共查询到20条相似文献,搜索用时 0 毫秒
1.
Winifred O. Idigo Svetlana Reilly Mei Hua Zhang Yin Hua Zhang Raja Jayaram Ricardo Carnicer Mark J. Crabtree Jean-Luc Balligand Barbara Casadei 《The Journal of biological chemistry》2012,287(52):43665-43673
Myocardial constitutive No production depends on the activity of both endothelial and neuronal NOS (eNOS and nNOS, respectively). Stimulation of myocardial β3-adrenergic receptor (β3-AR) produces a negative inotropic effect that is dependent on eNOS. We evaluated whether nNOS also plays a role in β3-AR signaling and found that the β3-AR-mediated reduction in cell shortening and [Ca2+]i transient amplitude was abolished both in eNOS−/− and nNOS−/− left ventricular (LV) myocytes and in wild type LV myocytes after nNOS inhibition with S-methyl-l-thiocitrulline. LV superoxide (O2˙̄) production was increased in nNOS−/− mice and reduced by l-Nω-nitroarginine methyl ester (l-NAME), indicating uncoupling of eNOS activity. eNOS S-glutathionylation and Ser-1177 phosphorylation were significantly increased in nNOS−/− myocytes, whereas myocardial tetrahydrobiopterin, eNOS Thr-495 phosphorylation, and arginase activity did not differ between genotypes. Although inhibitors of xanthine oxidoreductase (XOR) or NOX2 NADPH oxidase caused a similar reduction in myocardial O2˙̄, only XOR inhibition reduced eNOS S-glutathionylation and Ser-1177 phosphorylation and restored both eNOS coupled activity and the negative inotropic and [Ca2+]i transient response to β3-AR stimulation in nNOS−/− mice. In summary, our data show that increased O2˙̄ production by XOR selectively uncouples eNOS activity and abolishes the negative inotropic effect of β3-AR stimulation in nNOS−/− myocytes. These findings provide unequivocal evidence of a functional interaction between the myocardial constitutive NOS isoforms and indicate that aspects of the myocardial phenotype of nNOS−/− mice result from disruption of eNOS signaling. 相似文献
2.
目的:研究右美托咪定(DEX)对离体红细胞变形性的影响,探讨其作用机制。方法:采健康志愿者人血5 m L制成2%红细胞悬液,分成空白对照组(C组),低浓度DEX组(DL组)、中浓度DEX组(DM组)、高浓度DEX组(DH组),单纯育亨宾(Y组)以及育亨宾+DEX组(YD组),每组9例。将各组样本放入37℃恒温震荡培养箱中60 min后取出,测定红细胞变形性指数(EI)、红细胞内一氧化氮(NO)含量以及内皮型一氧化氮合酶(e NOS)含量。结果:与C组比较,DL组、DM组、DH组及YD组中EI、红细胞内NO、e NOS的含量增高(P0.05);Y组含量差异无统计学意义(P0.05)。与YD组比较,DM组EI、红细胞内NO及e NOS含量增高(P0.05)。结论:DEX可以提高离体红细胞的变形能力,其可能机制是通过激活红细胞膜上肾上腺素受体,激活红细胞内e NOS使细胞内NO增多有关。 相似文献
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4.
