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.
Dong
A
A
Department of Biochemistry and Molecular Biology, Colorado State University, Fort Collins 80523, USA.
Huang
P
P
Caughey
W S
WS
D000818:Animals; D001665:Binding Sites; D002417:Cattle; D017666:Deuterium Oxide; D006639:Histidine / Q000737:chemistry; D066298:In Vitro Techniques; D008958:Models, Molecular; D010084:Oxidation-Reduction; D011487:Protein Conformation; D017433:Protein Structure, Secondary; D012996:Solutions; D017550:Spectroscopy, Fourier Transform Infrared; D013482:Superoxide Dismutase / Q000737:chemistry* / Q000378:metabolism; D014867:Water
D012996:Solutions; D014867:Water; D006639:Histidine; D013482:Superoxide Dismutase; D017666:Deuterium Oxide