ELECTRONIC-STRUCTURE OF THE PERTURBED BLUE COPPER SITE IN NITRITE REDUCTASE - SPECTROSCOPIC PROPERTIES, BONDING, AND IMPLICATIONS FOR THE ENTATIC RACK STATE/

Low-temperature optical absorption, circular dichroism, magnetic circu lar dichroism, and sulfur K-edge X-ray absorption spectra have been me asured for the green ''blue'' copper center (type 1) in Achromobacter cycloclastes nitrite reductase. Combined with density functional calcu lations, the results of these spectroscopies have been used to define the extremely ''perturbed'' electronic structure of this site relative to that of the prototypical ''classic'' site found in plastocyanin. E xperimentally calibrated density functional calculations have been fur ther used to determine the specific geometric distortions which genera te the perturbed electronic structure. These studies indicate that the principal electronic structure changes in nitrite reductase, relative to plastocyanin, are a rotation of the Cu d(x2-y2) half-filled, highe st occupied molecular orbital (HOMO) and an increase in the ligand fie ld strength at the Cu center. The HOMO rotation increases the pseudo-s igma interaction and decreases the pi interaction of the cysteine (Cys ) sulfur with Cu d(x2-y2). Furthermore, significant methionine (Met) s ulfur character is mixed into the HOMO due to increased overlap with C u d(x2-y2). These changes in Cu-ligand interactions result in the redi stribution of absorption intensity in the charge transfer and ligand f ield transitions. Additionally, the new S(Met)-Cu interaction accounts for the unexpectedly high sulfur covalency in the HOMO. The increase in ligand field strength shifts all the d --> d transitions in nitrite reductase to similar to 1000 cm(-1) higher energy than their counterp arts in plastocyanin, which accounts for the EPR spectral differences between the type 1 sites in these complexes. The geometric distortion primarily responsible for the electronic structure changes in nitrite reductase, relative to plastocyanin, is determined to involve a couple d angular movement of the Cys and Met residues toward a more flattened tetrahedral (toward square planar) structure. This movement is consis tent with a tetragonal Jahn-Teller distortion resulting from the short er Cu-S(Met) bond in nitrite reductase relative to plastocyanin. This increased Jahn-Teller distortion implies that the type 1 site is ''les s entatic'' than that in plastocyanin.