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Query: UNIPROT:P17174 (aspartate aminotransferase)
 14,872 document(s) hit in 31,850,051 MEDLINE articles (0.00 seconds)

beta-Methyleneaspartate, a specific inhibitor of aspartate aminotransferase (EC 2.6.1.1.), was used to investigate the role of the malate-aspartate shuttle in rat brain synaptosomes. Incubation of rat brain cytosol, "free" mitochondria, synaptosol, and synaptic mitochondria, with 2 mM beta-methyleneaspartate resulted in inhibition of aspartate aminotransferase by 69%, 67%, 49%, and 76%, respectively. The reconstituted malate-aspartate shuttle of "free" brain mitochondria was inhibited by a similar degree (53%). As a consequence of the inhibition of the aspartate aminotransferase, and hence the malate-aspartate shuttle, the following changes were observed in synaptosomes: decreased glucose oxidation via the pyruvate dehydrogenase reaction and the tricarboxylic acid cycle; decreased acetylcholine synthesis; and an increase in the cytosolic redox state, as measured by the lactate/pyruvate ratio. The main reason for these changes can be attributed to decreased carbon flow through the tricarboxylic acid cycle (i.e., decreased formation of oxaloacetate), rather than as a direct consequence of changes in the NAD+/NADH ratio. Malate/glutamate oxidation in "free" mitochondria was also decreased in the presence of 2 mM beta-methyleneaspartate. This is probably a result of decreased glutamate transport into mitochondria as a result of low levels of aspartate, which are needed for the exchange with glutamate by the energy-dependent glutamate-aspartate translocator.
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PMID:Influence of the malate-aspartate shuttle on oxidative metabolism in synaptosomes. 336 10

In lymphocytes of the rat, pyruvate kinase, phosphoenolpyruvate carboxykinase and NADP+-linked malate dehydrogenase (decarboxylating) are distributed almost exclusively in the cytosol whereas pyruvate carboxylase is distributed almost entirely in the mitochondria. For NAD+-linked malate dehydrogenase and aspartate aminotransferase approximately 80% and 40%, respectively, are in the cytosolic compartment. Since glutaminase is present in the mitochondria, glutamine is converted to malate within the mitochondria but further metabolism of the malate is likely to occur in the cytosol. Hence pyruvate produced from this malate, via oxaloacetate and phosphoenolpyruvate carboxykinase, may be rapidly converted to lactate, so restricting the entry of pyruvate into the mitochondria and explaining why very little glutamine is completely oxidised in these cells despite a high capacity of the Krebs cycle.
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PMID:Intracellular distribution of some enzymes of the glutamine utilisation pathway in rat lymphocytes. 374 15

The role of glucocorticoids in the increase by cold-exposure of the effect of alanine on the malate-aspartate shuttle was studied in perfused rat liver. The capacity of the shuttle was evaluated by measurement of changes in both the rate of glucose production from sorbitol and the ratio of lactate to pyruvate during ethanol oxidation (Biomed. Res. 6, Suppl., 1986). The effect of alanine on the shuttle capacity was decreased by adrenalectomy. When 1.5 mg/kg dexamethasone sulfate was administrated to adrenalectomized rats kept at 24 or 4 degrees C, once daily for 5 days, the effect of alanine on the shuttle increased its capacity to the level of sham-operated rats that had been exposed to 4 degrees C for 5 days. The effects of dexamethasone were blocked by the coadministration of tetracycline with the agent. Cold exposure and steroid replacement had little effect on the alanine-induced elevation of the levels of aspartate, glutamate, and alpha-ketoglutarate in liver cells. The increase of the effect of alanine could not be explained only by changes in the activity of NAD+ malate dehydrogenase and aspartate aminotransferase. The results suggest that cold exposure and replacement treatment with glucocorticoids modulate equally the effect of alanine on the capacity of the malate-aspartate shuttle.
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PMID:Effects of alanine on malate-aspartate shuttle in perfused livers from cold-exposed rats. 376 24

