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1.
Nitric oxide is a signaling molecule in intercellular communication as well as a powerful weapon used by macrophages to kill tumor cells and pathogenic bacteria. Here, we show that when Escherichia coli cells are exposed to nitric oxide, its ferredoxin [2Fe-2S] cluster is nitrosylated, forming the dinitrosyl iron complex with a characteristic EPR signal at g(av) = 2.04. Such formed ferredoxin dinitrosyl iron complex is efficiently repaired in E. coli cells even in the absence of new protein synthesis. However, the repair activity is completely inactivated once E. coli cells are disrupted, indicating that repairing the ferredoxin dinitrosyl iron complex requires cellular reducing equivalents. In search of such cellular factors, we find that l-cysteine can effectively eliminate the EPR signal of the ferredoxin dinitrosyl iron complex and release the ferrous iron from the complex. In contrast, N-acetyl-l-cysteine and reduced glutathione are much less effective. l-Cysteine seems to have a general function, since it can also remove the otherwise stable dinitrosyl iron complexes from proteins in the cell extracts prepared from the E. coli cells treated with nitric oxide. We propose that l-cysteine is responsible for removing the dinitrosyl iron complexes from the nitric oxide-modified proteins into which a new iron-sulfur cluster will be reassembled. 相似文献
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3.
Iron-sulfur proteins are among the sensitive targets of the nitric oxide cytotoxicity. When Escherichia coli cells are exposed to nitric oxide, iron-sulfur clusters are modified forming protein-bound dinitrosyl iron complexes. Such modified protein dinitrosyl iron complexes are stable in vitro but are efficiently repaired in aerobically growing E. coli cells even without any new protein synthesis. Here we show that cysteine desulfurase encoded by the gene iscS of E. coli can directly convert the ferredoxin dinitrosyl iron complex to the ferredoxin [2Fe-2S] cluster in the presence of L-cysteine in vitro. A reassembly of the [2Fe-2S] cluster in the ferredoxin dinitrosyl iron complex does not require any addition of iron or other protein components. Furthermore, a complete removal of the dinitrosyl iron complex from ferredoxin prevents reassembly of the [2Fe-2S] cluster in the protein. The results suggest that cysteine desulfurase (IscS) together with L-cysteine can efficiently repair the nitric oxide-modified ferredoxin [2Fe-2S] cluster and that the iron center in the dinitrosyl iron complex may be recycled for the reassembly of iron-sulfur clusters in proteins. 相似文献
4.
K. B. Shumaev S. A. Gubkina A. F. Vanin D. Sh. Burbaev V. P. Mokh A. F. Topunov E. K. Ruuge 《Biophysics》2013,58(2):172-177
It has been shown that interaction of cysteine dinitrosyl iron complexes with methylglyoxal leads to the formation of a new type of dinitrosyl iron complexes, EPR spectrum of these complexes essentially differs from spectra of dinitrosyl iron complexes containing unmodified thiol. The products of the cysteine reaction with methylglyoxal are hemithioacetals, Schiff bases and thiazolidines, which most likely serve as ligands for the new type of dinitrosyl iron complexes. It has been shown that the new type of dinitrosyl iron complexes as cysteine dinitrosyl iron complexes, which are physiological donors of nitric oxide, exert a vasodilator effect. It has also been found that the oxidative destruction of the new type of dinitrosyl iron complexes occurs at normal oxygen partial pressure, but these dinitrosyl iron complexes remain rather stable under hypoxia modeling. An assumption that the destruction of the new type of dinitrosyl iron complexes is caused by the formation of a bound peroxynitrite-containing intermediate is made. 相似文献
5.
