Mössbauer and magnetic susceptibility studies on iron(II) metallothionein from rabbit liver: Evidence for the existence of an unusual type of [M3(CysS)9]3‐ cluster

The magnetic properties of the Fe(II)‐binding sites in Fe(II)₇‐metallothionein (MT) have been studied using Mössbauer spectroscopy and magnetic‐susceptibility measurements. In agreement with our previous results, simulation of the Mössbauer spectra showed the presence of paramagnetic and diamagnetic subspectra in the ratio 3:4. By comparison with Mössbauer spectra of the inorganic adamantane‐like (Et₄N)₂[Fe₄(SEt)₁₀] model compound, the diamagnetic component in Fe(II)₇‐MT has been assigned to a four‐metal cluster in which there is antiferromagnetic coupling between the high‐spin Fe(II) ions. It is suggested that the organization of this cluster is similar to that determined in the three‐dimensional structure of the protein, containing diamagnetic Zn(II) and/or Cd(II) ions. From magnetic‐susceptibility studies, an average magnetic moment of approximately 8.5 μ_B was obtained for the three remaining bound Fe(II) ions, responsible for the paramagnetic component observed in the Mössbauer studies. This value is slightly lower than that for three completely uncoupled Fe(II) ions, suggesting the existence of a three‐metal cluster within which there is weak exchange coupling between adjacent Fe(II) ions. The spin‐Hamiltonian formalism including, besides zero‐field and Zeeman interaction, also exchange interaction among the three Fe(II) ions in the three‐metal cluster, H = ‐J₁₂ (S₁·S₂)‐J₂₃ (S₂·S₃‐J₁₃ (S₁·S₃), was applied to simulate both magnetic‐Mössbauer and magnetic‐susceptibility data. Reasonable fits were achieved only with values |J₁₂|=|J₂₃|=|J₁₃|=|J| < 1 cm⁻¹. Such a situation could not be reconciled with the chair‐like geometry of the [M₃(CysS)₉]^3‐ cluster determined with paramagnetic metal ions, where significantly stronger coupling would be anticipated (|J|= 50–70 cm⁻¹). However, modest exchange‐coupling properties have been reported for a number of crystallographically characterized trinuclear [Fe₃(SR)₃X₆]^3‐ clusters (X = Cl, Br; R = Phe, p‐tolyl, 2,6‐Me₂C₆H₃) distinguished by the preferential formation of a planar Fe₃(μ₂‐SR)₃ ring [Whitener, M. A., Bashkin, J. A., Hagen, K. S., Girerd, J.‐J., Gamp, E. Edelstein, N. & Holm, R. H. (1986) J. Amer. Chem. Soc. 108, 5607–5620]. It is therefore more likely that a pseudo‐planar geometry rather than a chair‐like geometry is present in the Fe₃ cluster of Fe(II)₇‐MT. This would represent the first example of structural differences on binding divalent metal ions to this protein.