The RS-isomers of beta-mercapto-alpha-ketoglutarate, beta-methylmercapto-alpha-ketoglutarate and beta-methylmercapto-alpha-hydroxyglutarate have been synthesized. Beta-Mercapto-alpha-ketoglutarate was a potent inhibitor, competitive with isocitrate and noncompetitive with NADP+, of the mitochondrial NADP-specific isozyme from pig heart (Ki = 5 nM; Km (DL-isocitrate)/Ki(RS-beta-mercapto-alpha-ketoglutarate) = 650) and pig liver, the cytosolic isozyme from pig liver (I0.5 = 23 nM), and the NADP-linked enzymes from yeast (Ki = 58 nM) and Escherichia coli (Ki = 58 nM) at pH 7.4 and with Mg2+ as activator. beta-Mercapto-alpha-ketoglutarate was also an effective inhibitor of NADP-isocitrate-dehydrogenase activity in intact liver mitochondria. beta-Mercapto-alpha-ketoglutarate was a much less potent inhibitor for heart NAD-isocitrate dehydrogenase (Ki = 520 nM) than for the NADP-specific enzyme. beta-Methylmercapto-alpha-ketoglutarate (I0.5 = 10 microM) was a much less effective inhibitor than the beta-mercapto derivative for heart NADP-isocitrate dehydrogenase. The beta-sulfur substituted alpha-ketoglutarates were substrates for the oxidation of NADPH by heart NADP-isocitrate dehydrogenase without requiring CO2. beta-Methylmercapto-alpha-hydroxyglutarate, the expected product of reduction of beta-methylmercapto-alpha-ketoglutarate, did not cause reduction of NADP+ but it was an inhibitor competitive with isocitrate for NADP-isocitrate dehydrogenase. The beta-sulfur substituted alpha-ketoglutarate derivatives were alternate substrates for alpha-ketoglutarate dehydrogenase and the cytosolic and mitochondrial isozymes of heart aspartate aminotransferase but had no effect on glutamate dehydrogenase or alanine aminotransferase.
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PMID:beta-Sulfur substituted alpha-ketoglutarates as inhibitors and alternate substrates for isocitrate dehydrogenases and certain other enzymes. 394 94

Current evidence suggests that mitochondrial matrix enzymes exist in solid-state, multienzyme complexes in vivo. Addition of polyethylene glycol to a solution containing malate dehydrogenase and citrate synthase generates such a solid-state, enzyme complex in vitro at enzyme concentrations permitting kinetic measurements. Suspensions of the isolated, solid-state, hetero-complex of these enzymes were used to study the coupled reactions of citrate synthesis from malate, NAD, and CoASAc. The particles appear to be about 1 microgram in diameter. Considering the ratio of enzyme to oxalacetate molecules in or at the surface of the solid-state particles, one would expect oxalacetate to be converted to citrate within a few molecular distances of the site of oxalacetate generation. This model of "substrate channeling" (or alternatively a direct transfer of oxalacetate between enzymes) is supported by experiments with excess aspartate aminotransferase and glutamate added to the solution phase to give a reaction competing with the synthase for bulk phase oxalacetate. Quantities of aminotransferase that reduce the citrate reaction rate with soluble dehydrogenase and synthase by 90% do not significantly affect rates with comparable amounts of the dehydrogenase-synthase complex. We suggest that similar substrate channeling can occur in vivo and discuss the possible advantages provided thereby.
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PMID:Substrate channeling of oxalacetate in solid-state complexes of malate dehydrogenase and citrate synthase. 406 62