A beneficial effect of dinitrosyl iron complexes (DNIC) with thiol-containing ligands on penile cavernus tissue was shown in rats subjected to penile denervation. Histological and histochemical investigations demonstrated that intracavernous injections of dinitrosyl iron complexes (2 times per one week during 6 months) blocked the reinforcement of endothelial cell proliferation in the tissue characteristic of the cavernous tissue when the penile nerve was removed. On the other hand, treatment with dinitrosyl iron complexes led to the preservation of mitotic activity of smooth myocytes and protected against the appearance in these cells of collagenase, an indicator of muscle transformation into fibrous tissue. It was shown that the process of fibrous transformation of myocytes correlates with a decrease in the mitotic activity of fibroblasts in the adventive part of cavernosa. The mitotic activity increased in cavernous tissue in the absence of dinitrosyl iron complexes. The efficiency of long-term action of dinitrosyl iron complexes on the erection in both intact animals and animals subjected to neuroectomy of cavernous tissue nerve was shown. The injection of low-molecular dinitrosyl iron complexes to the cavernous tissue resulted in the formation of protein-bound dinitrosyl iron complexes in the tissue, which were detected by the EPR technique. It is assumed that these dinitrosyl iron complexes function as a depot of nitric oxide, providing long-lasting penis erection. 相似文献
6.
A F Vanin 《Biofizika》1987,32(1):128-131
Treatment of perfused mouse liver with nitric oxide does not change the intensities of ESR signals of iron-sulfur proteins characteristic of this tissue. Proceeding from this evidence and also from the ratio between the iron content in these proteins and dinitrosyl iron complexes (complexes 2.03) formed in the liver when it contacts with NO, it is concluded that iron-sulfur proteins are not involved in the formation of complexes 2.03. It seems that only the loosely bound form of non-heme iron-free iron is involved in this process. 相似文献
7.
A F Vanin G B Men'shikov I A Moroz P I Mordvintcev V A Serezhenkov D Sh Burbaev 《Biochimica et biophysica acta》1992,1135(3):275-279
In cultured macrophages (J 774 line) a decrease in iron-sulfur centers (ISC) was not observed after 5 min treatment with nitric oxide (NO) (10(-7) M NO/10(7) cells). The content of these centers was measured by electron spin resonance (ESR) spectroscopy at 16-60 K. However, the appearance of a characteristic ESR signal at g(av) = 2.03 indicated the formation of dinitrosyl iron complex (DNIC) in these cells. These findings suggest that loosely bound non-heme iron (free iron) but not iron from ISC is mainly involved in DNIC formation. ISC might release iron for DNIC formation after their destruction induced by the products of NO oxidation (NO2, N2O3, etc). 相似文献
8.
H Hori F Masuya M Tsubaki S Yoshikawa Y Ichikawa 《The Journal of biological chemistry》1992,267(26):18377-18381
Low temperature photolysis of nitric oxide from the nitrosyl complexes of ferric cytochrome P450scc was examined by EPR spectroscopy to elucidate the stereochemical interaction between heme-bound ligand and side-chain of cholesterol or its hydroxylated analogues at the substrate-binding site. The photoproducts of the NO complexes trapped at 5 K exhibited new EPR absorptions providing information on the steric crowding of the distal heme moiety. Without substrate, the photoproduct exhibited a broad EPR absorption at g-8 due to magnetic dipole-dipole interaction between the photo-dissociated NO (S = 1/2) and the ferric iron (S = 5/2). This indicates that the photo-dissociated NO can move far away from the heme iron in the less restricted distal heme moiety of the substrate-free cytochrome P450scc. In the presence of substrates, such as cholesterol, 20(S)-hydroxycholesterol, 22(S)-hydroxycholesterol, 22(R)-hydroxycholesterol, and 25-hydroxycholesterol, the EPR spectra of the photoproducts exhibited many variations having broad g-8 absorptions and/or the widespread signals together with zero-field absorption. Among the steroid complexes used, 20(S)-hydroxycholesterol complex exhibited a conspicuously widespread EPR signal with a distinct zero-field absorption due to a spin-coupled interaction between the ferric iron (S = 5/2) and the photolyzed NO (S = 1/2). These results indicate that the 20(S)-hydroxycholesterol complex has restricted substrate-binding structure and that the hydroxylation of the cholesterol side-chain at the 22R position is necessary to proceed the side-chain cleavage reaction properly in cytochrome P450scc. 相似文献
9.
