ± 0.069 and 10.5 ± 0.32 M −1 min −1 . The A. niger enzyme is more stable toward hydrogen peroxide-induced inactivation than the liver enzyme. The A niger enzyme is markedly destabilized by 20 mM ethanol, whereas the inactivation of the liver enzyme is unaffected by ethanol. Reaction of bovine liver catalase with ethyl hydroperoxide produced the characteristic absorption spectrum of compound I and in the absence of ethanol the spectrum associated with compound II. In contrast, the fungal enzyme developed compound I spectrum but spectral changes that might be ascribed to compound II were barely detected in the Soret region. Spectral changes for A. niger catalase in the visible region were modified by the presence of ethanol but could not be clearly correlated with the bovine catalase compound II spectra either in the presence or absence of ethanol. The stability of the fungal and bovine catalases in the presence of hydrogen peroxide is quantitatively documented. The enzymes are also shown to be different in their response to ethanol and in the formation of compound II-like species with ethyl hydroperoxide. It appears unlikely that compound II is an intermediate in the hydrogen peroxide-mediated inactivation reaction of either catalase under catalatic assay conditions.