Nitric oxide and oxidative stress in brain and heart of normal rats treated with doxorubicin: role of aminoguanidine 总被引:2,自引:0,他引:2
Doxorubicin (DOX) is a potent antitumor antibiotic drug known to cause severe cardiac toxicity. Moreover, its adverse effects were found to be extended to the cerebral tissue. Several mechanisms for this toxicity have been ascribed. Currently, one of the most accepted mechanisms is through free radicals; however, the exact role of nitric oxide (NO) is still unclear. Accordingly, a NO-synthase inhibitor with some antioxidant property, aminoguanidine (AG), was selected to examine its potential protective effect against DOX-induced toxicity. Male Wistar albino rats (150-200 g) were allocated into a normal control group, DOX-induced toxicity group, and DOX + AG-treated group. DOX was injected i.p. at a dose of 10 mg/kg divided into four equal injections over a period of 2 weeks. AG was injected i.p. at a dose of 100 mg/kg 1 h before each DOX injection. The animals were sacrificed 24 h after the last DOX injection and the following parameters were measured: serum lactate dehydrogenase (LDH) and creatine phosphokinase (CPK) activities, cardiac and cerebral contents of malondialdehyde (MDA), conjugated diene (CD), glutathione (GSH), NO, and cytosolic calcium, as well as superoxide dismutase (SOD) and glutathione peroxidase (GSHP(X)) activities. Cardiotoxicity was manifested by a marked increase in serum LDH and CPK in addition to the sharp increase in MDA reaching eightfolds the basal level. This was accompanied by significant increase in CD, NO, cytosolic calcium, SOD, and GSHP(X) content/activity by 69, 85, 76, 125, and 41% respectively as compared to normal control. On the other hand, GSH was significantly depressed. In brain, only significant increase in MDA and GSHP(X) and decrease in GSH were obtained but to a lesser extent than the cardiac tissue. AG treatment failed to prevent the excessive release of cardiac enzymes; however, it alleviated the adverse effects of DOX in heart. AG administration resulted in marked decrease in the elevated levels of MDA, NO, SOD, and GSHP(X), however, MDA level was still pathological. The altered parameters in brain were restored by AG. It is concluded that, AG could not provide complete protection against DOX-induced toxicity. Therefore, it is recommended that, maintenance of the endogenous antioxidant, GSH, and regulation of calcium homeostasis must be considered, rather than NO formation, to guard against DOX-induced toxicity. 相似文献
5.
Freire-Garabal M Núñez MJ Riveiro P Balboa J López P Zamorano BG Rodrigo E Rey-Méndez M 《Life sciences》2002,72(2):173-183
We studied the effects of 1, 2, 5, 10 and 20 mg/kg of fluoxetine on the activity of phagocytosis in mice subjected to a chronic auditory stressor. Both the in vitro and in vivo activity of phagocytosis, measured using the zymosan-particle uptake method and the carbon clearance test, respectively, were reduced after 2, 4, 8 and 16 days of stress exposure. A partial recovery on the in vivo activity of phagocytosis was found on day 16th. Daily treatment with fluoxetine partially reversed the adverse effects of stress in a dose-dependent manner on both parameters but did not significantly affect the activity of phagocytosis in unstressed mice. Significant differences appeared when fluoxetine was administered at 2 mg/kg. Maximum effect was reached at 5 mg/kg. 相似文献
6.
Christina L. Grek Jie Zhang Yefim Manevich Danyelle M. Townsend Kenneth D. Tew 《The Journal of biological chemistry》2013,288(37):26497-26504
Post-translational S-glutathionylation occurs through the reversible addition of a proximal donor of glutathione to thiolate anions of cysteines in target proteins, where the modification alters molecular mass, charge, and structure/function and/or prevents degradation from sulfhydryl overoxidation or proteolysis. Catalysis of both the forward (glutathione S-transferase P) and reverse (glutaredoxin) reactions creates a functional cycle that can also regulate certain protein functional clusters, including those involved in redox-dependent cell signaling events. For translational application, S-glutathionylated serum proteins may be useful as biomarkers in individuals (who may also have polymorphic expression of glutathione S-transferase P) exposed to agents that cause oxidative or nitrosative stress. 相似文献
7.
Latika Nagpal Mohammad M. Haque Amit Saha Nirmalya Mukherjee Arnab Ghosh Brindaban C. Ranu Dennis J. Stuehr Koustubh Panda 《The Journal of biological chemistry》2013,288(27):19685-19697
Overproduction of nitric oxide (NO) by inducible nitric-oxide synthase (iNOS) has been etiologically linked to several inflammatory, immunological, and neurodegenerative diseases. As dimerization of NOS is required for its activity, several dimerization inhibitors, including pyrimidine imidazoles, are being evaluated for therapeutic inhibition of iNOS. However, the precise mechanism of their action is still unclear. Here, we examined the mechanism of iNOS inhibition by a pyrimidine imidazole core compound and its derivative (PID), having low cellular toxicity and high affinity for iNOS, using rapid stopped-flow kinetic, gel filtration, and spectrophotometric analysis. PID bound to iNOS heme to generate an irreversible PID-iNOS monomer complex that could not be converted to active dimers by tetrahydrobiopterin (H4B) and l-arginine (Arg). We utilized the iNOS oxygenase domain (iNOSoxy) and two monomeric mutants whose dimerization could be induced (K82AiNOSoxy) or not induced (D92AiNOSoxy) with H4B to elucidate the kinetics of PID binding to the iNOS monomer and dimer. We observed that the apparent PID affinity for the monomer was 11 times higher than the dimer. PID binding rate was also sensitive to H4B and Arg site occupancy. PID could also interact with nascent iNOS monomers in iNOS-synthesizing RAW cells, to prevent their post-translational dimerization, and it also caused irreversible monomerization of active iNOS dimers thereby accomplishing complete physiological inhibition of iNOS. Thus, our study establishes PID as a versatile iNOS inhibitor and therefore a potential in vivo tool for examining the causal role of iNOS in diseases associated with its overexpression as well as therapeutic control of such diseases. 相似文献
8.
