Gene/Protein Disease Symptom Drug Enzyme Compound
Pivot Concepts: Gene/Protein Disease Symptom Drug Enzyme Compound   Target Concepts: Gene/Protein Disease Symptom Drug Enzyme Compound

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Query: EC:2.3.3.1 (citrate synthase)
4,488 document(s) hit in 31,850,051 MEDLINE articles (0.00 seconds)

Short-chain ubiquinone analogues act as electron acceptors and as inhibitors in the lymphoblast mitochondria of ND1/3460 mutants, which indicates structural changes in the ubiquinone-binding domain of Complex I in this mutant. The ND4/11778 mutant and two secondary ND5 mutants studied are associated with reductions of at least 50, 35 and 30% in the catalytic rate constant, respectively. However, the efficiency of oxidative phosphorylation is unaffected in all these ND mutants. The rate of respiration is only slightly limited by Complex I in lymphoblast mitochondria. Consequently, there is a 30-35% reduction in the electron flow through Complex I compared with that through Complex II, and an increased lactate/pyruvate ratio, in the ND1 and ND4 mutants, but these factors were unaffected in the secondary ND5 mutants. Energy metabolism is thus less severely affected in the secondary mutants than in the primary mutants, which supports the division into these two categories. An increased ubiquinone-10 content in the mitochondrial membrane of all the mutants, and enhanced succinate dehydrogenase and citrate synthase activities in the ND4 mutant, are proposed to be compensatory changes. The efficiency of these changes and the level of kinetic limitation of respiration by Complex I in each tissue are proposed to determine the clinical development of the disease.
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PMID:Catalytic activity of complex I in cell lines that possess replacement mutations in the ND genes in Leber's hereditary optic neuropathy. 870 9

Citrate synthase (citrate oxaloacetate-lyase, CoA-acetylating; EC 4.1.3.7, CS) was isolated and purified to homogeneity from a methylotrophic producer of polyhydroxybutyrate (PHB), Methylobacterium extorquens 15. The purification procedure includes streptomycin sulfate treatment of cell-free extract, ammonium sulfate fractionation, two steps of hydrophobic chromatography, and ion-exchange chromatography. The specific activity of the final enzyme preparation was 24 U/mg protein. The enzyme has apparent molecular weight 260 kD and consists of four 66-kD subunits. The enzyme shows a sigmoid saturation curve with CoASA (h = 1.3). Kinetic parameters are: K(m) = 84 microM for CoASA; K(m) = 12 microM for oxaloacetate; Vmax = 29.7 mumoles/min per mg protein. KCl at concentrations up to 80 mM activates the CS. ATP exerts a significant inhibitory effect on the enzyme activity, whereas NAD(P)H, isocitrate, alpha-ketoglutarate, ADP, acetoacetyl-CoA, glyoxylate, and glutamate have no influence. A possible role of the CS in coordinated control of CoASA transformation through the tricarboxylic acid cycle and PHB biosynthesis in this methylotroph is discussed.
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PMID:Purification and characterization of citrate synthase from Methylobacterium extorquens--a methylotrophic producer of polyhydroxybutyrate. 911 33

A gene encoding 544 amino acids for a subunit of group II chaperonin (thermosome) was cloned from a thermophilic methanogen, Methanococcus thermolithotrophicus. The deduced amino acid sequence showed 66.5, 56.1, and 20.1% similarities to those of Methanopyrus kandleri and Thermoplasma acidophilum and group I chaperonin of Escherichia coli, respectively. We call this chaperonin MTTS (M. thermolithotrophicus thermosome). The MTTS gene was expressed in E. coli. The purified recombinant MTTS seemed to be monomeric on gel filtration in the absence of Mg2+ and ATP. The monomer assembled to an oligomer (complex) in the presence of 50 mM MgCl2, 0.25 mM ATP, and 0.3 M (NH4)2SO4. It was eluted immediately before the elution volume of E. coli GroEL tetradecamer on gel filtration with a TSKgel G3000SWXL column. This reconstructed MTTS complex showed the cylindrical structure with two stacked rings in electron microscopy. The MTTS complex formed filamentous structures in the presence of Mg2+ and ATP at the protein concentration above 3.0 mg/ml. This filament formation was reversible. The MTTS filament was dissociated to the complex by dilution to the protein concentration of 0.2 mg/ml, even in the presence of Mg2+ and ATP. The MTTS complex exhibited weak ATPase activity with the hydrolysis rate of 74 mol of ATP hydrolysis/mol of MTTS complex/min at 70 degreesC. The MTTS complex promoted the refolding of chemically denatured thermophilic archaeal citrate synthase and glucose dehydrogenase at 50 degreesC in an ATP-dependent fashion. The analysis of nucleotide specificity of chaperone activity of MTTS suggested that it was coupled with hydrolysis of ATP, CTP, or UTP.
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PMID:Group II chaperonin in a thermophilic methanogen, Methanococcus thermolithotrophicus. Chaperone activity and filament-forming ability. 977 67

