Gene/Protein Disease Symptom Drug Enzyme Compound
Pivot Concepts:   Target Concepts:
Query: EC:2.3.3.1 (citrate synthase)
 4,488 document(s) hit in 31,850,051 MEDLINE articles (0.00 seconds)

To evaluate the peroxisomal requirement for beta-oxidation of hydroxyeicosatetraenoic acids (HETES), we tested 5-, 12- and 15-HETE oxidation in wild-type and mutant Chinese hamster ovary (CHO) cells. Mutant CHO cells contain peroxisomal ghosts, have random cytosolic localization of catalase and lack two of the enzymes necessary for peroxisomal beta-oxidation. Reverse-phase HPLC indicated that 33% of 12-HETE radioactivity was converted by wild-type CHO cells during a 2 h incubation to one major and several minor polar metabolites. Wild-type CHO cells also converted 15-HETE to one major and several minor polar metabolites. Neither 12- nor 15-HETE were converted to any metabolites by the mutant CHO cell lines, despite appreciable cellular uptake of these hydroxyeicosanoids. 5-HETE was not converted to any metabolic products by either the wild-type or the mutant CHO cells. Docosahexaenoic acid beta-oxidation was substantially reduced in the mutants as compared to the wild-type cells, palmitic acid beta-oxidation was reduced to an intermediate extent in the mutants, but octanoate beta-oxidation and citrate synthase activity were not impaired. Protein immunoblotting for mitochondrial manganese superoxide dismutase indicated a single band of identity at 20 kDa in both wild-type and mutant CHO cells. Since mutant CHO cells fail to convert 12- and 15-HETE to oxidative metabolites but contain normal mitochondrial enzymatic activities, intact peroxisomes appear to be the organelle responsible for HETE oxidation.
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PMID:Hydroxyeicosatetraenoic acid oxidation in Chinese hamster ovary cells: a peroxisomal metabolic pathway. 189 74

Local X-irradiation of mouse heart caused a large increase in manganese superoxide dismutase activity (MnSOD) in this organ but not in copper and zinc containing superoxide dismutase (Cu-Zn SOD) activity. MnSOD induction was both dose and time dependent. Another mitochondrial enzyme, citrate synthase, was not induced by X-irradiation. The amount of immunoreactive MnSOD also increased after X-irradiation, showing that the amount of MnSOD protein increased after X-irradiation. The response to X-irradiation was found to be biphasic--with one large peak and one smaller peak of manganese superoxide dismutase activity. The effect of various inhibitors of cellular activities on these two peaks of MnSOD activity was examined. Cycloheximide, a cytosolic protein synthesis inhibitor, abolished both peaks of MnSOD activity, while chloramphenicol, a mitochondrial protein synthesis inhibitor, has no effect on either peak. Actinomycin D, a RNA-synthesis inhibitor, lowered both peaks, but had more of an effect on the second peak than on the first. In vivo protein synthesis studies using [3H]arginine showed that an increase in new protein synthesis occurred during the time period of the second peak, but did not occur during the first peak. These results are consistent with the hypothesis that MnSOD induction occurs in two peaks with the first peak due to a preformed MnSOD protein or mRNA for MnSOD and the second peak due to an increase in new protein synthesis.
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PMID:Increase in manganese superoxide dismutase activity in the mouse heart after X-irradiation. 357 7

Amyotrophic lateral sclerosis is a fatal paralytic disorder of unknown cause. Recent evidence implicated the role of free radicals in the death of motor neurons in this disease. To investigate this hypothesis further, we measured the activity of the main free radical scavenging enzymes copper/zinc superoxide dismutase, manganese superoxide dismutase, catalase, and glutathione peroxidase in postmortem brain samples from 9 patients with sporadic amyotrophic lateral sclerosis and from 9 control subjects. We examined samples from the precentral gyrus of the cerebral cortex, a region affected in amyotrophic lateral sclerosis, and from the cerebellar cortex, a region not affected. The two groups did not differ in age or postmortem delay. In the precentral gyrus from amyotrophic lateral sclerosis samples, glutathione peroxidase activity as measured by spectrophotometric assay (13.8 +/- 2.6 nmol/min/mg protein [mean +/- standard error of mean]) was reduced significantly compared to the activity in the precentral gyrus from control samples (22.7 +/- 0.5 nmol/min/mg protein). In contrast, glutathione peroxidase activity was not significantly altered in the cerebellar cortex from amyotrophic lateral sclerosis patients compared to controls. Copper/zinc superoxide dismutase, manganese superoxide dismutase (corrected or not corrected for citrate synthase), and catalase were not significantly altered in the precentral gyrus or cerebellar cortex in the patient samples. This study indicated that glutathione peroxidase activity is reduced in a brain region affected in amyotrophic lateral sclerosis, thus suggesting that free radicals may be implicated in the pathogenesis of the disease.
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PMID:Brain superoxide dismutase, catalase, and glutathione peroxidase activities in amyotrophic lateral sclerosis. 896 46

