Deoxymyoglobin Is a Nitrite Reductase That Generates Nitric Oxide and Regulates Mitochondrial Respiration

Abstract

Abstract-Previous studies have revealed a novel interaction between deoxyhemoglobin and nitrite to generate nitric oxide (NO) in blood. It has been proposed that nitrite acts as an endocrine reservoir of NO and contributes to hypoxic vasodilation and signaling. Here, we characterize the nitrite reductase activity of deoxymyoglobin, which reduces nitrite approximately 36 times faster than deoxyhemoglobin because of its lower heme redox potential. We hypothesize that physiologically this reaction releases NO in proximity to mitochondria and regulates respiration through cytochrome c oxidase. Spectrophotometric and chemiluminescent measurements show that the deoxymyoglobin-nitrite reaction produces NO in a second order reaction that is dependent on deoxymyoglobin, nitrite and proton concentration, with a bimolecular rate constant of 12.4 mol/L -1 s -1 (pH 7.4, 37°C). Because the IC 50 for NO-dependent inhibition of mitochondrial respiration is approximately 100 nmol/L at physiological oxygen tensions (5 to 10 mol/L); we tested whether the myoglobin-dependent reduction of nitrite could inhibit respiration. Indeed, the addition of deoxymyoglobin and nitrite to isolated rat heart and liver mitochondria resulted in the inhibition of respiration, while myoglobin or nitrite alone had no effect. The addition of nitrite to rat heart homogenate containing both myoglobin and mitochondria resulted in NO generation and inhibition of respiration; these effects were blocked by myoglobin oxidation with ferricyanide but not by the xanthine oxidoreductase inhibitor allopurinol. These data expand on the paradigm that heme-globins conserve and generate NO via nitrite reduction along physiological oxygen gradients, and further demonstrate that NO generation from nitrite reduction can escape heme autocapture to regulate NO-dependent signaling. (Circ Res. 2007;100:654-661.)

Nitrite reductase activity of myoglobin regulates respiration and cellular viability in myocardial ischemia-reperfusion injury

“…Whereas XOR has been proposed as being responsible for the conversion of nitrite to NO • in the heart by Webb et al (11), Zweier et al (19), and recently Baker et al (20), no such effect was observed by our groups (14,15). We have recently demonstrated that a decrease in myocardial oxygen tension drives the conversion of oxymyoglobin to deoxyMb, which reacts with nitrite to generate NO • , regulating cardiac energetics and function (14,15). Whether deoxyMb or XOR is responsible for nitrite reduction to NO • in the ischemic heart to mediate cytoprotection remains unclear.…”

“…The exact mechanisms behind the cytoprotective effects of exogenous nitrite, however, are still a matter of debate. Whereas XOR has been proposed as being responsible for the conversion of nitrite to NO • in the heart by Webb et al (11), Zweier et al (19), and recently Baker et al (20), no such effect was observed by our groups (14,15). We have recently demonstrated that a decrease in myocardial oxygen tension drives the conversion of oxymyoglobin to deoxyMb, which reacts with nitrite to generate NO • , regulating cardiac energetics and function (14,15).…”

“…In myoglobin ϩ/ϩ hearts, NO • generation from nitrite inhibits mitochondrial respiration during ischemia, which may allow the tissue to conserve the little oxygen present. In contrast, the lack of inhibition in myoglobin Ϫ/Ϫ hearts may lead to a steeper oxygen gradient within the tissue and hence a greater oxygen deficit deeper in the tissue (14).…”

“…Nitrite is present at concentrations of 0.3-1.0 M in plasma and 1-20 M in tissue (3)(4)(5), and has been proposed to participate in hypoxic vasodilation and signaling (6)(7)(8)(9). In vivo, nitrite may be converted to NO • by several mechanisms including acidic disproportionation (10), and enzymatic conversion via xanthine-oxidoreductase (XOR) (11), mitochondrial enzymes (12), deoxyhemoglobin (6,9,13), and deoxymyoglobin (deoxyMb) (14,15).…”

Mitochondrial dysfunction and oxidative stress in the pathogenesis of alcohol- and obesity-induced fatty liver diseases

“…good or bad NO, the complex interplay between NO and mitochondria remains to be defined in the context of fatty liver disease from alcohol and obesity. Moreover, the critical importance of understanding the regulation of the mitochondrial respiratory chain by NO in hepatic physiology and pathophysiology is supported by several studies demonstrating that localized NO production can occur within the organelle through a specific mitochondrial NOS isoform or via metabolism of nitrite to NO [119][120][121].…”