Manganese superoxide dismutase from Escherichia coli has been prepared in homogeneous Mn3+ and Mn2+ redox forms for characterization by a combination of optical absorption, circular dichroism (CD), magnetic circular dichroism (MCD), and EPR spectroscopies. MCD spectra of the unliganded Mn3+ protein displays a strikingly simple pattern, a pair of bands of equal magnitude but oppositely signed intensity. Saturation magnetization curves for the native Mn site show a dramatic nesting that reflects large splittings in the paramagnetic ground state, suggesting postive zero-field splitting (D > 0). The ground-state behavior and the excited-state spectra can be interpreted in terms of a distorted trigonal-bipyramidal environment for the metal ion. Binding exogenous ligands (F−, N3−) perturbs the Mn3+ site, leading to a distinctly different pattern of MCD intensity, a pseudo-A-term feature at high energy that exhibits a strong magnetic field saturation at low temperature. Saturation magnetization curves for the anion complexes are less broadly nested than for the native enzyme and appear to arise from a rhombically split non-Kramers doublet lowest in the quintet ground state (D < 0). A change in the nature of the orbital ground state evidently occurs as a consequence of the metal complex distorting when small molecules coordinate. A Berry pseudorotation model is presented for the distorting Mn3+ site to account for the spectroscopic changes associated with anion binding. On reduction, absorption and CD signals are lost for the Mn center, but weak temperature-dependent MCD features are observed for the d5 metal ion, providing a probe of the enzyme in this redox form. The reduced active site also responds to anion interactions, reflected in the EPR spectra for the Mn2+ center, suggesting that small molecules coordinate to both oxidized and reduced metal centers in MnSD.
ASJC Scopus subject areas
- Colloid and Surface Chemistry