Malignant mesothelioma cells contain elevated levels of manganese superoxide dismutase (MnSOD) and are highly resistant to oxidants compared to non-malignant mesothelial cells. Since the level of cellular free radicals may be important for cell survival, we hypothesized that the increase of MnSOD in the mitochondria of mesothelioma cells may alter the free radical levels of these organelles. First, MnSOD activity was compared to the activities of two constitutive mitochondrial enzymes; MnSOD activity was 20 times higher in the mesothelioma cells than in the mesothelial cells, whereas the activities of citrate synthase and cytochrome c oxidase did not differ significantly in the two cell lines. This indicates that the activity of MnSOD per mitochondrion was increased in the mesothelioma cells. Superoxide production was assayed in the isolated mitochondria of these cells using lucigenin chemiluminescence. Mitochondrial superoxide levels were significantly lower (72%) in the mesothelioma cells compared to the mesothelial cells. Oxidant production in intact cells, assayed by fluorimetry using 2',7'-dichlorodihydrofluorescein as a fluorescent probe, did not differ significantly between these cells. We conclude that mitochondrial superoxide levels are lower in mesothelioma cells compared to nonmalignant mesothelial cells, and that this difference may be explained by higher MnSOD activity in the mitochondria of these cells. Oxidant production was not different in these cells, which may be due to the previously observed increase in H2O2-scavenging mechanisms of mesothelioma cells.
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PMID:Generation of reactive oxygen species by human mesothelioma cells. 1038 73

We report here that estrogen (E(2)) modulates mitochondrial function in the vasculature. Mitochondrial dysfunction is implicated in the etiology of vascular disease; thus, vasoprotection by estrogen may involve hormonal effects on the mitochondria. To test this hypothesis, mitochondria were isolated from cerebral blood vessels obtained from ovariectomized female rats, with or without E(2) replacement. Estrogen receptor-alpha (ER-alpha) was detected in mitochondria by immunoblot and confocal imaging of intact vessels. E(2) treatment in vivo increased the levels of specific proteins in cerebrovascular mitochondria, such as ER-alpha, cytochrome c, subunit IV of complex IV, and manganese superoxide dismutase, all encoded in the nuclear genome, and subunit I of complex IV, encoded in the mitochondrial genome. Levels of glutathione peroxidase-1 and catalase, however, were not affected. Functional assays of mitochondrial citrate synthase and complex IV, key rate-limiting steps in energy production, showed that E(2) treatment increased enzyme activity. In contrast, mitochondrial production of hydrogen peroxide was decreased in vessels from E(2)-treated animals. In vitro incubation of cerebral vessels with 10 nM 17beta-estradiol for 18 h also elevated levels of mitochondrial cytochrome c. This effect was blocked by the estrogen receptor antagonist fulvestrant (ICI-182,780, Faslodex) but was unaffected by inhibitors of nitric-oxide synthase or phosphoinositide-3-kinase. Nuclear respiratory factor-1 protein, a primary regulator of nuclear gene-encoded mitochondrial genes, was significantly increased by long-term estrogen treatment in vivo. In summary, these novel findings suggest that vascular protection by E(2) is mediated, in part, by modulation of mitochondrial function, resulting in greater energy-producing capacity and decreased reactive oxygen species production.
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PMID:Estrogen increases mitochondrial efficiency and reduces oxidative stress in cerebral blood vessels. 1599 67

