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1.
The activity and induction patterns of the isoenzymes of aspartate aminotransferase (AsAT) of the liver of male rats during growth, development, and senescence were studied. The activity of both the cytoplasmic and mitochondrial isoenzymes increased significantly until adulthood and remained constant thereafter. Adrenalectomy decreased, and hydrocortisone administration increased, the activity of the cytoplasmic AsAT of the liver of young-immature, adult, and senescent rats. However, the degree of these responses decreased with increasing age of the animal. The hormone-mediated induction of this isoenzyme is actinomycin D-sensitive. Mitochondrial AsAT on the other hand is irresponsive towards these treatments throughout the life span of the rat. 相似文献
2.
Intracellular distribution of aspartate aminotransferase (AAT) in Spirodela polyrhiza (Lemnaceae), strain SJ, has been studied by differential centrifugation. The bulk of the enzyme (73% of total cellular content) was localized in the cytoplasm and 24% activity was localized in chloroplasts. Purified cytoplasmic and chloroplastic isozymes differed by their affinity for substrates. The reaction balance was shifted towards direct and reverse transamination in the cytoplasm and chloroplast, respectively. Competitive inhibition of AAT by excessive substrates and enzyme affinity modulation by certain intermediates of the tricarboxylic acid cycle (isocitrate, succinate, and citrate) were observed. Possible involvement of AAT isozymes in the coordination of carbon and nitrogen metabolism through the regulation of 2-oxoglutarate synthesis and utilization in different cellular compartments is discussed. 相似文献
3.
E C Cornish C M Cussen F J Hird P E Todd 《Comparative biochemistry and physiology. B, Comparative biochemistry》1978,61(3):375-378
1. The activities of aminotransferases catalysing the transfer of amino groups from aspartate, alanine and leucine to 2-oxoglutarate in different tissues of the rat, pigeon and trout have been determined. 2. Alanine-2-oxoglutarate aminotransferase was high in the liver of the rat and trout and low in that of the pigeon. 3. Aspartate-2-oxoglutarate aminotransferase was usually the dominant aminotransferase in all tissues and was highest in oxidative tissues where the TCA cycle is active. Its activity in the various livers is not correlated with the function of aspartate in nitrogen excretion. 4. The activity of aspartate-2-oxoglutarate aminotransferase in oxidative tissues argues that aspartate in conjunction with this enzyme serves as a buffer of oxaloacetate to keep the TCA cycle running and/or to mediate the transfer of reducing equivalents across mitochondrial membranes. 相似文献
4.
The activity of lactate dehydrogenase (LDH), indophenol oxidase, aspartate aminotransferase (AsAT), alkaline phosphatase, acid phosphatase and aldolase at different stages of rat development was measured. We have also determined changes in the activity of these enzymes resulting from transplantation of embryonic nerve tissue (ENT) into the brain of adult animals. During development from the embryo to the adult animal, LDH and AsAT activities increased, while alkaline phosphatase activity diminished. After ENT transplantation, the most prominent changes were in the alkaline phosphatase activity whereas the activity of LDH, AsAT and acid phosphatase remained unchanged and similar to that in the brain cortex of intact adult animals. Changes in the enzyme activity resulting from ENT transplantation changed in a manner characteristic of the transplant. Local brain damage did not change the activity of the studied enzymes fifty days after surgery. 相似文献
5.
In pea seedlings transferred to a medium containing 3% NaCl, NADP-dependent isocitrate dehydrogenase (NADP-IDH) activity in leaves increased more than twofold during the first hour, decreased, and increased again (approximately 1.5 times) by the 14th hour of exposure. Some catalytic and regulatory properties of NADP-IDH were studied using enzyme preparations purified from normal plants and plants exposed to salt stress. The results showed that K
m for NADP-IDH in reactions with isocitrate and NADP decreased under stress by factors of 2 and 3, respectively. Specific features of enzyme activity regulation by citrate and trans-aconitate under these conditions were revealed. The inhibitory effects of -aminobutyric acid (GABA) and 2-oxoglutarate on NADP-IDH from plants exposed to salt stress was stronger than that on the enzyme from normal plants. Glutamine and glutamate slightly activated the enzyme in both cases. 相似文献
6.
