Nitric-oxide Reductase

• Published in: The Journal of biological chemistry Vol. 277; no. 26; pp. 23407 - 23413
• Main Authors: Pinakoulaki, Eftychia, Gemeinhardt, Sabine, Saraste, Matti, Varotsis, Constantinos
• Format: Journal Article
• Language: English
• Published: Elsevier Inc 28.06.2002; American Society for Biochemistry and Molecular Biology
• ISSN: 0021-9258; 1083-351X
• DOI: 10.1074/jbc.M201913200

We have applied resonance Raman spectroscopy to investigate the properties of the dinuclear center of oxidized, reduced, and NO-bound nitric-oxide reductase from Paracoccus denitrificans. The spectra of the oxidized enzyme show two distinct νas(Fe-O-Fe) modes at 815 and 833 cm−1 of the heme/non-heme diiron center. The splitting of the Fe-O-Fe mode suggests that two different conformations (open and closed) are present in the catalytic site of the enzyme. We find evidence from deuterium exchange experiments that in the dominant conformation (833 cm−1mode, closed), the Fe-O-Fe unit is hydrogen-bonded to distal residue(s). The ferric nitrosyl complex of nitric-oxide reductase exhibits the ν(Fe3+-NO) and ν(N-O) at 594 and 1904 cm−1, respectively. The nitrosyl species we detect is photolabile and can be photolyzed to generate a new form of oxidized enzyme in which the proximal histidine is ligated to hemeb3, in contrast to the resting form. Photodissociation of the NO ligand yields a five-coordinate high-spin heme b3. Based on the findings reported here, the structure and properties of the dinuclear center of nitric- oxide reductase in the oxidized, reduced, and NO-bound form as well as its photoproduct can be described with certainty.