Concerted Nitric Oxide Formation and Release from the Simultaneous Reactions of Nitrite with Deoxy- and Oxyhemoglobin

Grubina, Rozalina; Huang, Zhi; Shiva, Sruti; Joshi, Mahesh S.; Azarov, Ivan; Basu, Swati; Ringwood, Lorna A.; Jiang, Alice; Hogg, Neil; Kim‐Shapiro, Daniel B.; Gladwin, Mark T.

doi:10.1074/jbc.m700546200

Cited by 139 publications

(192 citation statements)

References 59 publications

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“…1A are consistent with the intermediate formation of ferrylHb that achieves a measurable level during the rapid propagation phase. This is consistent with earlier observations using oxymyoglobin (14) and oxyhemoglobin (15). Finally, Fig.…”

Section: Methodssupporting

confidence: 82%

The Reaction between Nitrite and Oxyhemoglobin

Keszler

Piknová

Schechter

et al. 2008

Journal of Biological Chemistry

Self Cite

101

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The nitrite anion (NO 2 ؊ ) has recently received much attention as an endogenous nitric oxide source that has the potential to be supplemented for therapeutic benefit. One major mechanism of nitrite reduction is the direct reaction between this anion and the ferrous heme group of deoxygenated hemoglobin. However, the reaction of nitrite with oxyhemoglobin (oxyHb) is well established and generates nitrate and methemoglobin (metHb). Several mechanisms have been proposed that involve the intermediacy of protein-free radicals, ferryl heme, nitrogen dioxide (NO 2 ), and hydrogen peroxide (H 2 O 2 ) in an autocatalytic free radical chain reaction, which could potentially limit the usefulness of nitrite therapy. In this study we show that none of the previously published mechanisms is sufficient to fully explain the kinetics of the reaction of nitrite with oxyHb. Based on experimental data and kinetic simulation, we have modified previous models for this reaction mechanism and show that the new model proposed here is consistent with experimental data. The important feature of this model is that, whereas previously both H 2 O 2 and NO 2 were thought to be integral to both the initiation and propagation steps, H 2 O 2 now only plays a role as an initiator species, and NO 2 only plays a role as an autocatalytic propagatory species. The consequences of uncoupling the roles of H 2 O 2 and NO 2 in the reaction mechanism for the in vivo reactivity of nitrite are discussed.Elevated levels of nitrite in blood can trigger the oxidation of hemoglobin, leading to methemoglobinemia (1, 2). The mechanism of nitrite-dependent oxidation of deoxyhemoglobin (deoxyHb) 2 under anaerobic conditions, and its physiological potential in hypoxic vasodilation has recently been established (3-5). However, the mechanism of reaction between oxyhemoglobin (oxyHb) and nitrite has not yet been fully elucidated. Existing mechanistic approaches, starting with the earliest observations of Gamgee in 1868 (6), were recently reviewed (7). The end products of the reaction are methemoglobin (metHb) and nitrate. The kinetics of this reaction, when nitrite is in excess, are complex and exhibit an initial slow phase (often referred to as the lag phase) that accelerates into a rapid phase (referred to as the propagation phase) of oxidation. This reaction profile has been modeled as an autocatalytic, free radical chain reaction. The chain carrier radical and the autocatalytic steps have been variously hypothesized. Doyle et al. (8) found that hydrogen peroxide (H 2 O 2 ) accelerated oxyHb decay, whereas both catalase and superoxide dismutase elongated the half-time of the reaction. Based on these observations they suggested a mechanism according to which H 2 O 2 and superoxide played crucial roles. At the same time, Kosaka et al. (9) observed that catalase could extend the slow phase, but observed no effect of superoxide dismutase. In addition, they detected a transient protein free radical by EPR spectroscopy, and proposed a model with the intermediacy of H 2...

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Section: Methodssupporting

confidence: 82%

The Reaction between Nitrite and Oxyhemoglobin

Keszler

Piknová

Schechter

et al. 2008

Journal of Biological Chemistry

Self Cite

101

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“…In short, CO diffuses freely out of RBCs into the solution upon the addition of extracellular oxyHb, whereas no NO-export out of the RBCs-enclosed HbFe II NO was observed for three hours. These results confirm that: (1) the bioavailability of NO is not conserved under this condition, which is consistent with previous findings [47]; and (2) …”