Carbohydrate deprivation reduces NADPH-production in fish liver but not in adipose tissue 总被引:3,自引:0,他引:3
Barroso JB Peragón J García-Salguero L de la Higuera M Lupiáñez JA 《The international journal of biochemistry & cell biology》2001,33(8):785-796
Little is known about the way in which carnivorous fish such as salmonids mobilise and metabolise dietary carbohydrates, which are essential to lipid metabolism. Thus we have studied changes caused by the absence of dietary carbohydrates to the kinetics and molecular behaviour of the four cellular NADPH-production systems [glucose 6-phosphate dehydrogenase (G6PDH); 6-phosphogluconate dehydrogenase (6PGDH); malic enzyme (ME); and isocitrate dehydrogenase NADP-dependent (NADP-IDH)] in the liver and adipose tissue of rainbow trout (Oncorhynchus mykiss). We used spectrophotometry to study enzyme kinetics and nucleic acid concentrations, and immunoblot analysis to determine specific protein concentrations. The absence of carbohydrate reduced specific enzyme activity, maximum rate and catalytic efficiency by almost 65% in G6PDH and 6PGDH, by more than 50% in ME, and by almost 25% in NADP-IDH but caused no significant changes in the K(m) values or activity ratios in any of these hepatic enzymes. Molecular analysis clearly showed that this kinetic behaviour reflected concomitant changes in intracellular enzyme concentrations, produced by protein-induction/repression processes rather than changes in the activity of pre-existing enzymes. We conclude that the absence of carbohydrates significantly reduces intracellular concentrations of G6PDH, ME and NADP-IDH in trout liver in percentages similar to those recorded for enzyme activity. We found no such variations in the concentrations of any of these enzymes in adipose tissue and no change in the levels of their activity, suggesting that the liver and adipose tissues are subject to different regulation systems with regard to carbohydrates and play distinct roles in lipid metabolism. 相似文献
7.
Abstract— Mitochondrial and cytoplasmic forms of aspartate aminotransferase were purified from rat brain homogenates and tested for their ability to catalyze transamination of various aromatic amino acids. The mitochondrial enzyme exhibited activity toward tyrosine and phenylalanine with 2-oxoglutar-ate as acceptor, although the specific activities were less than 1% of the corresponding aspartate activity when all substrates were 10 mM. Even less activity was seen with DOPA, 5-hydroxytryptophan and tryptophan. The cytoplasmic aspartate aminotransferase was active toward tryptophan, 5-hydroxytryptophan and DOPA, but these transaminations were favored by pyruvate or oxaloacetate rather than 2-oxoglutarate as keto acid. Based on co-migration of aromatic activities with the respective aspartate aminotransferases during isoelectric focusing and based on equal sensitivities of aromatic transamination and aspartate transamination to inhibition by vinylglycine, it was concluded that all activities resided in the aspartate aminotransferase enzymes. Some doubt exists, however, as to the physiological significance of these alternate activities in view of the requirement that aromatic amino acids must compete with aspartate for transamination by these enzymes. 相似文献
8.
Ethanol or acetaldehyde orally administered (15% and 2% respectively in drinking water) to male Wistar rats for three months induced alterations in the main liver enzymes responsible for ethanol metabolism, aspartate and alanine aminotransferases and NAD glutamate dehydrogenase. Ethanol produced a significant decrease in the activity of soluble alcohol dehydrogenase, while acetaldehyde induced alterations both in soluble and mitochondrial aldehyde dehydrogenases: soluble activity was significantly higher than in the control and ethanol-treated groups, and mitochondrial activity was significantly diminished. Both soluble aspartate and alanine aminotransferases showed pronounced increases by the chronic effect of acetaldehyde, while mitochondrial activities were practically unchanged by the effect of ethanol or acetaldehyde. Mitochondrial NAD glutamate dehydrogenase showed a rise in its activity both by the effect of chronic ethanol and acetaldehyde consumption. The level of metabolites assayed in liver extracts showed marked differences between ethanol and acetaldehyde treatment which indicates that ethanol produced a remarkable increase in glutamate, aspartate and free ammonia together with marked decrease in pyruvate and 2-oxoglutarate concentrations. Acetaldehyde consumption induced a significant decrease in 2-oxoglutarate and pyruvate concentrations. These observations suggest that ethanol has an important effect on the urea cycle enzymes, while the effect of acetaldehyde contributes to the impairment of the citric acid cycle. 相似文献
9.
Some kinetic and other properties of the isoenzymes of aspartate aminotransferase isolated from sheep liver.
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A method for the purification of mitochondrial isoenzyme of sheep liver aspartate aminotransferase (EC 2.6.1.1) is described. The final preparation is homogeneous by ultracentrifuge analyses and polyacrylamide-gel electrophoresis and has a high specific activity (182 units/mg). The molecular weight determined by sedimentation equilibrium is 87,100 +/- 680. The amino acid composition is presented; it is similar to that of other mitochondrial isoenzymes, but with a higher content of tyrosine and threonine. Subforms have been detected. On isoelectric focusing a broad band was obtained, with pI 9.14. The properties of the mitochondrial aspartate aminotransferase are compared with those of the cytoplasmic isoenzyme. The Km for L-aspartate and 2-oxoglutarate for the cytoplasmic enzyme were 2.96 +/- 0.20 mM and 0.093 +/- 0.010 mM respectively; the corresponding values for the mitochondrial form were 0.40 +/- 0.12 mM and 0.98 +/- 0.14 mM. Cytoplasmic aspartate aminotransferase showed substrate inhibition by concentrations of 2-oxoglutarate above 0.25 mM in the presence of aspartate up to 2mM. The mitochondrial isoenzyme was not inhibited in this way. Pi at pH 7.4 inhibited cytoplasmic holoenzyme activity by up to about 60% and mitochondrial holoenzyme activity up to 40%. The apparent dissociation constants for pyridoxal 5'-phosphate were 0.23 micrometer (cytoplasmic) and 0.062 micrometer (mitochondrial) and for pyridoxamine 5'-phosphate they were 70 micrometer (cytoplasmic) and 40 micrometer (mitochondrial). Pi competitively inhibited coenzyme binding to the apoenzymes; the inhibition constants at 37 degree C were 32 micrometer for the cytoplasmic isoenzyme and 19.5 micrometer for the mitochondrial form. 相似文献
10.
