EPR spectra at liquid helium
temperatures
and MCD spectra at room temperature and 4.2 K are presented for fully
oxidized
nitric oxide reductase (NOR) from Pseudomonas stutzeri. The MCD
spectra show that the enzyme contains three heme groups at equivalent
concentrations
but distinctive in their axial coordination. Two, in the low-spin
ferric
state at all temperatures, give rise to infrared charge-transfer
transitions
which show the hemes to have bis-histidine and histidine-methionine
ligation,
respectively. The EPR spectra show them to be magnetically isolated.
The
third heme has an unusual temperature-dependent spin state and
spectroscopic
features which are consistent with histidine-hydroxide coordination. No
EPR signals have been detected from this heme. Together with its
unusual
near-infrared MCD, this suggests a spin-spin interaction between this
heme
and another paramagnet. The three hemes account for only 75% of the
iron
content, and it is concluded that the additional paramagnet is a
mononuclear
ferric ion. These results provide further evidence that NOR is indeed
structurally
related to heme-copper oxidases and that it contains a heme/non-heme
iron
spin-coupled pair at the active site.