Post-translational modification of mitochondrial proteins by phosphorylation or dephosphorylation plays an essential role in numerous cell signaling pathways involved in regulating energy metabolism and in mitochondrion-induced apoptosis. Here we present a phosphoproteomic screen of the mitochondrial matrix proteins and begin to establish the protein phosphorylations acutely associated with calcium ions (Ca(2+)) signaling in porcine heart mitochondria. Forty-five phosphorylated proteins were detected by gel electrophoresis-mass spectrometry of Pro-Q Diamond staining, while many more Pro-Q Diamond-stained proteins evaded mass spectrometry detection. Time-dependent (32)P incorporation in intact mitochondria confirmed the extensive matrix protein phosphoryation and revealed the dynamic nature of this process. Classes of proteins that were detected included all of the mitochondrial respiratory chain complexes, as well as enzymes involved in intermediary metabolism, such as pyruvate dehydrogenase (PDH), citrate synthase, and acyl-CoA dehydrogenases. These data demonstrate that the phosphoproteome of the mitochondrial matrix is extensive and dynamic. Ca(2+) has previously been shown to activate various dehydrogenases, promote the generation of reactive oxygen species (ROS), and initiate apoptosis via cytochrome c release. To evaluate the Ca(2+) signaling network, the effects of a Ca(2+) challenge sufficient to release cytochrome c were evaluated on the mitochondrial phosphoproteome. Novel Ca(2+)-induced dephosphorylation was observed in manganese superoxide dismutase (MnSOD) as well as the previously characterized PDH. A Ca(2+) dose-dependent dephosphorylation of MnSOD was associated with an approximately 2-fold maximum increase in activity; neither the dephosphorylation nor activity changes were induced by ROS production in the absence of Ca(2+). These data demonstrate the use of a phosphoproteome screen in determining mitochondrial signaling pathways and reveal new pathways for Ca(2+) modification of mitochondrial function at the level of MnSOD.
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PMID:Mitochondrial matrix phosphoproteome: effect of extra mitochondrial calcium. 1648 45

We hypothesized that specific muscular transcript level adaptations participate in the improvement of endurance performances following intermittent hypoxia training in endurance-trained subjects. Fifteen male high-level, long-distance runners integrated a modified living low-training high program comprising two weekly controlled training sessions performed at the second ventilatory threshold for 6 wk into their normal training schedule. The athletes were randomly assigned to either a normoxic (Nor) (inspired O2 fraction = 20.9%, n = 6) or a hypoxic group exercising under normobaric hypoxia (Hyp) (inspired O2 fraction = 14.5%, n = 9). Oxygen uptake and speed at second ventilatory threshold, maximal oxygen uptake (VO2 max), and time to exhaustion (Tlim) at constant load at VO2 max velocity in normoxia and muscular levels of selected mRNAs in biopsies were determined before and after training. VO2 max (+5%) and Tlim (+35%) increased specifically in the Hyp group. At the molecular level, mRNA concentrations of the hypoxia-inducible factor 1alpha (+104%), glucose transporter-4 (+32%), phosphofructokinase (+32%), peroxisome proliferator-activated receptor gamma coactivator 1alpha (+60%), citrate synthase (+28%), cytochrome oxidase 1 (+74%) and 4 (+36%), carbonic anhydrase-3 (+74%), and manganese superoxide dismutase (+44%) were significantly augmented in muscle after exercise training in Hyp only. Significant correlations were noted between muscular mRNA levels of monocarboxylate transporter-1, carbonic anhydrase-3, glucose transporter-4, and Tlim only in the group of athletes who trained in hypoxia (P < 0.05). Accordingly, the addition of short hypoxic stress to the regular endurance training protocol induces transcriptional adaptations in skeletal muscle of athletic subjects. Expressional adaptations involving redox regulation and glucose uptake are being recognized as a potential molecular pathway, resulting in improved endurance performance in hypoxia-trained subjects.
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PMID:Exercise training in normobaric hypoxia in endurance runners. III. Muscular adjustments of selected gene transcripts. 1654 Jul 10