1. A method is described for extracting separately mitochondrial and extramitochondrial enzymes from fat-cells prepared by collagenase digestion from rat epididymal fat-pads. The following distribution of enzymes has been observed (with the total activities of the enzymes as units/mg of fat-cell DNA at 25 degrees C given in parenthesis). Exclusively mitochondrial enzymes: glutamate dehydrogenase (1.8), NAD-isocitrate dehydrogenase (0.5), citrate synthase (5.2), pyruvate carboxylase (3.0); exclusively extramitochondrial enzymes: glucose 6-phosphate dehydrogenase (5.8), 6-phosphogluconate dehydrogenase (5.2), NADP-malate dehydrogenase (11.0), ATP-citrate lyase (5.1); enzymes present in both mitochondrial and extramitochondrial compartments: NADP-isocitrate dehydrogenase (3.7), NAD-malate dehydrogenase (330), aconitate hydratase (1.1), carnitine acetyltransferase (0.4), acetyl-CoA synthetase (1.0), aspartate aminotransferase (1.7), alanine aminotransferase (6.1). The mean DNA content of eight preparations of fat-cells was 109mug/g dry weight of cells. 2. Mitochondria showing respiratory control ratios of 3-6 with pyruvate, about 3 with succinate and P/O ratios of approaching 3 and 2 respectively have been isolated from fat-cells. From studies of rates of oxygen uptake and of swelling in iso-osmotic solutions of ammonium salts, it is concluded that fat-cell mitochondria are permeable to the monocarboxylic acids, pyruvate and acetate; that in the presence of phosphate they are permeable to malate and succinate and to a lesser extent oxaloacetate but not fumarate; and that in the presence of both malate and phosphate they are permeable to citrate, isocitrate and 2-oxoglutarate. In addition, isolated fat-cell mitochondria have been found to oxidize acetyl l-carnitine and, slowly, l-glycerol 3-phosphate. 3. It is concluded that the major means of transport of acetyl units into the cytoplasm for fatty acid synthesis is as citrate. Extensive transport as glutamate, 2-oxoglutarate and isocitrate, as acetate and as acetyl l-carnitine appears to be ruled out by the low activities of mitochondrial aconitate hydratase, mitochondrial acetyl-CoA hydrolyase and carnitine acetyltransferase respectively. Pathways whereby oxaloacetate generated in the cytoplasm during fatty acid synthesis by ATP-citrate lyase may be returned to mitochondria for further citrate synthesis are discussed. 4. It is also concluded that fat-cells contain pathways that will allow the excess of reducing power formed in the cytoplasm when adipose tissue is incubated in glucose and insulin to be transferred to mitochondria as l-glycerol 3-phosphate or malate. When adipose tissue is incubated in pyruvate alone, reducing power for fatty acid, l-glycerol 3-phosphate and lactate formation may be transferred to the cytoplasm as citrate and malate.
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PMID:The intracellular localization of enzymes in white-adipose-tissue fat-cells and permeability properties of fat-cell mitochondria. Transfer of acetyl units and reducing power between mitochondria and cytoplasm. 439 82

Treatment of rat liver mitochondria with digitonin followed by differential centrifugation was used to resolve the intramitochondrial localization of both soluble and particulate enzymes. Rat liver mitochondria were separated into three fractions: inner membrane plus matrix, outer membrane, and a soluble fraction containing enzymes localized between the membranes plus some solublized outer membrane. Monoamine oxidase, kynurenine hydroxylase, and rotenone-insensitive NADH-cytochrome c reductase were found primarily in the outer membrane fraction. Succinate-cytochrome c reductase, succinate dehydrogenase, cytochrome oxidase, beta-hydroxybutyrate dehydrogenase, alpha-ketoglutarate dehydrogenase, lipoamide dehydrogenase, NAD- and NADH-isocitrate dehydrogenase, glutamate dehydrogenase, aspartate aminotransferase, and ornithine transcarbamoylase were found in the inner membrane-matrix fraction. Nucleoside diphosphokinase was found in both the outer membrane and soluble fractions; this suggests a dual localization. Adenylate kinase was found entirely in the soluble fraction and was released at a lower digitonin concentration than was the outer membrane; this suggests that this enzyme is localized between the two membranes. The inner membrane-matrix fraction was separated into inner membrane and matrix by treatment with the nonionic detergent Lubrol, and this separation was used as a basis for calculating the relative protein content of the mitochondrial components. The inner membrane-matrix fraction retained a high degree of morphological and biochemical integrity and exhibited a high respiratory rate and respiratory control when assayed in a sucrose-mannitol medium containing EDTA.
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PMID:Enzymatic properties of the inner and outer membranes of rat liver mitochondria. 569 70