Although it is well known that Bcl-2 can prevent apoptosis, the Bcl-2's anti-apoptotic mechanism is not fully understood. Here, we investigate the mechanism of oxidant-induced cell death and to investigate the role of Bcl-2 in the tert-butyl hydroperoxide (t-BuOOH)-induced oxidant injury in Rat-1 fibroblasts and their bcl-2 transfected counterparts, b5 cells. Treatment with t-BuOOH causes mitochondrial disfunction and induced morphological features consistent with apoptosis more markedly in Rat-1 cells than in b5 cells. The hydroperoxide t-BuOOH at concentrations less than 100 nM for as long as 48 h or with higher concentrations (up to 100 microM) for only 3 h induces death in Rat-1 cells, whereas their bcl-2 transfectants were significantly resistant to cytotoxicity by both time and all concentration other than 100 microM. The similar results were obtained also for DNA strand cleavages as detected by TUNEL stain. The bcl-2 transfectants significantly suppressed t-BuOOH-induced increases in both lipid peroxidation and caspase-3 activation 3 and 1 h after t-BuOOH exposure, respectively, but failed to suppress either caspase-1 activation or an enhanced production of the intracellular reactive oxygen species (ROS). Intracellular uptake of [1-(14)C] ascorbic acid (Asc) into the bcl-2 transfectants was superior to that into the non-transfectants always under examined conditions regardless of serum addition to culture medium and cell density. Upregulation of Bcl-2 proteins was rapidly induced after t-BuOOH exposure in the transfectants, but not in non-transfectants, and restored till 24 h to the normal Bcl-2 level. Thus suppressions of both lipid peroxidation and the subsequent cell death events such as caspase-3 activation and DNA cleavage were concerned with the inhibitory effects of Bcl-2 on the t-BuOOH-induced cytotoxicity. And some of these events may correlate with Bcl-2 expression-induced partial enhanced anti-oxidant cellular ability including enrichment of intracellular Asc and oxidative stress-induced upregulation of Bcl-2 protein. On the other hand, ROS production and caspase-1 activation were not related to cytoprotection by Bcl-2. 相似文献
10.
The beneficial action of dinitrosyl iron complex with glutathione on conjunctive veins of eyes in rabbits with experimental thrombosis of conjunctive veins has been demonstrated. Aqueous solutions of dinitrosyl iron complexes were added subconjunctively at doses of 5.4-8.1 micromole per eye. The average duration of thrombosis by the action of dinitrosyl iron complex decreased from 6.4 days in control animals to 2 days. The addition of dinitrosyl iron complex resulted in blood flow recovery in occlusive vessels and prevented ischemia and necrosis of tissues. The enhancement of hemorrhagic activity induced by dinitrosyl iron complexes was abrogated with combined addition of the nonselective NO synthase inhibitor N-nitro-L-arginine. In contrast, S-nitrosoglutathione affected adversely the veins: the duration of thrombosis in experimental thrombosis of conjunctive veins increased to 7 days. Intensive hemorhage developed in the conjunctive. The formation of protein-bound dinitrosyl iron complexes was observed by the EPR method in eye tissues after the subconjunctive or parabulbar addition of dinitrosyl iron complex with glutathione. This was not the case when the complex was injected intravenously. It was shown that dinitrosyl iron complex with glutathione induces the blockade of pellet aggregation or strengthens the fibrinolytic activity of plasma of patients with eye vessel pathology. The beneficial action of dinitrosyl iron complexes on conjunctive veins was proposed to be due to the capacity of dinitrosyl iron complexes to donate NO primarily to its biological targets. The release of free NO molecules in large amounts is not characteristic for dinitrosyl iron complexes. This process is characteristic of S-nitrosoglutathione, which sharply increases the probability of the accumulation of peroxynitrite, which produces a toxic effect on cells and tissues. 相似文献
11.
L N Kubrina P I Mordvintsev A F Vanin 《Biulleten' eksperimental'no? biologii i meditsiny》1989,107(1):31-33
EPR evidence was obtained that more intensive formation of mononitrosyl non-heme iron complexes with diethyl-dithiocarbamate (DETC) took place in mouse liver when inflammation process was initiated in mice by the lipopolysaccharide isolated from Salmonella typhimurium bacterium wall DETC intraperitoneally injected bound with endogenous non-heme iron resulted with DETC-Fe complex formation. These complexes were as a traps of nitric oxide appeared in animal tissues, and NO-Fe-DETC complexes were observed. Phenazone known as a free radical process inhibitor lowered NO production in animal organism. The free radical processes were suggested to intensify under inflammation reactions and to cause the various amino groups oxidation to nitroso groups which were capable to release free nitric oxide. 相似文献
12.