Section: Conservation Of Co But Not No By Rbcssupporting

confidence: 93%

Interactions of nitrosylhemoglobin and carboxyhemoglobin with erythrocyte

2008

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Nitrosylhemoglobin (HbFe II NO) has been detected in vivo, and its role in NO transport and preservation has been discussed. To gain insight into the potential role of HbFe II NO, we performed in vitro experiments to determine the effect of oxygenated red blood cells (RBCs) on the dissociation of cell-free HbFe II NO, using carboxyhemoglobin (HbFe II CO) as a comparison. Results show that the apparent half-life of the cell-free HbFe II CO was reduced significantly in the presence of RBCs at 1 % hematocrit. In contrast, RBC did not change the apparent half-life of extracellular HbFe II NO, but caused a shift in the HbFe II NO dissociation product from methemoglobin (metHbFe III ) to oxyhemoglobin (HbFe II O 2 ). Extracellular hemoglobin was able to extract CO from HbFe II COcontaining RBC, but not NO from HbFe II NO-containing RBC. Although these results appear to suggest some unusual interactions between HbFe II NO and RBC, the data are explainable by simple HbFe II NO dissociation and hemoglobin oxidation with known rate constants. A kinetic model consisting of these reactions shows that i) deoxyhemoglobin is an intermediate in the reaction of HbFe II NO oxidation to metHbFe III , ii) the rate-limiting step of HbFe II NO decay is the dissociation of NO from HbFe II NO, iii) the magnitude of NO diffusion rate constant into RBC is estimated to be ∼10 4 M -1 s -1 , consistent with previous results determined from a competition assay, and iv) no additional chemical reactions are required to explain these data.

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“…This group includes hemoglobin and myoglobin (43)(44)(45)(46)(47). The rate of NO production by Cco increases with decreasing pH and increasing NO 2 Ϫ concentration (21), suggesting that that Cco functions in a nitrite reductase reaction (NO 2 Ϫ ϩ FeII ϩ H ϩ 3 NO ϩ FeIII ϩ OH Ϫ ) similar to that proposed for hemoglobin and myoglobin (48)(49)(50)(51)(52). Unlike the Cco oxidase reaction, which requires four electrons, the Cco NO 2 Ϫ reductase reaction involves a one-electron transfer.…”

Section: Discussionmentioning

confidence: 90%

Oxygen-regulated isoforms of cytochrome c oxidase have differential effects on its nitric oxide production and on hypoxic signaling

Castello

Woo

Ball

et al. 2008

Proc. Natl. Acad. Sci. U.S.A.

102

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Recently, it has been reported that mitochondria possess a novel pathway for nitric oxide (NO) synthesis. This pathway is induced when cells experience hypoxia, is nitrite (NO 2 ؊ )-dependent, is independent of NO synthases, and is catalyzed by cytochrome c oxidase (Cco). It has been proposed that this mitochondrially produced NO is a component of hypoxic signaling and the induction of nuclear hypoxic genes. In this study, we examine the NO 2 ؊ -dependent NO production in yeast engineered to contain alternative isoforms, Va or Vb, of Cco subunit V. Previous studies have shown that these isoforms have differential effects on oxygen reduction by Cco, and that their genes (COX5a and COX5b, respectively) are inversely regulated by oxygen. Here, we find that the Vb isozyme has a higher turnover rate for NO production than the Va isozyme and that the Vb isozyme produces NO at much higher oxygen concentrations than the Va isozyme. We have also found that the hypoxic genes CYC7 and OLE1 are induced to higher levels in a strain carrying the Vb isozyme than in a strain carrying the Va isozyme. Together, these results demonstrate that the subunit V isoforms have differential effects on NO 2 ؊ -dependent NO production by Cco and provide further support for a role of Cco in hypoxic signaling. These findings also suggest a positive feedback mechanism in which mitochondrially produced NO induces expression of COX5b, whose protein product then functions to enhance the ability of Cco to produce NO in hypoxic/anoxic cells.hypoxia ͉ mitochondria ͉ reactive oxygen species ͉ nitrite ͉ yeast I t has been known for quite some time that the mitochondrial respiratory chain is capable of generating reactive oxygen species (ROS) that account for much of the oxidative stress experienced by cells (1-3). The levels of these ROS increase when electron flow through the respiratory chain is inhibited by respiratory inhibitors (4-6) or altered by uncoupling electron transport from oxidative phosphorylation (7,8). Several studies have shown that exposure of cells and tissues to hypoxia increases ROS levels and oxidative stress (9-11). This increase in oxidative stress during exposure to hypoxia depends on a functional mitochondrial respiratory chain (10). It is currently unclear whether this increase is the result of increased generation or decreased destruction of ROS under hypoxic conditions. In yeast cells, the increase in ROS levels and oxidative stress is transient, as determined by shifting cells from normoxia to anoxia (10). During this shift, cells experience a continuum of decreasing oxygen concentrations and a transient increase in the levels of carbonylation of both mitochondrial and cytosolic protein, an increase in 8-OH-dG levels in mtDNA, and an increase in expression of the SOD1 gene. Many of the proteins that are carbonylated during a shift from normoxia to anoxia are the same proteins that are carbonylated when cells are exposed to menadione, a redox recycling agent that produces elevated intracellular levels of superoxide (12). ...

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Concerted Nitric Oxide Formation and Release from the Simultaneous Reactions of Nitrite with Deoxy- and Oxyhemoglobin

Cited by 139 publications

References 59 publications

The Reaction between Nitrite and Oxyhemoglobin

The Reaction between Nitrite and Oxyhemoglobin

Interactions of nitrosylhemoglobin and carboxyhemoglobin with erythrocyte

Oxygen-regulated isoforms of cytochrome c oxidase have differential effects on its nitric oxide production and on hypoxic signaling

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