Inducible Nitric-oxide Synthase Generates Superoxide from the Reductase Domain

Xia, Yong; Roman, Linda J.; Masters, Bettie Sue Siler; Zweíer, Jay L.

doi:10.1074/jbc.273.35.22635

Cited by 366 publications

(230 citation statements)

References 29 publications

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“…The production of NO from L-arginine by NOS occurs via two sequential monooxygenation events, consuming 1.5 equivalents of NADPH for every NO produced [12]. Our laboratory and several others have demonstrated that besides synthesizing NO, all three isoforms of NOS can also generate O 2 .-, depending on substrate and cofactor availability [13][14][15][16][17][18]. With low levels or total absence of L-arginine, purified nNOS catalyzes the reduction of oxygen, with O 2 .-production [13,14].…”

Section: Introductionmentioning

confidence: 99%

Dose dependent effects of reactive oxygen and nitrogen species on the function of neuronal nitric oxide synthase

Sun

Druhan

Zweíer

2008

Archives of Biochemistry and Biophysics

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Reactive nitrogen species (RNS) and oxygen species (ROS) have been reported to modulate the function of nitric oxide synthase (NOS); however, the precise dosedependent effects of specific RNS and ROS on NOS function are unknown. Questions remain unanswered regarding whether pathophysiological levels of RNS and ROS alter NOS function, and if this alteration is reversible. We measured the effects of peroxynitrite (ONOO -), superoxide (O 2 .-), hydroxyl radical ( . OH), and H 2 O 2 on nNOS activity. The results showed that NO production was inhibited in a dosedependent manner by all four oxidants, but only O 2 . -and ONOO -were inhibitory at pathophysiological concentrations (≤ 50 μM). Subsequent addition of tetrahydrobiopterin (BH 4 ) fully restored activity after O 2 .-exposure, while BH 4 partially rescued the activity decrease induced by the other three oxidants. Furthermore, treatment with either ONOO -or O 2 .-stimulated nNOS uncoupling with decreased NO and enhanced O 2 .-generation. Thus, nNOS is reversibly uncoupled by O 2 .-(≤ 50 μM), but irreversibly uncoupled and inactivated by ONOO -. Additionally, we observed that the mechanism by which oxidative stress alters nNOS activity involves not only BH 4 oxidation, but also nNOS monomerization as well as possible degradation of the heme.

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Section: Introductionmentioning

confidence: 99%

Dose dependent effects of reactive oxygen and nitrogen species on the function of neuronal nitric oxide synthase

Sun

Druhan

Zweíer

2008

Archives of Biochemistry and Biophysics

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“…NOS1 co-localizes with xanthine oxidase [4], a superoxide (O 2 .-) producing enzyme, and may potentially lead to the production of low levels of peroxynitrite (ONOO − ), as nitric oxide and superoxide react with a very high rate constant [5]. When expressed, NOS2 is capable of producing large amounts of both nitric oxide and superoxide [6,7]. Additionally, NADPH oxidase and xanthine oxidoreductase increase superoxide production during these pathophysiological conditions of the myocardium [8,9], thus leading to the formation of high levels of peroxynitrite.…”

Section: Introductionmentioning

confidence: 99%

Biphasic effect of SIN-1 is reliant upon cardiomyocyte contractile state

Kohr¹,

Wang²,

Wheeler³

et al. 2008

Free Radical Biology and Medicine

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Many studies have demonstrated a biphasic effect of peroxynitrite in the myocardium, but few studies have investigated this biphasic effect on β-adrenergic responsiveness and its dependence on contractile state. We have previously shown that high SIN-1 (source of peroxynitrite, 200 μmol/L) produced significant anti-adrenergic effects during maximal β-adrenergic stimulation in cardiomyocytes. In the current study, we hypothesize that the negative effects of high SIN-1 will be greatest during high contractile states, while the positive effects of low SIN-1 (10 μmol/L) will predominate during low contractility. Isolated murine cardiomyocytes were field stimulated at 1 Hz and [Ca 2+ ] i transients and shortening were recorded. Following submaximal ISO (β-adrenergic agonist, 0.01 μmol/L) stimulation, 200 μmol/L SIN-1 induced two distinct phenomena. Cardiomyocytes undergoing a large response to ISO showed a significant reduction in contractility, while cardiomyocytes exhibiting a modest response to ISO showed a further increase in contractility. Additionally, 10 μmol/L SIN-1 always increased contractility during low ISO stimulation, but had no effect during maximal ISO (1 μmol/L) stimulation. 10 μmol/L SIN-1 also increased basal contractility. Interestingly, SIN-1 only produced a contractile effect under one condition in phospholamban knockout cardiomyocytes, providing a potential mechanism for the biphasic effect of peroxynitrite. These results provide clear evidence for a biphasic effect of peroxynitrite, with high peroxynitrite modulating high levels of β-adrenergic responsiveness and low peroxynitrite regulating basal function and low levels of β-adrenergic stimulation.