The mechanism of ammonia assimilation in Methanosarcina barkeri and Methanobacterium thermoautotrophicum was documented by analysis of enzyme activities, 13NH3 incorporation studies, and comparison of growth and enzyme activity levels in continuous culture. Glutamate accounted for 65 and 52% of the total amino acids in the soluble pools of M. barkeri and M. thermoautotrophicum. Both organisms contained significant activities of glutamine synthetase, glutamate synthase, glutamate oxaloacetate transaminase, and glutamate pyruvate transaminase. Hydrogen-reduced deazaflavin-factor 420 or flavin mononucleotide but not NAD, NADP, or ferredoxin was used as the electron donor for glutamate synthase in M. barkeri. Glutamate dehydrogenase activity was not detected in either organism, but alanine dehydrogenase activity was present in M. thermoautotrophicum. The in vivo activity of the glutamine synthetase was verified in M. thermoautotrophicum by analysis of 13NH3 incorporation into glutamine, glutamate, and alanine. Alanine dehydrogenase and glutamine synthetase activity varied in response to [NH4+] when M. thermoautotrophicum was cultured in a chemostat with cysteine as the sulfur source. Alanine dehydrogenase activity and growth yield (grams of cells/mole of methane) were highest when the organism was cultured with excess ammonia, whereas growth yield was lower and glutamine synthetase was maximal when ammonia was limiting.
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PMID:Ammonia assimilation and synthesis of alanine, aspartate, and glutamate in Methanosarcina barkeri and Methanobacterium thermoautotrophicum. 612 78

Pregnant F-344 rats were exposed by intubation to a single dose (35 mg/kg) of 1,2-dimethylhydrazine dihydrochloride or to an acetate buffer on day 14 of gestation. A detailed examination of the effects of this dose of 1,2-dimethylhydrazine on brush border enzymes in the offspring was performed. Changes in the liver and colon levels of the alpha-glycerophosphate and malate/aspartate substrate cycle enzymes were measured during the development; at days 17 and 20 of gestation and at 2, 6, 13, 20, 27, 55, 110 days and 1 year after birth. It is concluded that metabolic energy enzymes, glutamate oxaloacetate transaminase and malate dehydrogenase, are more sensitive to 1,2-dimethylhydrazine treatment than are the brush border hydrolases or NAD- and Fp-linked alpha-glycerophosphate dehydrogenases.
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PMID:Development of rat liver and colon substrate cycle enzymes in offspring with and without prenatal exposure to a carcinogenic dose of 1,2-dimethylhydrazine. 614 Jan 24

The activity of certain key enzymes involved in glutamic acid metabolism was studied in purified brain mitochondria and in mitochondrial subfractions separated in a discontinuous 1.2--1.6 mol/l sucrose gradient. Alanine aminotransferase and glutamate dehydrogenase were found to be matrix enzymes and aspartate aminotransferase to be associated with the inner mitochondrial membranes. After the purified mitochondria had been separated into 5 subfractions, aspartate aminotransferase and NAD+-dependent isocitrate dehydrogenase were found to be bound to the lighter mitochondrial subfractions settling at the 1.4--1.5 mol/l sucrose boundary while alanine aminotransferase, 4-aminobutyrate transaminase and glutamate dehydrogenase were associated with the heavier subfractions settling below 2.4 mol/l sucrose. The highest specific activity of the given enzymes was found in the subfraction settling at the 1.4--1.5 mol/l sucrose boundary, the only exception being alanine aminotransferase activity, whose maximum was found in the subfractions settling in 1.5 and 1.6 mol/l sucrose. It was concluded that alanine aminotransferase, in conjunction with glutamate dehydrogenase, is linked to NH3 binding and to the oxidation of reduced adenine nucleotides; in addition, alanine aminotransferase is presumed to have the function of transporting glutamate from the mitochondria to the extramitochondrial space.
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PMID:Alanine aminotransferase and some other enzymes in different populations of free brain cortex mitochondria. 645 52


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