Studies on the EPR and Light Absorption Spectra of Horse Erythrocyte Catalase and Its Derivatives

Abstract

1. The EPR spectra of horse erythrocyte catalase [EG 1.11.1.6] and its derivatives were measured at liquid nitrogen temperature. Free catalase and HF-catalase gave EPR spectra having two intense signals with g-values of 6.6 and 5.4 at low magnetic field and a weak signal with g-value of 2.03 at high magnetic field. The EPR spectrum of HCN-catalase showed three signals with g-values of 1.66, 2.25 and 2.84, and that of HN₃-catalase exhibited signals with g-values of 6.7, 5.2, 2.80, 2.18 and 1.74. From the data, the electronic states of the hematin-irons of free catalase and HF-catalase were of a high-spin type, and those of HCN-catalase and HN₃-catalase were of low-spin type and of a mixture of high- and low-spin types, respectively.

2. The optical absorption spectra of free catalase and the HF-and HCN-complexes measured at liquid nitrogen temperature were essentially the same as those obtained at room temperature. However, the optical absorption spectrum of HN₃-catalase at liquid nitrogen temperature was of a mixture of high- and low-spin types, although the absorption spectrum of this complex at room temperature was of high-spin type. The absorption maxima, which are characteristic of the low-spin type, appeared at temperatures below\emdash 80°C; that is, in HN₃-catalase, there seemed to be thermal equilibrium between the low-and high-spin forms.

3. The energy differences between the ξ and η or ζ orbitals of the HCN- and HN₃-complexes were calculated from the EPR signals of the low-spin type. It was found that the energy difference between the ξ and η orbitals of de is 900 cm⁻¹ for HGN-catalase and 1000 cm'1 for HN₃-catalase, and the difference between the ξ and ζ orbitals of de is 2000 cm-1 for HCN-catalase and 2700 cm⁻¹ for HN₃-catalase. From the data on the EPR spectra of the high-spin type, the energy difference, E_y—E_x, was found to be 600 cm⁻¹ for free catalase and HF-catalase, and 700 cm⁻¹ for HN₃-catalase.