The geometric and electronic structure of NO bound to reduced protocatechuate 3,4-dioxygenase and its substrate (3,4-dihydroxybenzoate, PCA) complex have been examined by X-ray absorption (XAS), UV-vis absorption (Abs), magnetic circular dichroism (MCD), and variable temperature variable field (VTVH) MCD spectroscopies. The results are compared to those previously published on model complexes described as {FeNO}(7) systems in which an S = 5/2 ferric center is antiferromagnetically coupled to an S = 1 NO-. XAS pre-edge analysis indicates that the Fe-NO units in (FePCD)-P-III{NO-} and (FePCD)-P-III{PCA,NO-} lack the greatly increased pre-edge intensity representative of most {FeNO}(7) model sites. Furthermore, from extended X-ray absorption fine structure (EXAFS) analysis, the (FePCD)-P-III{NO-} and (FePCD)-P-III{PCA,NO-} active sites are shown to have an Fe-NO distance of at least 1.91 Angstrom, approximate to0.2 Angstrom greater than those found in the model complexes. The weakened Fe-NO bond is consistent with the overall lengthening of the bond lengths and the fact that VTVH MCD data show that NO- --> Fe-III CT transitions are no longer polarized along the z-axis of the zero-field splitting tensor. The weaker Fe-NO bond derives from the strong donor interaction of the endogenous phenolate and substrate catecholate ligands, which is observed from the increased intensity in the CT region relative to that of {FeNO}(7) model complexes, and from the shift in XAS edge position to lower energy. As NO is an analogue of O-2, the effect of endogenous ligand donor strength on the Fe-NO bond has important implications with respect to O-2 activation by non-heme iron enzymes.