N.V.Shokhirev, F.A.Walker.
Analysis of the temperature dependence of the H-1 contact shifts in low-spin
Fe(III)
model hemes and heme proteins: Explanation of ''Curie'' and ''anti-Curie'' behavior
within the same molecule.
Journal of Physical Chemistry. 99: (50) 17795-17804 DEC 14 1995
Abstract:
The reasons that the temperature dependence of the NMR isotropic shifts
of model ferrihemes and ferriheme proteins deviate from Curie behavior
have been analyzed by considering the energies of the valence orbitals of
the metal and the porphyrinate. For low-spin Fe(III), overlap of the
e-symmetry pi orbitals of a symmetrical porphyrin ring and the
d(p) orbitals of the metal produces two
low-energy molecular orbitals that are mainly porphyrin in character and
are filled and two high-energy (valence) molecular orbitals that are
mainly metal in character and contain three electrons. The odd electron
in the valence set thus gives rise to the spin delocalization that
results in the observed contact shift pattern of these systems.
Unsymmetrical substitution and/or presence of a planar axial ligand that
is prevented from rotation removes the degeneracy of these
e(p) orbitals, producing a system in
which the energy separation between the two formerly degenerate pi orbitals, Delta
E(pi), is of the order of only tens of cm-1
for the former or quite large (several times k(B)T) for the latter. In
either case, both orbitals are utilized for spin delocalization to a
significant extent as the temperature is varied, according to their
varying Boltzmann populations. Such a two-level system obeys a modified
Curie law that takes into account the thermal population of the two
levels as a function of temperature. In fact, the temperature dependence
of some of the contact shifts of model hemes or heme proteins may show
anti-Curie behavior if Delta E(pi) is large compared to kBT at
ambient emperatures. Such anti-Curie behavior has been observed for two
of the heme methyl resonances of several cytochromes c and b(5) and
cyanometmyoglobins or -hemoglobins, where the axial methionine pi-symmetry
lone pair or histidine imidazole plane orientation, respectively, is
believed to be the important factor in determining
D E(p).
Assuming reasonable energy separations of the two valence
e(p) orbitals, from very small to quite
large (similar to 1000 cm-1), the expected temperature
dependence of the contact shifts has been calculated for an assumed set
of valence MO coefficients. These results have then been compared to the
experimental isotropic shifts of several model heme systems having
unsymmetrical substitution patterns and/or one fixed axial ligand and to
several heme proteins. Using a computer program developed to fit the
observed isotropic shifts to the two-level equation,
D E(p) was
estimated from the temperature dependence of the isotropic shifts of the
protons of the beta-pyrrole substituents of the above-mentioned systems.
In the case of the proteins investigated, Aplysia cyanometmyoglobin and
cytochrome b(5), the values of D
E(p), obtained from analysis of proton
isotropic shifts are similar to those calculated from EPR g values
measured at low temperatures, while for model hemins, the values of
D E(p)
obtained are smaller than those predicted and vary in accord with the
expectations as to the rigidity, or lack thereof, of the orientation of
at least one planar axial ligand, indicating that thermal averaging of
the two levels due to rapid rotation (or libration) of the axial ligand
is fast on the NMR time scale. This same two-level approach could be
applied to any system in which there is a thermal equilibrium between two
states separated by an energy within several factors of kBT.