N-O bond cleavage mechanism(s) in nitrous oxide reductase

Quantum chemical calculations of active-site models of nitrous oxide reductase (N ₂OR) have been undertaken to elucidate the mechanism of N-O bond cleavage mediated by the supported tetranuclear Cu ₄S core (Cu _Z) found in the enzymatic active site. Using either a minimal model previously employed by Gorelsky et al. (J. Am. Chem. Soc. 128:278-290, 2006) or a more extended model including key residue side chains in the active-site second shell, we found two distinct mechanisms. In the first model, N ₂O binds to the fully reduced Cu _Z in a bent μ-(1,3)-O,N bridging fashion between the Cu _I and Cu _IV centers and subsequently extrudes N ₂ while generating the corresponding bridged μ-oxo species. In the second model, substrate N ₂O binds loosely to one of the coppers of Cu _Z in a terminal fashion, i.e., using only the oxygen atom; loss of N ₂ generates the same μ-oxo copper core. The free energies of activation predicted for these two alternative pathways are sufficiently close to one another that theory does not provide decisive support for one over the other, posing an interesting problem with respect to experiments that might be designed to distinguish between the two. Effects of nearby residues and active-site water molecules are also explored.