Chen CA Lin CH Druhan LJ Wang TY Chen YR Zweier JL 《The Journal of biological chemistry》2011,286(33):29098-29107
An increase in production of reactive oxygen species resulting in a decrease in nitric oxide bioavailability in the endothelium contributes to many cardiovascular diseases, and these reactive oxygen species can oxidize cellular macromolecules. Protein thiols are critical reducing equivalents that maintain cellular redox state and are primary targets for oxidative modification. We demonstrate endothelial NOS (eNOS) oxidant-induced protein thiyl radical formation from tetrahydrobiopterin-free enzyme or following exposure to exogenous superoxide using immunoblotting, immunostaining, and mass spectrometry. Spin trapping with 5,5-dimethyl-1-pyrroline N-oxide (DMPO) followed by immunoblotting using an anti-DMPO antibody demonstrated the formation of eNOS protein radicals, which were abolished by superoxide dismutase and L-NAME, indicating that protein radical formation was due to superoxide generation from the eNOS heme. With tetrahydrobiopterin-reconstituted eNOS, eNOS protein radical formation was completely inhibited. Using mass spectrometric and mutagenesis analysis, we identified Cys-908 as the residue involved in protein radical formation. Mutagenesis of this key cysteine to alanine abolished eNOS thiyl radical formation and uncoupled eNOS, leading to increased superoxide generation. Protein thiyl radical formation leads to oxidation or modification of cysteine with either disulfide bond formation or S-glutathionylation, which induces eNOS uncoupling. Furthermore, in endothelial cells treated with menadione to trigger cellular superoxide generation, eNOS protein radical formation, as visualized with confocal microscopy, was increased, and these results were confirmed by immunoprecipitation with anti-eNOS antibody, followed by immunoblotting with an anti-DMPO antibody. Thus, eNOS protein radical formation provides the basis for a mechanism of superoxide-directed regulation of eNOS, involving thiol oxidation, defining a unique pathway for the redox regulation of cardiovascular function. 相似文献
9.
Schildknecht S Pape R Müller N Robotta M Marquardt A Bürkle A Drescher M Leist M 《The Journal of biological chemistry》2011,286(7):4991-5002
Minocycline prevents oxidative protein modifications and damage in disease models associated with inflammatory glial activation and oxidative stress. Although the drug has been assumed to act by preventing the up-regulation of proinflammatory enzymes, we probed here its direct chemical interaction with reactive oxygen species. The antibiotic did not react with superoxide or •NO radicals, but peroxynitrite (PON) was scavenged in the range of ∼1 μm minocycline and below. The interaction of pharmacologically relevant minocycline concentrations with PON was corroborated in several assay systems and significantly exceeded the efficacy of other antibiotics. Minocycline was degraded during the reaction with PON, and the resultant products lacked antioxidant properties. The antioxidant activity of minocycline extended to cellular systems, because it prevented neuronal mitochondrial DNA damage and glutathione depletion. Maintenance of neuronal viability under PON stress was shown to be solely dependent on direct chemical scavenging by minocycline. We chose α-synuclein (ASYN), known from Parkinsonian pathology as a biologically relevant target in chemical and cellular nitration reactions. Submicromolar concentrations of minocycline prevented tyrosine nitration of ASYN by PON. Mass spectrometric analysis revealed that minocycline impeded nitrations more effectively than methionine oxidations and dimerizations of ASYN, which are secondary reactions under PON stress. Thus, PON scavenging at low concentrations is a novel feature of minocycline and may help to explain its pharmacological activity. 相似文献
10.
