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Spectroscopic and computational study of a nonheme iron nitrosyl center in a biosynthetic model of nitric oxide reductase.

ABSTRACT: A major barrier to understanding the mechanism of nitric oxide reductases (NORs) is the lack of a selective probe of NO binding to the nonheme FeB center. By replacing the heme in a biosynthetic model of NORs, which structurally and functionally mimics NORs, with isostructural ZnPP, the electronic structure and functional properties of the FeB nitrosyl complex was probed. This approach allowed observation of the first S=3/2 nonheme {FeNO}(7) complex in a protein-based model system of NOR. Detailed spectroscopic and computational studies show that the electronic state of the {FeNO}(7) complex is best described as a high spin ferrous iron (S=2) antiferromagnetically coupled to an NO radical (S=1/2) [Fe(2+)-NO(.)]. The radical nature of the FeB -bound NO would facilitate N-N bond formation by radical coupling with the heme-bound NO. This finding, therefore, supports the proposed trans mechanism of NO reduction by NORs.

SUBMITTER: Chakraborty S

PROVIDER: S-EPMC4048663 | biostudies-literature | 2014 Feb

REPOSITORIES: biostudies-literature

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Spectroscopic and computational study of a nonheme iron nitrosyl center in a biosynthetic model of nitric oxide reductase.

Chakraborty Saumen S Reed Julian J Ross Matthew M Nilges Mark J MJ Petrik Igor D ID Ghosh Soumya S Hammes-Schiffer Sharon S Sage J Timothy JT Zhang Yong Y Schulz Charles E CE Lu Yi Y

Angewandte Chemie (International ed. in English) 20140131 9

A major barrier to understanding the mechanism of nitric oxide reductases (NORs) is the lack of a selective probe of NO binding to the nonheme FeB center. By replacing the heme in a biosynthetic model of NORs, which structurally and functionally mimics NORs, with isostructural ZnPP, the electronic structure and functional properties of the FeB nitrosyl complex was probed. This approach allowed observation of the first S=3/2 nonheme {FeNO}(7) complex in a protein-based model system of NOR. Detail ...[more]

PMID: 24481708

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Project description:A series of tetranuclear iron complexes displaying a site-differentiated metal center was synthesized. Three of the metal centers are coordinated to our previously reported ligand, based on a 1,3,5-triarylbenzene motif with nitrogen and oxygen donors. The fourth (apical) iron center is coordinatively unsaturated and appended to the trinuclear core through three bridging pyrazolates and an interstitial μ4-oxide moiety. Electrochemical studies of complex [LFe3(PhPz)3OFe][OTf]2 revealed three reversible redox events assigned to the Fe(II)4/Fe(II)3Fe(III) (-1.733 V), Fe(II)3Fe(III)/Fe(II)2Fe(III)2 (-0.727 V), and Fe(II)2Fe(III)2/Fe(II)Fe(III)3 (0.018 V) redox couples. Combined Mössbauer and crystallographic studies indicate that the change in oxidation state is exclusively localized at the triiron core, without changing the oxidation state of the apical metal center. This phenomenon is assigned to differences in the coordination environment of the two metal sites and provides a strategy for storing electron and hole equivalents without affecting the oxidation state of the coordinatively unsaturated metal. The presence of a ligand-binding site allowed the effect of redox modulation on nitric oxide activation by an Fe(II) metal center to be studied. Treatment of the clusters with nitric oxide resulted in binding of NO to the apical iron center, generating a {FeNO}(7) moiety. As with the NO-free precursors, the three reversible redox events are localized at the iron centers distal from the NO ligand. Altering the redox state of the triiron core resulted in significant change in the NO stretching frequency, by as much as 100 cm(-1). The increased activation of NO is attributed to structural changes within the clusters, in particular, those related to the interaction of the metal centers with the interstitial atom. The differences in NO activation were further shown to lead to differential reactivity, with NO disproportionation and N2O formation performed by the more electron-rich cluster.

Project description:Denitrifying NO reductases are transmembrane protein complexes that are evolutionarily related to heme/copper terminal oxidases. They utilize a heme/nonheme diiron center to reduce two NO molecules to N2O. Engineering a nonheme Fe(B) site within the heme distal pocket of sperm whale myoglobin has offered well-defined diiron clusters for the investigation of the mechanism of NO reduction in these unique active sites. In this study, we use FTIR spectroscopy to monitor the production of N2O in solution and to show that the presence of a distal Fe(B)(II) is not sufficient to produce the expected product. However, the addition of a glutamate side chain peripheral to the diiron site allows for 50% of a productive single-turnover reaction. Unproductive reactions are characterized by resonance Raman spectroscopy as dinitrosyl complexes, where one NO molecule is bound to the heme iron to form a five-coordinate low-spin {FeNO}(7) species with ν(FeNO)(heme) and ν(NO)(heme) at 522 and 1660 cm(-1), and a second NO molecule is bound to the nonheme Fe(B) site with a ν(NO)(FeB) at 1755 cm(-1). Stopped-flow UV-vis absorption coupled with rapid-freeze-quench resonance Raman spectroscopy provide a detailed map of the reaction coordinates leading to the unproductive iron-nitrosyl dimer. Unexpectedly, NO binding to Fe(B) is kinetically favored and occurs prior to the binding of a second NO to the heme iron, leading to a (six-coordinate low-spin heme-nitrosyl/FeB-nitrosyl) transient dinitrosyl complex with characteristic ν(FeNO)(heme) at 570 ± 2 cm(-1) and ν(NO)(FeB) at 1755 cm(-1). Without the addition of a peripheral glutamate, the dinitrosyl complex is converted to a dead-end product after the dissociation of the proximal histidine of the heme iron, but the added peripheral glutamate side chain in Fe(B)Mb2 lowers the rate of dissociation of the promixal histidine which in turn allows the (six-coordinate low-spin heme-nitrosyl/Fe(B)-nitrosyl) transient dinitrosyl complex to decay with production of N2O at a rate of 0.7 s(-1) at 4 °C. Taken together, our results support the proposed trans mechanism of NO reduction in NORs.

Dataset Information

Spectroscopic and computational study of a nonheme iron nitrosyl center in a biosynthetic model of nitric oxide reductase.

Publications

Spectroscopic and computational study of a nonheme iron nitrosyl center in a biosynthetic model of nitric oxide reductase.

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