The archaeon Methanopyrus kandleri is the most thermophilic methanogen presently known. It contains a chaperonin (thermosome) which represents a 951 kDa homo-hexadecameric protein complex with NH4+-dependent ATPase activity. Since its synthesis is not increased upon heat shock, we set out to test its chaperone function. In order to obtain the chaperonin in amounts sufficient for functional investigations, the gene encoding the 60 kDa subunit was expressed in E. coili BL21 (DE3) cells. Purification yielded soluble, high-molecular-mass double-ring complexes, indistinguishable from the natural thermosome. In order to study the functional properties of the recombinant protein complex, pig citrate synthase, yeast alcohol dehydrogenase, yeast alpha-glucosidase, bovine insulin, and Thermotoga phosphoglycerate kinase were used as model substrates. The results demonstrate that the recombinant M. kandleri thermosome possesses a chaperone-like activity in vitro, inhibiting aggregation as the major off-pathway-reaction during thermal unfolding and refolding of proteins after chemical denaturation. However, the chaperonin only forms dead-end complexes with its non-native substrates, no release is detectable at temperatures between 25 and 60 degrees C.
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PMID:The recombinant thermosome from the hyperthermophilic archaeon Methanopyrus kandleri: in vitro analysis of its chaperone activity. 1006 37

Mitochondrial NAD-dependent (IDH) and cytosolic NADP-dependent isocitrate dehydrogenases have been considered as candidates for the production of 2-oxoglutarate required by the glutamine synthetase/glutamate synthase cycle. The increase in IDH transcripts in leaf and root tissues, induced by nitrate or NH4+ resupply to short-term N-starved tobacco (Nicotiana tabacum) plants, suggested that this enzyme could play such a role. The leaf and root steady-state mRNA levels of citrate synthase, acotinase, IDH, and glutamine synthetase were found to respond similarly to nitrate, whereas those for cytosolic NADP-dependent isocitrate dehydrogenase and fumarase responded differently. This apparent coordination occurred only at the mRNA level, since activity and protein levels of certain corresponding enzymes were not altered. Roots and leaves were not affected to the same extent either by N starvation or nitrate addition, the roots showing smaller changes in N metabolite levels. After nitrate resupply, these organs showed different response kinetics with respect to mRNA and N metabolite levels, suggesting that under such conditions nitrate assimilation was preferentially carried out in the roots. The differential effects appeared to reflect the C/N status after N starvation, the response kinetics being associated with the nitrate assimilatory capacity of each organ, signaled either by nitrate status or by metabolite(s) associated with its metabolism.
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PMID:Simultaneous expression of NAD-dependent isocitrate dehydrogenase and other krebs cycle genes after nitrate resupply to short-term nitrogen-starved tobacco 1039 6

A specially designed model reactor based on a 42-L laboratory fermentor was equipped with six stirrers (Rushton turbines) and five cylindrical disks. In this model reactor, the mixing time, Theta(90), turned out to be 13 times longer compared with the 42-L standard laboratory fermentor fitted with two Rushton turbines and four wall-fixed longitudinal baffles. To prove the suitability of the model reactor for scaledown studies of mixing-time-dependent processes, parallel exponential fed-batch cultivations were carried out with the leucine-auxotrophic strain, Corynebacterium glutamicum DSM 5715, serving as a microbial test system. L‐Leucine, the process-limiting substrate, was fed onto the liquid surface of both reactors. Cultivations were conducted using the same inoculum material and equal oxygen supply. The model reactor showed reduced sugar consumption (-14%), reduced ammonium consumption (-19%), and reduced biomass formation (-7%), which resulted in a decrease in L-lysine formation (-12%). These findings were reflected in less specific enzyme activity, which was determined for citrate synthase (CS), phosphoenolpyruvate carboxylase (PEP-C), and aspartate kinase (AK). The reduced specific activity of CS correlated with lower CO(2) evolution (-36%) during cultivation. The model reactor represents a valuable tool to simulate the conditions of poor mixing and inhomogeneous substrate distribution in bioreactors of industrial scale. Copyright 1999 John Wiley & Sons, Inc.
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PMID:A special reactor design for investigations of mixing time effects in a scaled-down industrial L-lysine fed-batch fermentation process 1040 40

