PROTON ENDOR SPECTROSCOPY OF CYANIDE-INHIBITED COPPER-ZINC SUPEROXIDE-DISMUTASE IN RANDOMLY ORIENTED SAMPLES

Electron nuclear double resonance (ENDOR) spectroscopy of frozen solut ions of samples of CN-inhibited superoxide dismutase (CN-SOD) was appl ied in order to unravel the structural changes and the changes in elec tronic parameters which occur in native SOD upon CN- ligation, Two sam ples were prepared from bovine sources in H2O and in D2O in order dete ct buffer exchangeable protons in hydrogen bonds. Two further samples from human sources were employed which contained specifically deuterat ed histidine carbon-bound proton positions. Measurements were taken at about 10 K from the proton ENDOR response at different magnetic field positions of the electron paramagnetic resonance spectrum. The experi mental results from the samples at different deuteration levels were r eproduced by simulations of orientation-selected ENDOR powder-type spe ctra. These data were integrated into a new structure of the copper si te of CN-SOD together with positions of the g tensor axes and the spin densities in the complex. Comparison is made in relation to the nativ e SOD in both structural and electronic parameters aspects and with th e structural proposal from X-ray determination of CN-SOD single crysta ls. (C) 1998 Elsevier Science S.A. All rights reserved.