EPR and ligand field studies of iron superoxide dismutases and iron-substituted manganese superoxide dismutases: Relationships between electronic structure of the active site and activity

The problem of metal selectivity of iron/manganese superoxide dismutases (S ODs) is addressed through the electronic structures of active sites using e lectron paramagnetic resonance and ligand field calculations. Studies of wi ld-type iron(III) SOD (FeSOD) from Escherichia coil and from Methanobacteri um thermoautotrophicum and iron-substituted manganese(III) SOD (Fe(sub)MnSO D) from E. coli and from Serratia marcescens are reported. EPR spectroscopy of wild-type enzymes shows transitions within all three Kramers doublets i dentified by their g values. From the temperature dependence of the observe d transitions, the zero-field splitting is found to be negative, D = -2 +/- 0.2 cm(-1). The electronic structure is typical of a distorted trigonal bi pyramid, all the EPR features being reproduced by ligand field analysis. Th is unique and necessary electronic structure characterizes wild-type enzyme s whatever their classification from the amino acid sequence into iron or m anganese types, as E. coli FeSOD or M. thermoautotrophicum FeSOD. In iron-s ubstituted manganese SODs, reduced catalytic activity is found. We describe how inhomogeneity of all reported substituted MnSODs might explain the act ivity decrease. EPR spectra of substituted enzymes show several overlapping components. From simulation of these spectra, one component is identified which shares the same electronic structure of the wild-type FeSODs, with th e proportion depending on pH. Ligand field calculations were performed to i nvestigate distortions of the active site geometry which induce variation o f the excitation energy of the lowest quartet state. The corresponding coup ling between the ground state and the elicited state is found to be maximum in the geometry of the native SODs. We conjecture that such couplings shou ld be considered in the electron-transfer process and in the contribution o f the typical electronic structure of FeSOD to the activity.