Diaphragmatic antioxidant enzymes are upregulated following acute and long-term treadmill exercise, but the effect of lifelong voluntary exercise (E) on diaphragmatic antioxidants is unknown. Therefore, 10-week old Fisher 344 rats were assigned to either: (a) sedentary ad libitum (AL) fed (24AL; n = 6); (b) E + 8% caloric restriction (24ECR; n = 9); or (c) sedentary + 8% caloric restriction (24CR; n = 9) groups. Diaphragms were harvested from animals at 24 months of age. Heme oxygenase-1 (HO-1) mRNA in addition to catalase (CAT), glutathione peroxidase (GPX), copper-zinc superoxide dismutase (Cu-ZnSOD) and manganese superoxide dismutase (MnSOD) mRNA and protein levels were measured. Reduced glutathione (GSH) and citrate synthase (CS) activity were measured to assess antioxidant status and oxidative capacity, respectively. The 24CR group demonstrated increased GPX, HO-1, MnSOD, and CAT mRNA compared to 24AL and 24ECR. Interestingly, the increased mRNA in 24CR animals did not result in elevated protein levels. No group differences in Cu-ZnSOD mRNA, CS activity, or GSH were observed, although GSH was 30% greater in 24CR animals (p = 0.085). In summary, although CR elevated the mRNA of key antioxidant enzymes in the diaphragm, lifelong CR alone or in combination with voluntary exercise did not alter diaphragm CS activity, antioxidant protein quantity, or GSH levels.
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PMID:Moderate caloric restriction increases diaphragmatic antioxidant enzyme mRNA, but not when combined with lifelong exercise. 1667 98

A myopathy characterized by mitochondrial pathology and oxidative stress is present in patients with peripheral arterial disease (PAD). Patients with PAD differ in disease severity, mode of presentation, and presence of comorbid conditions. In this study, we used a mouse model of hindlimb ischemia to isolate and directly investigate the effects of chronic inflow arterial occlusion on skeletal muscle microanatomy, mitochondrial function and expression, and oxidative stress. Hindlimb ischemia was induced by staged ligation/division of the common femoral and iliac arteries in C57BL/6 mice, and muscles were harvested 12 wk later. Muscle microanatomy was examined by bright-field microscopy, and mitochondrial content was determined as citrate synthase activity in muscle homogenates and ATP synthase expression by fluorescence microscopy. Electron transport chain (ETC) complexes I through IV were analyzed individually by respirometry. Oxidative stress was assessed as total protein carbonyls and 4-hydroxy-2-nonenal (HNE) adducts and altered expression and activity of manganese superoxide dismutase (MnSOD). Ischemic muscle exhibited histological features of myopathy and increased mitochondrial content compared with control muscle. Complex-dependent respiration was significantly reduced for ETC complexes I, III, and IV in ischemic muscle. Protein carbonyls, HNE adducts, and MnSOD expression were significantly increased in ischemic muscle. MnSOD activity was not significantly changed, suggesting MnSOD inactivation. Using a mouse model, we have demonstrated for the first time that inflow arterial occlusion alone, i.e., in the absence of other comorbid conditions, causes myopathy with mitochondrial dysfunction and increased oxidative stress, recapitulating the muscle pathology of PAD patients.
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PMID:Chronically ischemic mouse skeletal muscle exhibits myopathy in association with mitochondrial dysfunction and oxidative damage. 1848 Feb 38

Hypoxia, as one suboptimal environmental condition, can affect the physiological state of shrimp during pond aquaculture. To better understand the mechanism of response to hypoxic stress in Chinese shrimp Fenneropenaeus chinensis, proteome research approach was utilized. Differentially expressed proteins of hepatopancreas in adult Chinese shrimp between the control and hypoxia-stressed groups were screened. By 2-DE analysis, 67 spots showed obvious changes after hypoxia. Using LC-ESI-MS/MS, 51 spots representing 33 proteins were identified including preamylase, arginine kinase, phosphopyruvate hydratase, citrate synthase, ATP synthase alpha subunit, chymotrypsin BI, chitinase, ferritin, C-type lectin receptors, transketolase, formylglutathione hydrolase, formyltetrahydrofolate dehydrogenase, aldehyde dehydrogenase, glutathione peroxidase, cytosolic manganese superoxide dismutase, protein disulfide isomerase, beta-actin, oncoprotein nm23, crustacyanin-C1 and so on. These proteins could be functionally classified into several groups such as proteins related to energy production, metabolism-related proteins, immune-related proteins, antioxidant proteins, chaperones, cytoskeleton proteins and ungrouped proteins. The transcription levels of ten selected genes encode the identified proteins were analyzed by real-time PCR at different sampling times of hypoxia. This study is the first analysis of differentially expressed proteins in the hepatopancreas of shrimp after hypoxia and provides a new insight for further study in hypoxic stress response of shrimp at the protein level.
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PMID:Comparative proteomic profiles of the hepatopancreas in Fenneropenaeus chinensis response to hypoxic stress. 1957 23


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