Glucocorticoids near term are known to upregulate many important enzyme systems prior to birth. Glutamate dehydrogenase (GDH) is a mitochondrial enzyme that catalyzes both the reversible conversion of ammonium nitrogen into organic nitrogen (glutamate production) and the oxidative deamination of glutamate resulting in 2-oxoglutarate. The activity of this enzyme is considered to be of major importance in the development of catabolic conditions leading to gluconeogenesis prior to birth. Ovine hepatic GDH mRNA expression and activity were determined in near-term (130 days of gestation, term 147 +/- 4 days) control and acutely dexamethasone-treated (0.07 mg(-1) hr(-1) for 26 hr) fetuses. Dexamethasone infusion had no effect on placental or fetal liver weights. Dexamethasone infusion for 26 hr significantly increased hepatic GDH mRNA expression. This increased GDH mRNA expression was accompanied by an increase in hepatic mitochondrial GDH activity, from 30.0 +/- 7.4 to 58.2 +/- 8.1 U GDH/U CS (citrate synthase), and there was a significant correlation between GDH mRNA expression and GDH activity. The generated ovine GDH sequence displayed significant similarity with published human, rat, and murine GDH sequence. These data are consistent with the in vivo studies that have shown a redirection of glutamine carbon away from net hepatic glutamate release and into the citric acid cycle through the forward reaction catalyzed by GDH, i.e., glutamate to oxoglutarate.
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PMID:Induction of glutamate dehydrogenase in the ovine fetal liver by dexamethasone infusion during late gestation. 1252 80

The utilization of some agro-industrial wastes as soil conditioners to provide free-living nitrogen-fixing bacterial populations (e.g. Azospirillum spp.) with carbon and energy sources, may be an interesting perspective for agriculture. However, the presence of ammonium nitrogen in cultivated soils and/or various wastes could inhibit the growth of the nitrogen-fixing populations. The present investigation shows that growth of Azospirillum lipoferum was restricted at a dissolved oxygen (DO) concentration equal to 135 microM, when the initial NH4Cl concentration increased from 0.5 to 0.9 g/l. The activities of both citrate synthase (CS) and isocitrate dehydrogenase were significantly decreased in the presence of 0.9 g/l NH4Cl (e.g., 40% and 66%, respectively, in cells incubated for 95 h), while ammonium assimilation occurred via the glutamate dehydrogenase reaction. Furthermore, growth limitation occurred even in the presence of 0.5 g/l NH4Cl, when the DO concentration decreased from 135 to 30 microM. The activities of both CS and succinate dehydrogenase were dramatically decreased in cells grown at the lower DO concentration (e.g., 90% and 93% respectively, in a 95 h incubation), while ammonium assimilation was limited due to the low activities of both glutamate dehydrogenase and glutamate synthase. It is concluded that the threshold of ammonium concentration at which growth of A. lipoferum is limited, depends on the DO concentration in the medium.
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PMID:Metabolic activities in Azospirillum lipoferum grown in the presence of NH4+. 1276 47

Insulin resistance increases and muscle oxidative capacity decreases during aging, but lifestyle changes-especially physical activity-may reverse these trends. Here we report the effect of a 16-week aerobic exercise program (n = 65) or control activity (n = 37) performed by men and women aged 21-87 years on insulin sensitivity and muscle mitochondria. Insulin sensitivity, measured by intravenous glucose tolerance test, decreased with age (r = -0.32) and was related to abdominal fat content (r = -0.65). Exercise increased peak oxygen uptake (VO(2peak); 10%), activity of muscle mitochondrial enzymes (citrate synthase and cytochrome c oxidase, 45-76%) and mRNA levels of mitochondrial genes (COX4, ND4, both 66%) and genes involved in mitochondrial biogenesis (PGC-1alpha, 55%; NRF-1, 15%; TFAM, 85%). Exercise also increased muscle GLUT4 mRNA and protein (30-52%) and reduced abdominal fat (5%) and plasma triglycerides (25%). None of these changes were affected by age. In contrast, insulin sensitivity improved in younger people but not in middle-aged or older groups. Thus, the muscle mitochondrial response to 4 months of aerobic exercise training was similar in all age-groups, although the older people did not have an improvement in insulin sensitivity.
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PMID:Impact of aerobic exercise training on age-related changes in insulin sensitivity and muscle oxidative capacity. 1288 2

Based on the effects of different ammonium sulfate concentrations on meilingmycin biosynthesis, the results show that lower concentration of ammonium ions stimulates the biosynthesis of meilingmycin, while the concentration of higher than 5mmol/L inhibits the mycelial growth and the biosynthesis of the products. However, the sugar consumption rate increases with the elevating concentration of ammonium sulfate. On this basis, six enzymes, which are greatly related to the meilingmycin biosynthesis and the glucose metabolism, were measured and analyzed during the meilingmycin fermentation process. The results suggest that glucose-6-phosphate dehydrogenase, citrate synthase, succinate dehydrogenase and fatty acid synthase are stimulated by higher concentration of ammonium ions, while valine dehydrogenase and methylmalonyl-CoA carboxyltransferase are inhibited. From the results it follows that ammonium ions favors primary metabolism, that is, the HMP passway and the TCA circle is enhanced, as well as the source of the precursors for the biosynthesis of meilingmycin is restricted, which bring about the lower production of meilingmycin.
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PMID:[Regulatory effects of ammonium ions on the biosynthesis of meilingmycin]. 1598 36


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