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 dism utase 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 chang e in the oxidation state of copper at the active site. Quantitative an alysis of these spectra indicates that both reduced (Cu-I,Zn-II) and o xidized (Cu-II,Zn-II) enzymes are composed of about 35% antiparallel b eta-sheet, 45% unordered/loop, and 20% beta-turn structures. Significa nt 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 H is 44, 46, and 118 within the beta-sheet structure and also to the imi dazolate side chain of His 61 associated with unordered/loop structure . This study provides the first experimental evidence that an unordere d structure can exhibit bands in more than one region, one near 1658 c m(-1) and another near 1648 cm(-1) in both H2O and D2O solutions. The detected changes in protein conformation are expected to be critical t o the catalytic function of this enzyme. (C) 1995 Academic Press, Inc.