Endogenous superoxide production and the nitrite/nitrate ratio control the concentration of bioavailable free nitric oxide in leaves 总被引:7,自引:0,他引:7
Vanin AF Svistunenko DA Mikoyan VD Serezhenkov VA Fryer MJ Baker NR Cooper CE 《The Journal of biological chemistry》2004,279(23):24100-24107
We have quantitatively measured nitric oxide production in the leaves of Arabidopsis thaliana and Vicia faba by adapting ferrous dithiocarbamate spin tapping methods previously used in animal systems. Hydrophobic diethyldithiocarbamate complexes were used to measure NO interacting with membranes, and hydrophilic N-methyl-d-glucamine dithiocarbamate was used to measure NO released into the external solution. Both complexes were able to trap levels of NO, readily detectable by EPR spectroscopy. Basal rates of NO production (in the order of 1 nmol g(-) (1) h(-1)) agreed with previous studies. However, use of methodologies that corrected for the removal of free NO by endogenously produced superoxide resulted in a significant increase in trapped NO (up to 18 nmol g(-) (1) h(-1)). Basal NO production in leaves is therefore much higher than previously thought, but this is masked by significant superoxide production. The effects of nitrite (increased rate) and nitrate (decreased rate) are consistent with a role for nitrate reductase as the source of this basal NO production. However, rates under physiologically achievable nitrite concentrations never approach that reported following pathogen induction of plant nitric-oxide synthase. In Hibiscus rosa sinensis, the addition of exogenous nitrite generated sufficient NO such that EPR could be used to detect its production using endogenous spin traps (forming paramagnetic dinitrosyl iron complexes). Indeed the levels of this nitrosylated iron pool are sufficiently high that they may represent a method of maintaining bioavailable iron levels under conditions of iron starvation, thus explaining the previously observed role of NO in preventing chlorosis under these conditions. 相似文献
13.
The formation of dinitrosyl iron complexes with thiol-containing ligands in plant tissues (parsley and apple leaves) in the presence of nitric monoxide was demonstrated using electron paramagnetic resonance. In two types of tissues dinitrosyl iron complexes are predominantly represented by the binuclear diamagnetic form. This diamagnetic form can be transformed in EPR-detectable mononitrosyl iron complexes with diethyldithiocarbamate due to the ability of diethyldithiocarbamate to accept the iron-mononitrosyl groups from iron-dinitrosyl fragments of binuclear complexes. A similar transformation was observed under the effect of diethyldithiocarbamate on a mononuclear paramagnetic form of dinitrosyl iron complexes. The significant amount of binuclear dinitrosyl iron complexes found in plant tissues suggests that these complexes can be considered as a “working form” of nitric monoxide, which is recognized now as a universal regulator of metabolic processes in plants as well as in other organisms. 相似文献
14.
V. A. Serezhenkov I. S. Kuznetsov T. I. Romantsova M. I. Kuznetsova A. F. Vanin 《Biophysics》2011,56(6):1088-1095
It is shown that metformin, which is a drug used for treatment of type 2 diabetes mellitus, is metabolized in vivo in the
intestine and liver of mice with the release of nitric oxide. Subsequently the released nitric oxide forms paramagnetic mono-
and dinitrosyl iron complexes which can be registered by EPR. It is suggested that nitric oxide is responsible for the multifarious
therapeutic action of metformin such as lowering of blood glucose level, reduction of arterial hypertension, and other biological
effects. 相似文献
15.