Mauriz JL Linares P Macias RI Jorquera F Honrado E Olcoz JL González P González-Gallego J 《Free radical research》2003,37(8):841-848
The objective of this study is to determine if treatment with the angiogenesis inhibitor TNP-470 results in impairment of oxidative stress, inhibition of nuclear factor kappa B (NF-κB) activation and decrease of nitric oxide production in an experimental model of rat hepatocarcinogenesis. Tumour was induced by diethylnitrosamine and promoted by two-thirds hepatectomy plus acetaminofluorene administration. Experiments were carried out at 28 weeks after initiating the treatment. TNP-470 was administered at 30 mg/kg, three times per week from 20 to 28 weeks. Carcinomatous tissue growing outside dysplastic nodules and a marked expression of placental glutathione S-transferase were detected in rats with induced carcinogenesis. Liver concentrations of thiobarbituric acid reactive substances, reduced glutathione (GSH) and glutathione disulfide (GSSG) were significantly higher than those of controls and there was a significant increase in the GSSG/GSH ratio. Tumour growth was accompanied by augmented expression of inducible nitric oxide synthase, activation of (NF-κB) and proteolysis of IkappaB. All these effects were absent in animals receiving TNP-470. Our results indicate that TNP-470 inhibits oxidative stress, nitric oxide production and NF-κB activation induced by experimental hepatocarcinogenesis. These changes would contribute to the beneficial effects of TNP-470 in cancer treatment. 相似文献
11.
Chen W Li L Brod T Saeed O Thabet S Jansen T Dikalov S Weyand C Goronzy J Harrison DG 《The Journal of biological chemistry》2011,286(16):13846-13851
Tetrahydrobiopterin (BH(4)) is an essential co-factor for the nitric-oxide (NO) synthases, and in its absence these enzymes produce superoxide (O(2)(·-)) rather than NO. The rate-limiting enzyme for BH(4) production is guanosine triphosphate cyclohydrolase-1 (GTPCH-1). Because endogenously produced NO affects T cell function, we sought to determine whether antigen stimulation affected T cell GTPCH-1 expression and ultimately BH(4) levels. Resting T cells had minimal expression of inducible NOS (NOS2), endothelial NOS (NOS3), and GTPCH-1 protein and nearly undetectable levels of BH(4). Anti-CD3 stimulation of T cells robustly stimulated the coordinated expression of NOS2, NOS3, and GTPCH-1 and markedly increased both GTPCH-1 activity and T cell BH(4) levels. The newly expressed GTPCH-1 was phosphorylated on serine 72 and pharmacological inhibition of casein kinase II reduced GTPCH-1 phosphorylation and blunted the increase in T cell BH(4). Inhibition of GTPCH-1 with diaminohydroxypyrimidine (1 mmol/liter) prevented T cell BH(4) accumulation, reduced NO production, and increased T cell O(2)(·-) production, due to both NOS2 and NOS3 uncoupling. GTPCH-1 inhibition also promoted TH(2) polarization in memory CD4 cells. Ovalbumin immunization of mice transgenic for an ovalbumin receptor (OT-II mice) confirmed a marked increase in T cell BH(4) in vivo. These studies identify a previously unidentified consequence of T cell activation, promoting BH(4) levels, NO production, and modulating T cell cytokine production. 相似文献
12.
Karin Wetzelberger Shahid P. Baba Mahesh Thirunavukkarasu Ye-Shih Ho Nilanjana Maulik Oleg A. Barski Daniel J. Conklin Aruni Bhatnagar 《The Journal of biological chemistry》2010,285(34):26135-26148
Aldose reductase (AR) is a multifunctional enzyme that catalyzes the reduction of glucose and lipid peroxidation-derived aldehydes. During myocardial ischemia, the activity of AR is increased due to the oxidation of its cysteine residues to sulfenic acids. It is not known, however, whether the activated, sulfenic form of the protein (AR-SOH) is converted back to its reduced, unactivated state (AR-SH). We report here that in perfused mouse hearts activation of AR during 15 min of global ischemia is completely reversed by 30 min of reperfusion. During reperfusion, AR-SOH was converted to a mixed disulfide (AR-SSG). Deactivation of AR and the appearance of AR-SSG during reperfusion were delayed in hearts of mice lacking glutathione S-transferase P (GSTP). In vitro, GSTP accelerated glutathiolation and inactivation of AR-SOH. Reduction of AR-SSG to AR-SH was facilitated by glutaredoxin (GRX). Ischemic activation of AR was increased in GRX-null hearts but was attenuated in the hearts of cardiospecific GRX transgenic mice. Incubation of AR-SSG with GRX led to the regeneration of the reduced form of the enzyme. In ischemic cardiospecific AR transgenic hearts, AR was co-immunoprecipitated with GSTP, whereas in reperfused hearts, the association of AR with GRX was increased. These findings suggest that upon reperfusion of the ischemic heart AR-SOH is converted to AR-SSG via GSTP-assisted glutathiolation. AR-SSG is then reduced by GRX to AR-SH. Sequential catalysis by GSTP and GRX may be a general redox switching mechanism that regulates the reduction of protein sulfenic acids to cysteines. 相似文献
13.
