Evaluation of 13C and 1H Fermi contact shifts in horse cytochrome c. The origin of the anti-Curie effect.

Many ferricytochromes c exhibit a peculiar effect in which the 1H chemical shifts of the haem methyl groups appear in pairs and, although the paramagnetic shifts of the two groups with the larger shifts decrease with temperature, those of the pair with the smaller shifts actually increase. Recent NMR studies [Santos, H. and Turner, D. L. (1992) Eur. J. Biochem. 206, 721-728] gave 1H and 13C assignments for most of the haem substituents and the axial ligands in horse cytochrome c at 30 degrees C and 50 degrees C in both oxidation states. These data are used together with an empirically determined magnetic susceptibility tensor to evaluate the Fermi contact contribution to the paramagnetic shift and hence map the delocalization of the unpaired electron. The anti-Curie effect is explained by a Boltzmann distribution between partially filled porphyrin 3e(pi) molecular orbitals with an energy difference of 3 kJ/mol. The fact that the energy gap is small with respect to the energy of binding to the electron transfer partners calls into question the significance of the asymmetry of the electron distribution in the electron transfer process.