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 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 microB 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 magnitude of J12 = magnitude of J23 = magnitude of J13 = magnitude of 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 (magnitude of 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(mu 2-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.

Evidence for polynuclear aggregates of ferric daunomycin. A Mössbauer, EPR, X-ray absorption spectroscopy and magnetic susceptibility study.

The interaction of the antitumor agent daunomycin (DN) with ferric iron has been analysed by Mössbauer spectroscopy, EPR, extended X-ray absorption fine structure (EXAFS), and magnetic susceptibility measurements. In contrast to literature data, at millimolar iron and anthracycline concentrations no solitary Fe(DN)3 complexes are formed in appreciable amounts. The Mössbauer spectroscopic analysis revealed severe dependencies on temperature, on the preparation procedure, the time allowed for equilibration, and on the metal/ligand ratio. The Mössbauer spectra exhibit two components: a broad magnetic sextet and a quadrupole doublet at an Fe/DN molar ratio of 1:3 and exclusively a doublet at a molar ratio of 1:20, indicating an equilibrium of these two spectral components. The EPR spectra are dominated by a signal at g(eff) = 2. Double integration of the EPR signals enabled the determination of their spin density and a correlation between EPR and Mössbauer spectra. The Mössbauer sextet species is EPR invisible and corresponds to magnetically ordered polynuclear aggregates with high magnetic anisotropy. EXAFS and susceptibility measurements provide additional evidence for the formation of polynuclear aggregates of ferric daunomycin. The quadrupole doublet species in the Mössbauer spectra correlates with the g = 2 signal in EPR. This species is also related to a magnetically ordered system, exhibiting, however, superparamagnetic behavior due to less magnetic anisotropy. Since daunomycin forms dimers in aqueous solution at millimolar concentrations, we conclude that the cooperative phenomena observed in EPR and Mössbauer spectra are a consequence of its stacking effects.