Daniel R. Gonzalez Adriana V. Treuer Jorge Castellanos Raul A. Dulce Joshua M. Hare 《The Journal of biological chemistry》2010,285(37):28938-28945
S-Nitrosylation is a ubiquitous post-translational modification that regulates diverse biologic processes. In skeletal muscle, hypernitrosylation of the ryanodine receptor (RyR) causes sarcoplasmic reticulum (SR) calcium leak, but whether abnormalities of cardiac RyR nitrosylation contribute to dysfunction of cardiac excitation-contraction coupling remains controversial. In this study, we tested the hypothesis that cardiac RyR2 is hyponitrosylated in heart failure, because of nitroso-redox imbalance. We evaluated excitation-contraction coupling and nitroso-redox balance in spontaneously hypertensive heart failure rats with dilated cardiomyopathy and age-matched Wistar-Kyoto rats. Spontaneously hypertensive heart failure myocytes were characterized by depressed contractility, increased diastolic Ca2+ leak, hyponitrosylation of RyR2, and enhanced xanthine oxidase derived superoxide. Global S-nitrosylation was decreased in failing hearts compared with nonfailing. Xanthine oxidase inhibition restored global and RyR2 nitrosylation and reversed the diastolic SR Ca2+ leak, improving Ca2+ handling and contractility. Together these findings demonstrate that nitroso-redox imbalance causes RyR2 oxidation, hyponitrosylation, and SR Ca2+ leak, a hallmark of cardiac dysfunction. The reversal of this phenotype by inhibition of xanthine oxidase has important pathophysiologic and therapeutic implications. 相似文献
14.
Moonhee Lee Valerio Tazzari Daniela Giustarini Ranieri Rossi Anna Sparatore Piero Del Soldato Edith McGeer Patrick L. McGeer 《The Journal of biological chemistry》2010,285(23):17318-17328
The main lesion in Parkinson disease (PD) is loss of substantia nigra dopaminergic neurons. Levodopa (l-DOPA) is the most widely used therapy, but it does not arrest disease progression. Some possible contributing factors to the continuing neuronal loss are oxidative stress, including oxidation of l-DOPA, and neurotoxins generated by locally activated microglia and astrocytes. A possible method of reducing these factors is to produce l-DOPA hybrid compounds that have antioxidant and antiinflammatory properties. Here we demonstrate the properties of four such l-DOPA hybrids based on coupling l-DOPA to four different hydrogen sulfide-donating compounds. The donors themselves were shown to be capable of conversion by isolated mitochondria to H2S or equivalent SH− ions. This capability was confirmed by in vivo results, showing a large increase in intracerebral dopamine and glutathione after iv administration in rats. When human microglia, astrocytes, and SH-SY5Y neuroblastoma cells were treated with these donating agents, they all accumulated H2S intracellularly as did their derivatives coupled to l-DOPA. The donating agents and the l-DOPA hybrids reduced the release of tumor necrosis factor-α, interleukin-6, and nitric oxide from stimulated microglia, astrocytes as well as the THP-1 and U373 cell lines. They also demonstrated a neuroprotective effect by reducing the toxicity of supernatants from these stimulated cells to SH-SY5Y cells. l-DOPA itself was without effect in any of these assays. The H2S-releasing l-DOPA hybrid molecules also inhibited MAO B activity. They may be useful for the treatment of PD because of their significant antiinflammatory, antioxidant, and neuroprotective properties. 相似文献
15.
