Redox-Dependent Magnetic Alignment of Clostridium pasteurianum Rubredoxin: Measurement of Magnetic Susceptibility Anisotropy and Prediction of Pseudocontact Shift Contributions

An analysis of the magnetic field dependence of one-bond couplings has yielded the magnetic susceptibility anisotropies for Clostridium pasteurianum rubredoxin (Rdx) in its oxidized Fe(III) and reduced Fe(II) states. Experimental one-bond 1HN−15N and 1Hα−13Cα couplings were measured at two field strengths (corresponding to 400 and 750 MHz 1H frequencies) and decomposed into their field-independent scalar (1J) and field-dependent dipolar (1D) components. The total numbers of measured dipolar couplings (1HN−15N plus 1Hα−13Cα) were 50 for oxidized Rdx and 49 for reduced Rdx. The atom pairs giving rise to these signals are located >11 A from the iron; those closer to the iron are too broad to be resolved in two-dimensional NMR spectra and may exhibit large Fermi contact shifts. A five-dimensional grid search and Powell minimization of the difference between each set of measured dipolar couplings and those calculated from an X-ray crystal structure of Fe(III) Rdx yielded the magnitude and orientation of the ma...