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“…After hearts were subjected to periods of global ischemia (30,45, or 60 min) followed by 30 min of reflow, basal CF values were measured and compared with the preischemic values. CF was decreased after ischemia, and this reduction in flow was proportional to the duration of ischemia (Fig.…”

Section: Effect Of Ischemia/reperfusion On Coronary Flow (Cf)mentioning

confidence: 99%

Myocardial ischemia results in tetrahydrobiopterin (BH ₄ ) oxidation with impaired endothelial function ameliorated by BH ₄

Dumitrescu¹,

Biondi

Xia³

et al. 2007

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

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Coronary vasodilation is impaired in the postischemic heart with a loss of endothelial nitric oxide synthase (eNOS) activity, but the mechanisms underlying ischemia-induced eNOS dysfunction are not understood. For nitric oxide (NO) synthesis, eNOS requires the redox-sensitive cofactor tetrahydrobiopterin (BH 4); however, the role of BH 4 in ischemia-induced endothelial dysfunction remains unknown. Therefore, isolated rat hearts were subjected to varying durations of ischemia, and the alterations in NOS-dependent vasodilation were measured and correlated with assays of eNOS activity and cardiac BH 4 concentrations. Ischemia timedependently decreased cardiac BH 4 content with 85, 95, or 97% irreversible degradation after 30, 45, or 60 min of ischemia, respectively. Paralleling the decreases in BH 4, reductions of eNOS activity were seen of 58, 86, or 92%, and NOS-derived superoxide production was greatly increased. Addition of 10 M BH4 enhanced eNOS activity in nonischemic hearts and partially restored activity after ischemia. It also suppressed NOS-derived superoxide production. Impaired coronary flow during postischemic reperfusion was improved by BH 4 infusion. Thus, BH4 depletion contributes to postischemic eNOS dysfunction, and BH 4 treatment is effective in partial restoration of endothelium-dependent coronary flow. Supplementation of BH 4 may therefore be an important therapeutic approach to reverse endothelial dysfunction in postischemic tissues.ischemia reperfusion injury ͉ nitric oxide ͉ nitric oxide synthase uncoupling ͉ superoxide ͉ vascular function N itric oxide synthase (NOS) converts L-arginine and O 2 to nitric oxide (NO) and L-citrulline. This enzymatic process consumes NADPH and requires Ca 2ϩ /calmodulin, flavin adenine dinucleotide, flavin mononucleotide, and tetrahydrobiopterin (BH 4 ) as NOS cofactors. Endothelial NO synthase (eNOS) contributes to the regulation of vasomotor tone and blood pressure by producing NO that activates soluble guanylate cyclase in vascular smooth muscle, resulting in vasorelaxation (1-3).Endothelial dysfunction is a prognostic marker of cardiovascular disease (4). It has been suggested that limited availability of BH 4 contributes to eNOS dysfunction in hypercholesterolemia, diabetes, atherosclerosis, hypertension, and heart failure (5-9). It was also observed previously that eNOS function is impaired in ischemic hearts (10). In vivo coronary artery occlusion triggers endothelial dysfunction and decreased eNOSdependent vasoreactivity, although reactivity is preserved to exogenous NO (11,12). Endothelial-dependent coronary vasoreactivity is impaired in hearts subjected to periods of global ischemia and reperfusion (10). Endothelium-dependent vasodilators induce a relatively high increase in coronary flow in control hearts or in those made ischemic for short times, but longer periods of ischemia result in an abrupt decline in vasodilatory response.In addition to impairing eNOS-mediated NO formation, BH 4 depletion may have additional detrimental effects in postischem...

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Inducible Nitric-oxide Synthase Generates Superoxide from the Reductase Domain

Abstract: In the absence of L-arginine, the heme center of the oxygenase domain of neuronal nitric-oxide synthase reduces molecular oxygen to superoxide (O 2 .

Cited by 366 publications

References 29 publications

Dose dependent effects of reactive oxygen and nitrogen species on the function of neuronal nitric oxide synthase

Dose dependent effects of reactive oxygen and nitrogen species on the function of neuronal nitric oxide synthase

Biphasic effect of SIN-1 is reliant upon cardiomyocyte contractile state

Myocardial ischemia results in tetrahydrobiopterin (BH ₄ ) oxidation with impaired endothelial function ameliorated by BH ₄

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Inducible Nitric-oxide Synthase Generates Superoxide from the Reductase Domain

Abstract: In the absence of L-arginine, the heme center of the oxygenase domain of neuronal nitric-oxide synthase reduces molecular oxygen to superoxide (O 2 .

Cited by 366 publications

References 29 publications

Dose dependent effects of reactive oxygen and nitrogen species on the function of neuronal nitric oxide synthase

Dose dependent effects of reactive oxygen and nitrogen species on the function of neuronal nitric oxide synthase

Biphasic effect of SIN-1 is reliant upon cardiomyocyte contractile state

Myocardial ischemia results in tetrahydrobiopterin (BH 4 ) oxidation with impaired endothelial function ameliorated by BH 4

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Myocardial ischemia results in tetrahydrobiopterin (BH ₄ ) oxidation with impaired endothelial function ameliorated by BH ₄