Leon Grayfer Erick Garcia Garcia Miodrag Belosevic 《The Journal of biological chemistry》2010,285(31):23537-23547
Unlike mammals, bony fish have two type II interferons, IFNγ and IFNγrel, whose pro-inflammatory functions have not been fully characterized. To elucidate the distinct roles of these type II interferons of bony fish, we examined the effects of recombinant goldfish (rg) IFNγ and IFNγrel on the macrophage antimicrobial responses, immune gene expression, and their signaling pathways. Our findings indicate that rgIFNγ and rgIFNγrel possess unique capacities to mediate each of the above processes. Q-PCR analysis revealed similar expression of both cytokines in tissues and immune cell populations of the goldfish, although IFNγ mRNA levels were generally higher in most tissues and cell types. Whereas rgIFNγ had long-lasting effects on the priming of goldfish monocyte ROI production, the rgIFNγrel had relatively short-lived ROI priming potential and eventually down-regulated the priming of ROI production induced by rgIFNγ or rgTNFα2. Whereas rgIFNγ induced relatively modest phagocytic and nitric oxide responses of goldfish macrophages, rgIFNγrel induced significantly higher phagocytosis, iNOSA and iNOSB gene expression and nitric oxide production compared with rgIFNγ. The rgIFNγ and rgIFNγrel induced different gene expression profiles in goldfish monocytes. These differences included significantly higher induction of TNFα2, CXCL8, ceruloplasmin, and interferon regulatory factor (IRFs) expression after activation of monocytes with rgIFNγrel. The rgIFNγrel was more abundant in whole cell lysates compared with rgIFNγ. Both cytokines induced the phosphorylation of Stat1, while the nuclear localization of Stat1 was only observed following treatment of monocytes with rgIFNγ. Our findings suggest the presence of functional segregation of the induction of macrophage antimicrobial functions by type II interferons of bony fish. 相似文献
16.
A large body of evidence supports a role for the NO-cGMP-protein kinase G pathway in the regulation of synaptic transmission
and plasticity, brain development and neuroprotection. Circumstancial evidence implicates natriuretic peptide-stimulated cGMP
formation in the same CNS functions. In addition to neurons, both cGMP-mediated pathways are functional in glial cells and
an increasing number of reports indicate that they may control important aspects of glial cell physiology relevant to neuronal
function. In this article we briefly review the regulation of cGMP formation in glial cells and summarize recent evidence
indicating that cGMP-mediated pathways can play important roles in astroglial and microglial function in normal and diseased
brain.
Special issue article in honor of Dr. Anna Maria Giuffrida-Stella. 相似文献
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18.
Regulation of human mitochondrial aldehyde dehydrogenase (ALDH-2) activity by electrophiles in vitro
Oelze M Knorr M Schell R Kamuf J Pautz A Art J Wenzel P Münzel T Kleinert H Daiber A 《The Journal of biological chemistry》2011,286(11):8893-8900
Recently, mitochondrial aldehyde dehydrogenase (ALDH-2) was reported to reduce ischemic damage in an experimental myocardial infarction model. ALDH-2 activity is redox-sensitive. Therefore, we here compared effects of various electrophiles (organic nitrates, reactive fatty acid metabolites, or oxidants) on the activity of ALDH-2 with special emphasis on organic nitrate-induced inactivation of the enzyme, the biochemical correlate of nitrate tolerance. Recombinant human ALDH-2 was overexpressed in Escherichia coli; activity was determined with an HPLC-based assay, and reactive oxygen and nitrogen species formation was determined by chemiluminescence, fluorescence, protein tyrosine nitration, and diaminonaphthalene nitrosation. The organic nitrate glyceryl trinitrate caused a severe concentration-dependent decrease in enzyme activity, whereas incubation with pentaerythritol tetranitrate had only minor effects. 4-Hydroxynonenal, an oxidized prostaglandin J(2), and 9- or 10-nitrooleate caused a significant inhibition of ALDH-2 activity, which was improved in the presence of Mg(2+) and Ca(2+). Hydrogen peroxide and NO generation caused only minor inhibition of ALDH-2 activity, whereas peroxynitrite generation or bolus additions lead to severe impairment of the enzymatic activity, which was prevented by the thioredoxin/thioredoxin reductase (Trx/TrxR) system. In the presence of glyceryl trinitrate and to a lesser extent pentaerythritol tetranitrate, ALDH-2 may be switched to a peroxynitrite synthase. Electrophiles of different nature potently regulate the enzymatic activity of ALDH-2 and thereby may influence the resistance to ischemic damage in response to myocardial infarction. The Trx/TrxR system may play an important role in this process because it not only prevents inhibition of ALDH-2 but is also inhibited by the ALDH-2 substrate 4-hydroxynonenal. 相似文献
19.
