Abstract:Summary. In this article, the coordinating behavior of nitric oxide (NO) as a biological ligand for hemeproteins and enzymes is presented by reference to the spectral and stereochemical properties of nitrosylhemeproteins and their model nitrosylheme. The structural change in the heme coordination environments on NO binding to hemeproteins is described primarily in relation to the mechanisms of activation and inhibition of the enzymes by NO.
“…These K d values are double that of the low-affinity phase (∼33 μM) observed with cytochrome bo . Furthermore, NO acts as a strong field ligand, and so its interaction with ferric hemes yields products that have been established by EPR experiments to be low spin ( S = 0) ( , ). The optical spectra of the Fe III heme nitrosyls are also characteristic of low-spin heme and completely different from the spectra of high-spin Fe III hemes ().…”
Section: Discussionmentioning
confidence: 99%
“…This evidence strongly attests to the sequential formation of Cu B II mono- and dinitrosyls. Note that binding of NO to the Fe III heme is expected to drive the heme group low-spin ( , ). 5 Titration of fast cytochrome bo with NO monitored by EPR spectroscopy.…”
Section: Stoichiometry Of No Binding To Fast Cytochrome Bomentioning
The reaction of nitric oxide (NO) with fast cytochrome bo from Escherichia coli has been studied by electronic absorption, MCD, and EPR spectroscopy. Titration of the enzyme with NO showed the formation of two distinct species, consistent with NO binding stoichiometries of 1:1 and 2:1 with observed dissociation constants at pH 7.5 of approximately 2.3 x 10(-)6 and 3.3 x 10(-)5 M. Monitoring the titration by EPR spectroscopy revealed that the broad EPR signals at g approximately 7.3, 3.7, and 2.8 due to magnetic interaction between high-spin heme o (S = 5/2) and CuBII (S = 1/2) are lost. A high-spin heme o signal at g = 6.0 appears as the 1:1 complex is formed but is lost again on formation of the 2:1 complex, which is EPR silent. The absorption spectrum shows that heme o remains in the high-spin FeIII state throughout the titration. These results are consistent with the binding of up to two NO molecules at CuBII. This has been confirmed by studies with the Cl- adduct of fast cytochrome bo. MCD evidence shows that heme o remains ligated by histidine and water. Addition of excess NO to the Cl- adduct leads to the appearance of a high-spin FeIII heme EPR signal. Hence chloride ion binds to CuB, blocking the binding of a second NO molecule. These results suggest a mechanism for the reduction of NO to nitrous oxide by cytochrome bo and cytochrome c oxidase in which the binding of two cis NO molecules at CuB permits the formation of an N-N bond and the abstraction of oxygen by the heme group.
“…These K d values are double that of the low-affinity phase (∼33 μM) observed with cytochrome bo . Furthermore, NO acts as a strong field ligand, and so its interaction with ferric hemes yields products that have been established by EPR experiments to be low spin ( S = 0) ( , ). The optical spectra of the Fe III heme nitrosyls are also characteristic of low-spin heme and completely different from the spectra of high-spin Fe III hemes ().…”
Section: Discussionmentioning
confidence: 99%
“…This evidence strongly attests to the sequential formation of Cu B II mono- and dinitrosyls. Note that binding of NO to the Fe III heme is expected to drive the heme group low-spin ( , ). 5 Titration of fast cytochrome bo with NO monitored by EPR spectroscopy.…”
Section: Stoichiometry Of No Binding To Fast Cytochrome Bomentioning
The reaction of nitric oxide (NO) with fast cytochrome bo from Escherichia coli has been studied by electronic absorption, MCD, and EPR spectroscopy. Titration of the enzyme with NO showed the formation of two distinct species, consistent with NO binding stoichiometries of 1:1 and 2:1 with observed dissociation constants at pH 7.5 of approximately 2.3 x 10(-)6 and 3.3 x 10(-)5 M. Monitoring the titration by EPR spectroscopy revealed that the broad EPR signals at g approximately 7.3, 3.7, and 2.8 due to magnetic interaction between high-spin heme o (S = 5/2) and CuBII (S = 1/2) are lost. A high-spin heme o signal at g = 6.0 appears as the 1:1 complex is formed but is lost again on formation of the 2:1 complex, which is EPR silent. The absorption spectrum shows that heme o remains in the high-spin FeIII state throughout the titration. These results are consistent with the binding of up to two NO molecules at CuBII. This has been confirmed by studies with the Cl- adduct of fast cytochrome bo. MCD evidence shows that heme o remains ligated by histidine and water. Addition of excess NO to the Cl- adduct leads to the appearance of a high-spin FeIII heme EPR signal. Hence chloride ion binds to CuB, blocking the binding of a second NO molecule. These results suggest a mechanism for the reduction of NO to nitrous oxide by cytochrome bo and cytochrome c oxidase in which the binding of two cis NO molecules at CuB permits the formation of an N-N bond and the abstraction of oxygen by the heme group.
“…Recently, lipopolysaccaride and interferon-γ have been used to produce a septic-shock model; the generated NO was trapped either in the abdomen of a living mouse or in the head of a living rat, using the Fe-(DETC) 2 complex (Yoshimura et al 1997).…”
Section: Esr Spectroscopy/imaging Of Endogenous Free Radicalsmentioning
Much outstanding progress concerning the application of ESR spectroscopy/imaging in the biomedical field has been made in recent years. The literature in this field has already been specifically covered by several reviews. The aim of this article is to provide an overview of the most important findings, obtained in the last four years, in the detection and localization of different exogenous free radicals, as well as of endogenous free radicals in diverse experimental animal models.
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