T Ochi 《Mutation research》1989,213(2):243-248
The effects of iron chelators and glutathione (GSH) depletion on the induction of chromosomal aberrations by tert-butyl hydroperoxide (t-BuOOH) were investigated in cultured Chinese hamster V79 cells. t-BuOOH in a concentration range of 0.1-1.0 mM induced chromosomal structural aberrations, consisting mainly of chromatid gaps and breaks, in a dose-dependent fashion. The divalent iron chelator o-phenanthroline almost completely suppressed the formation of chromosomal aberrations while the trivalent chelator desferrioxamine was less effective. GSH depletion did not affect the formation of chromosomal aberrations and DNA single-strand breaks (ssb) by t-BuOOH. DNA ssb by 0.5 mM t-BuOOH were repaired within 60 min of treatment in both GSH-depleted (GSH-) and non-depleted (GSH+) cells. In contrast, chromosomal aberrations increased a little during the 60 min after treatment in both GSH- and GSH+ cells. The aberrations were then repaired in GSH+ cells but those in GSH- cells were maintained to a great extent during 20 h of post-treatment incubation. These results indicate that divalent iron mediates the induction of chromosomal aberrations by t-BuOOH. That t-BuOOH-induced chromosomal aberrations remain even after DNA ssb were repaired suggests involvement of other lesions than DNA ssb in the formation of chromosomal aberrations by the hydroperoxide. 相似文献
16.
It is shown that metformin, which is a drug used for treatment of type 2 diabetes mellitus, metabolizes itself in vivo in the intestine and liver of mice with the release of nitric oxide. Subsequently the released nitric oxide forms paramagnetic mono- and dinitrosyl iron complexes which can be registered by EPR method. It is suggested that nitric oxide is just responsible for multifarious therapeutic action of metformin such as lowering of blood glucose level, reduction of arterial hypertension and other biological effects. 相似文献
17.
The effect of iron dinitrosyl complexes, S-nitrosoglutathione, and glutathione on free radical oxidation of rat heart mitochondria induced by tert-butyl hydroperoxide and metmyoglobin or their combination with ferritin was studied. It was shown that iron dinitrosyl complexes or the combination of S-nitrosoglutathione and glutathione inhibited most effectively the peroxidation of mitochondrial membranes. It was found that ferritin stimulated the prooxidant action of metmyoglobin. Using EPR spectroscopy, it was established that, in conditions of O2*- generation, the destruction of iron dinitrosyl complexes took place. Iron dinitrosyl complexes also inhibited the formation of thiyl radicals, which appeared during O2*- generation in the system containing glutathione and S-nitrosoglutathione. It is essential that the formation of iron dinitrosyl complexes in this reaction system took place with the involvement of ferritin. It was proposed that the prooxidant action of ferritin and myoglobin could be inverted to the antioxidant one. 相似文献
18.
Shumaev KB Gubkin AA Gubkina SA Gudkov LL Lakomkin VL Topunov AF Vanin AF Ruuge EK 《Biofizika》2007,52(3):534-538
It has been established that albumin-bound dinitrosyl iron complexes can be destroyed by superoxide radicals generated in a xanthine-xanthine oxidase system. It was shown that peroxynitrite also effectively destroyed albumin-bound dinitrosyl iron complexes. At the same time, hydrogen peroxide and tert-butyl hydroperoxide did not stimulate the destruction of albumin-bound dinitrosyl iron complexes up to concentrations one order higher than the content of NO. The data have been obtained indicating that dinitrosyl iron complexes possess the vasodilatory activity. It has been proposed that peroxynitrite and superoxide radical, by causing the destruction of albumin-bound dinitrosyl iron complexes, affect the physiological properties of nitric oxide. 相似文献
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20.
F X van Leeuwen F J Zuyderhoudt B F van Gelder J van Gool 《Biochimica et biophysica acta》1977,500(2):304-309
EPR spectra of the water-insoluble iron fraction, hemosiderin of human and rat liver are described. The homogenate of freshly prepared perfused rat liver shows a non-heme iron signal at g=4.3 and a high-spin heme-iron signal around g=6, whereas the washed and sonicated sample of the insoluble iron fraction shows solely a non-heme iron signal at g=4.3. This indicates that hemosiderin from rat liver does not contain heme iron. Human-liver preparations from post mortem obtained material show in the homogenates as well as in the washed and sonicated samples an intense high-spin heme iron signal at g=6.0 and a non-heme iron signal at g=4.3. A comparative experiment, carried out with "aged" rat liver preparations, reveals the same spectra as with the human preparations. It is concluded that that the heme present in the insoluble iron fraction is caused by degradation of hemoglobin in the obduction material, and that heme is not a constituent of the insoluble depot iron. 相似文献