Shigemoto Fujii Tomohiro Sawa Hideshi Ihara Kit I. Tong Tomoaki Ida Tatsuya Okamoto Ahmed Khandaker Ahtesham Yu Ishima Hozumi Motohashi Masayuki Yamamoto Takaaki Akaike 《The Journal of biological chemistry》2010,285(31):23970-23984
A nitrated guanine nucleotide, 8-nitroguanosine 3′,5′-cyclic monophosphate (8-nitro-cGMP), is formed via nitric oxide (NO) and causes protein S-guanylation. However, intracellular 8-nitro-cGMP levels and mechanisms of formation of 8-nitro-cGMP and S-guanylation are yet to be identified. In this study, we precisely quantified NO-dependent formation of 8-nitro-cGMP in C6 glioma cells via liquid chromatography-tandem mass spectrometry. Treatment of cells with S-nitroso-N-acetylpenicillamine led to a rapid, transient increase in cGMP, after which 8-nitro-cGMP increased linearly up to a peak value comparable with that of cGMP at 24 h and declined thereafter. Markedly high levels (>40 μm) of 8-nitro-cGMP were also evident in C6 cells that had been stimulated to express inducible NO synthase with excessive NO production. The amount of 8-nitro-cGMP generated was comparable with or much higher than that of cGMP, whose production profile slightly preceded 8-nitro-cGMP formation in the activated inducible NO synthase-expressing cells. These unexpectedly large amounts of 8-nitro-cGMP suggest that GTP (a substrate of cGMP biosynthesis), rather than cGMP per se, may undergo guanine nitration. Also, 8-nitro-cGMP caused S-guanylation of KEAP1 in cells, which led to Nrf2 activation and subsequent induction of antioxidant enzymes, including heme oxygenase-1; thus, 8-nitro-cGMP protected cells against cytotoxic effects of hydrogen peroxide. Proteomic analysis for endogenously modified KEAP1 with matrix-assisted laser desorption/ionization time-of-flight-tandem mass spectrometry revealed that 8-nitro-cGMP S-guanylated the Cys434 of KEAP1. The present report is therefore the first substantial corroboration of the biological significance of cellular 8-nitro-cGMP formation and potential roles of 8-nitro-cGMP in the Nrf2-dependent antioxidant response. 相似文献
20.
Alvarez C Lozano-Juste J Romero LC García I Gotor C León J 《The Journal of biological chemistry》2011,286(1):578-586
The last step of sulfur assimilation is catalyzed by O-acetylserine(thiol)lyase (OASTL) enzymes. OASTLs are encoded by a multigene family in the model plant Arabidopsis thaliana. Cytosolic OASA1 enzyme is the main source of OASTL activity and thus crucial for cysteine homeostasis. We found that nitrating conditions after exposure to peroxynitrite strongly inhibited OASTL activity. Among OASTLs, OASA1 was markedly sensitive to nitration as demonstrated by the comparative analysis of OASTL activity in nitrated crude protein extracts from wild type and different oastl mutants. Furthermore, nitration assays on purified recombinant OASA1 protein led to 90% reduction of the activity due to inhibition of the enzyme, as no degradation of the protein occurred under these conditions. The reduced activity was due to nitration of the protein because selective scavenging of peroxynitrite with epicatechin impaired OASA1 nitration and the concomitant inhibition of OASTL activity. Inhibition of OASA1 activity upon nitration correlated with the identification of a modified OASA1 protein containing 3-nitroTyr(302) residue. The essential role of the Tyr(302) residue for the catalytic activity was further demonstrated by the loss of OASTL activity of a Y302A-mutated version of OASA1. Inhibition caused by Tyr(302) nitration on OASA1 activity seems to be due to a drastically reduced O-acetylserine substrate binding to the nitrated protein, and also to reduced stabilization of the pyridoxal-5'-phosphate cofactor through hydrogen bonds. This is the first report identifying a Tyr nitration site of a plant protein with functional effect and the first post-translational modification identified in OASA1 enzyme. 相似文献