Redox-dependent changes in beta-sheet and loop structures of Cu,Zn superoxide dismutase in solution observed by infrared spectroscopy.

Redox-dependent conformational changes of bovine Cu,Zn superoxide dismutase in 20 mM phosphate buffer (pH 7.4) were studied at 20 degrees C using Fourier transform infrared spectroscopy. Amide I spectra provide evidence that conformational changes in the protein accompany a change in the oxidation state of copper at the active site. Quantitative analysis of these spectra indicates that both reduced (CuI,ZnII) and oxidized (CuII,ZnII) enzymes are composed of about 35% antiparallel beta-sheet, 45% unordered/loop, and 20% beta-turn structures. Significant redox-dependent changes occur in regions ascribed to beta-sheet and unordered/loop structures that are consistent with an active channel structure wherein the copper ion bonds to imidazolate side chains of His 44, 46, and 118 within the beta-sheet structure and also to the imidazolate side chain of His 61 associated with unordered/loop structure. This study provides the first experimental evidence that an unordered structure can exhibit bands in more than one region, one near 1658 cm-1 and another near 1648 cm-1 in both H2O and D2O solutions. The detected changes in protein conformation are expected to be critical to the catalytic function of this enzyme.