H218O isotope exchange studies on the mechanism of reduction of nitric oxide and nitrite to nitrous oxide by denitrifying bacteria. Evidence for an electrophilic nitrosyl during reduction of nitric oxide

Synthetic Modeling of Nitrite Binding and Activation by Reduced Copper Proteins. Characterization of Copper(I)−Nitrite Complexes That Evolve Nitric Oxide

“…Demonstration of their competence to evolve nitric oxide upon protonation further corroborates their suggested significance as intermediates in dissimilatory nitrite reduction by copper proteins. Several issues remain unresolved, however, such as the mechanism of N 2 O production 6 and the intermediacy of [CuNO] 10 species;15a these are the focus of ongoing efforts in our laboratory.…”

“…On the basis of the crystallographic analysis of nitrite-soaked NiR, , activity studies on type 2 depleted enzyme, EPR and ENDOR data for the related copper NiR from Alcaligenes xylosoxidans , effects of site-directed mutagenesis of copper ligands, and pulse radiolysis experiments interpreted to support intramolecular electron transfer, Cu-II has been suggested to be the locus for nitrite binding and subsequent activation, with Cu-I acting to shuttle electrons, perhaps via the Cys-His bridge, during catalysis. Drawing an analogy to mechanisms deduced for the more thoroughly studied iron heme-containing NiRs for which numerous synthetic models exist, Averill and co-workers proposed that a nitrite adduct to Cu-II in its reduced state [i.e., a Cu I −NO 2 - complex] is a key reaction intermediate (Figure ). ,, This intermediate may be envisioned to form via reduction of a Cu II −NO 2 - precursor or directly, via substrate binding to the prereduced Cu I site. Subsequent dehydration to yield an electrophilic copper−nitrosyl (originally written as Cu I −NO + ) 14a followed by release of NO, the principal enzyme product, is then postulated to generate the oxidized Cu-II center ready for reentry into the catalytic process.

2 Proposed mechanism for nitrite reduction by NiR.

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Synthesis, Structure, and Reactivity of Model Complexes of Copper Nitrite Reductase

“…While the activation of nitrite bound to Cu(I) on protonation was reported by Halfen and Tolman,16b we show here that protonation and dehydration of bound nitrite occur faster than its reduction. Although the existence of an intermediate Cu(I)−NO + species in the enzymatic reaction has never been directly proved, there is mechanistic evidence for its formation from the in vitro reactivity of the enzyme. , From the data in Table it is evident that the increase in the number of donor groups in the ligand enhances the activity of the Cu(I) complex in the reaction, in spite of the lower accessibility of the Cu(I) center to NO 2 - or NO + . It is also clear that the increase of donor strength of the nitrogen donors (1-BB or 2-BB vs TB) facilitates electron transfer from Cu(I) to the bound NO + residue.…”

Binding and activation of nitrite and nitric oxide by copper nitrite reductase and corresponding model complexes

“…and then react with a molecule of NO 2 − to form nitrous oxide (N 2 O) via a potential Cu(I)-N 2 O 3 intermediate that decomposes to N 2 O as shown in Scheme 2, top 10,46,47. Alternatively, Tolman suggested that NO disproportionation (cf.…”