Computational Study of the Non-Heme Iron Active Site in Superoxide Reductase and Its Reaction with Superoxide

Abstract

The ferrous square-pyramidal [Fe(NHis)4(SCys)] site of superoxide reductases (SORs) has been shown to reduce superoxide at a nearly diffusion-controlled rate. The final products of the reaction are hydrogen peroxide and the ferric hexacoordinated SOR site, with a carboxylate group from a conserved glutamate serving as the sixth ligand trans to the cysteine sulfur. A transient intermediate absorbing at approximately 600 nm in the reaction of the ferrous pentacoordinated site with superoxide has been proposed to be a ferric-(hydro)peroxo complex (Coulter, E.; Emerson, J.; Kurtz, D. M., Jr.; Cabelli, D. J. Am. Chem. Soc. 2000, 122, 11555-11556.). In the present study, DFT and ZINDO/S-CI results are shown to support the description of the 600-nm intermediate as an end-on, low-spin ferricperoxo or--hydroperoxo complex. Side-on peroxo coordination was found to be significantly less stable than end-on because of constraints on the imidazole ligand ring orientations imposed mostly by the protein. The modeled ferric-hydroperoxo complex had a decidedly nonplanar CysC beta-S-Fe-O-O geometry that appears to be imposed by the same constraints. A single prominent visible absorption of the (hydro)peroxo model is shown to be due mainly to a CysS-->Fe(III) pi charge transfer (CT) transition with a minor portion of His-->Fe(III) pi CT character and very little peroxo-->Fe(III) CT character. On the basis of calculations of models with various mono- and diprotonated peroxo ligands, protonation of the iron-bound peroxo oxygen is a key step in the decay of the ferric(hydro)peroxo complex favoring release of hydrogen peroxide over cleavage of the O-O bond, as occurs in the heme structural analogue, cytochrome P450.

Sulfur K-Edge X-ray Absorption Spectroscopy and Density Functional Theory Calculations on Superoxide Reductase: Role of the Axial Thiolate in Reactivity

“…This is in contrast to the results in Ref. 17 where a low-spin ground state was calculated to be more stable. Thus this computational model that predicts the correct ground state and thiolate covalency of both the resting high-spin and the CN − bound low-spin form of SOR predicts a high-spin ground state of an Fe III -OOH intermediate.…”

“…A different modeling approach was used in Ref. 17. Though neither the α carbon constraints nor the H-bonding interactions were included, this model reproduced the high-spin resting state using BP86 functional and a DN** basis set.…”

“…10 The reaction mechanism of SOR, involving transfer of an electron and two protons to superoxide to form hydrogen peroxide, has been studied in detail using kinetic and spectroscopic techniques. 12,13,14,15,16,17,18 Using kinetic data, Nivière et al and Kurtz et al have proposed the presence of at least one Fe III intermediate (henceforth referred to as intermediate I, characterized by a CT band at 600 nm) in the reaction. The formation of the intermediate was diffusion controlled and had no pH dependence or deuterium isotope effect and thus was proposed to be an Fe III -μ 2 -O 2 2− species.…”

“…A subsequent protonation driven H 2 O 2 release from this low-spin Fe III -OOH was proposed, though the energetic feasibility of such steps were not evaluated. 17 24 The trans effect of the thiolate has also been proposed to affect NO binding to this active site. 27 The thiolate may also play a major role in controlling the reduction potential of the active site.…”

Dioxygen Activation by Copper-Bleomycin: Theoretical Considerations

“…26 In fact the O−O bond in this copper(II)-peroxo model is distinctly longer than the 1.33 − 1.36 Å seen in related Fe(III)-peroxo models in heme and non-heme environment. 3,24,25 The geometrical parameters within the Cu−O−O−(H) moiety in 3, and especially the 1.47 Å O−O distance, are generally in line with previously reported data on related species, showing a clean Cu-hydroperoxo structure, 26 as also supported by the negative charge on the OOH (equal in magnitude to the charge computed for the superoxo ligand in 1) and by the negligible spin density (0.26 units). Figure 3 illustrates the spin density in model 2.…”