ANALYSIS OF THE TEMPERATURE-DEPENDENCE OF THE H-1 AND C-13 ISOTROPIC SHIFTS OF HORSE HEART FERRICYTOCHROME-C - EXPLANATION OF CURIE AND ANTI-CURIE TEMPERATURE-DEPENDENCE AND NONLINEAR PSEUDOCONTACT SHIFTS IN ACOMMON 2-LEVEL FRAMEWORK

The H-1 and C-13 hyperfine shifts of the heme methyls of horse heart f erricytochrome c have been measured over the temperature range 278-328 K in order to interpret the ''anomalous'' temperature-dependence of t he hyperfine shifts in terms of their pseudocontact and contact shifts . By taking advantage of the available pseudocontact shifts for protei n nuclei measured at 303 and 323 K (Santos, H.; Turner, D. L. fur. J. Biochem. 1992, 206, 721-728), the metal-centered pseudocontact shifts have been analyzed in terms of a thermally accessible excited state ly ing 355-590 cm(-1) to higher energy which has a magnetic susceptibilit y tensor with the rhombic anisotropy, Delta chi(rh), which is rotated by 90 degrees to that of the ground state. The metal-centered pseudoco ntact shifts have been evaluated at all temperatures at which the chem ical shifts were measured, and these calculated values were used to ev aluate the contact shifts of each heme methyl for the two nuclei. The temperature dependence of the heme methyl contact shifts for both H-1 and C-13, assuming a thermally accessible excited state, was then used to evaluate the spin density for the four beta-pyrrole heme carbons t o which the methyls are attached. The ligand-centered pseudocontact sh ifts have been estimated and found to give a modest contribution to th e experimental behavior. The H-1 and C-13 data are highly self-consist ent. The present analysis provides deep insight into the electron dist ribution on the porphyrin ring in low-spin Fe(III) hemes.