It is shown that, in the oxidized
state, heme c of Pseudomonas stutzeri (ZoBell strain)
cytochrome
cd1
has histidine-methionine ligation as observed for cytochrome
cd1
from Pseudomonas aeruginosa [Sutherland, J., Greenwood, C.,
Peterson,
J., and Thomson, A. J. (1986) Biochem. J. 233, 893-898]. However, the
X-ray
structure of Thiosphaera pantotropha cytochrome
cd1
reveals bis-histidine ligation for heme c. It is confirmed by
EPR
and near-infrared (NIR) MCD measurements that the bis-histidine
coordination
remains unaltered in the solution phase. Hence, the difference between
the heme c ligation states defines two distinct classes of oxidized
cytochromes
cd1 as isolated. A weak feature in the T.
pantotropha NIR MCD at 1900 nm suggests that a small population of
heme c has histidine-methionine coordination. The ligation
state
of heme d1 cannot be defined with the same level of
confidence,
because the porphyrin-to-Fe(III) charge-transfer (CT) bands are less
well
characterized for this class of partially reduced porphyrin ring.
However,
variable temperature absorption and MCD spectra show that, in the T.
pantotropha enzyme, heme d1 exists
in a
thermal low-spin/high-spin mixture with the low-spin as the ground
state,
whereas in P. stutzeri cytochrome
cd1,
and d1 heme is low-spin at all
temperatures.
A weak band, assigned as the heme d1
porphyrin-p(a1u,a2u)-to-ferric(d)
charge-transfer transition has been identified for the first time at
2170
nm. Its magnetic properties show the heme d1
to have an unusual (dxz,yz)4(dxy)1electronic
ground state as is found for low-spin Fe(III) chlorins [Cheesman, M.
R.,
and Walker, F. A. (1996) J. Am. Chem. Soc. 118, 7373-7380]. It is
proposed
that the localization of the Fe(III) unpaired d-electron in an orbital
lying in the heme plane may decrease the affinity of the Fe(III) heme
for
unsaturated ligands such as NO. Although heme d1
in the enzymes from P. stutzeri and
T. pantotropha shows
different temperature-dependent spin properties, the positions of the
low-spin
Fe(III) a-absorption
band, at ~
640 nm, are very similar to those observed for cytochromes cd1
from eight other sources, suggesting that all have similar strength
fields
from the axial ligands and, hence, that all have the same coordination,
namely histidine-tyrosine or possibly histidine-hydroxide at the heme.