| 19383478 |
Hyde JS, Bennett B, Walter ED, Millhauser GL, Sidabras JW, Antholine WE: EPR of Cu2+ prion protein constructs at 2 GHz using the g (perpendicular) region to characterize nitrogen ligation. Biophys J. 2009 Apr 22;96(8):3354-62.
A double octarepeat prion protein construct, which has two histidines, mixed with copper sulfate in a 3:2 molar ratio provides at most three imidazole ligands to each copper ion to form a square-planar Cu (2+) complex. This work is concerned with identification of the fourth ligand. A new (to our knowledge) electron paramagnetic resonance method based on analysis of the intense features of the electron paramagnetic resonance spectrum in the g ( perpendicular) region at 2 GHz is introduced to distinguish between three and four nitrogen ligands. The methodology was established by studies of a model system consisting of histidine imidazole ligation to Cu (2+). In this spectral region at 2 GHz (S-band), g-strain and broadening from the possible rhombic character of the Zeeman interaction are small. The most intense line is identified with the M (I) = +1/2 extra absorption peak. Spectral simulation demonstrated that this peak is insensitive to cupric A (x) and A (y) hyperfine interaction. The spectral region to the high-field side of this peak is uncluttered and suitable for analysis of nitrogen superhyperfine couplings to determine the number of nitrogens. The spectral region to the low-field side of the intense extra absorption peak in the g ( perpendicular) part of the spectrum is sensitive to the rhombic distortion parameters A (x) and A (y). Application of the method to the prion protein system indicates that two species are present and that the dominant species contains four nitrogen ligands. A new loop-gap microwave resonator is described that contains approximately 1 mL of frozen sample. |
33(0,1,1,3) |