Magnetic Susceptibility and EPR Measurements of Catalase and Its Derivatives

A Thermal Equilibrium between the High- and Low-spin States in the Catalase-azide Compound

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The molar paramagnetic susceptibilities (χ_mole) per Fe³⁺ ion of free catalase [EC 1. 11. 1. 6] and its fluoride, cyanide and azide compounds were measured with a magnetic torsion balance over the range from room temperature to liquid nitrogen temperature. From the slopes of the linear relationships for χ_mole-1/T the values of the effective Bohr magneton number (n_eff) were found to be 5.96 for free catalase, 5.78 for the fluoride compound and 2.63 for the cyanide compound. However, the curve for χ_mole-1/T of the azide compound deviated markedly from Curie's law. This can be explained by assuming that there is a thermal equilibrium between the high-spin (S=5/2) and low-spin (S=1/2) states of the hematin irons of this compound. From the data that the proportion of the low-spin form to the total concentration of the hematin irons of this compound increased on lowering the temperature, the energy level of the low-spin state seems to be lower than that of the high-spin state. The EPR spectrum of the azide compound was measured at 20°K and 77°K. The spec-trum at 20°K appeared to be more of low-spin type than at 77°K. This tendency accords with the results obtained from the magnetic susceptibility measurements.