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.