Medvedeva LV Popova TN Artyukhov VG Matasova LV Pinheiro de Carvalho MA 《Biochemistry. Biokhimii?a》2002,67(6):696-705
The intensity of free radical processes and the regulation of NADP-isocitrate dehydrogenase (EC 1.1.1.42; NADP-IDH) activity have been studied in the cytoplasmic fraction of normal and ischemized rat myocardium. Chemiluminescence parameters, such as the light sum (S) of slow flash and the tangent of the kinetic curve slope angle (tan1), which characterize the intensity of free radical processes, were increased in ischemia 2.1- and 20.0-fold, respectively. The slow flash intensity (Imax) was increased 22-fold. The contents of lipid peroxidation products–diene conjugates and malonic dialdehyde–were increased 11.9- and 4.7-fold, respectively, suggesting pronounced oxidative stress. Using homogenous enzyme preparations of NADP-IDH isolated from the normal and experimentally ischemized rat myocardium, a number of catalytic properties of the enzyme were characterized for normal and pathologic conditions. NADP-IDH from the normal and ischemized myocardium had the same electrophoretic mobility and was regulated similarly by Fe2+, Cu2+, Zn2+, and also with succinate and fumarate. However, under normal and pathologic conditions NADP-IDH was different in the affinity for substrates and in the sensitivity to inhibitory effects of hydrogen peroxide, reduced glutathione, and of Ca2+. The degree of synergy in the enzyme inhibition with Fe2+ and H2O2 was less pronounced in ischemia. The inhibitory effect of the reaction product 2-oxoglutarate was higher under normal conditions than in ischemia (the K
i values were 0.22 and 0.75 mM, respectively). The specific features of the NADP-IDH regulation in ischemia are suggested to promote the stimulation of the enzyme functioning during increased level of free radical processes, and this seems to be important for NADPH supplying for the glutathione reductase/glutathione peroxidase antioxidant system of cardiomyocytes. 相似文献
11.
Incubation of 50 mM D-glucose with aspartate aminotransferase (AST, EC 2.6.1.1) preparations (purified pig heart enzyme or a rat liver 20,000 x g supernatant) at 25 degrees C had no effect on enzyme activity. 50 mM D-fructose or D-ribose gradually inhibited pig heart AST under the same conditions to zero activity after 14 days. 50 mM DL-glyceraldehyde decreased enzyme activity to zero after 6 days of incubation. The inhibition of pig heart AST by 50 mM D-fructose or D-ribose was marked even at a temperature of 4 degrees C but it was less pronounced than at 25 degrees C. There was no effect of 0.5 mM 2-oxoglutarate on AST activity during incubation, while the presence of 25 mM L-aspartate decreased it rapidly. 0.5 mM 2-oxoglutarate partly prevented inhibition of AST by D-ribose or D-fructose, while an analogous experiment with 25 mM aspartate resulted in a rapid decline similar to that in the absence of sugars. 相似文献
12.
Jaroslav Dršata Martin Beránek VladimÍr Palička 《Journal of enzyme inhibition and medicinal chemistry》2013,28(1):31-36
Incubation of 50 mM d -glucose with aspartate aminotransferase (AST, EC 2.6.1.1) preparations (purified pig heart enzyme or a rat liver 20,000 × g supernatant) at 25°C had no effect on enzyme activity. 50 mM d -fructose or d -ribose gradually inhibited pig heart AST under the same conditions to zero activity after 14 days. 50 mM dl -glyceraldehyde decreased enzyme activity to zero after 6 days of incubation. The inhibition of pig heart AST by 50 mM d -fructose or d -ribose was marked even at a temperature of 4°C but it was less pronounced than at 25°C. There was no effect of 0.5 mM 2-oxoglutarate on AST activity during incubation, while the presence of 25 mM l -aspartate decreased it rapidly. 0.5 mM 2-oxoglutarate partly prevented inhibition of AST by d -ribose or d -fructose, while an analogous experiment with 25 mM aspartate resulted in a rapid decline similar to that in the absence of sugars. 相似文献
13.
The influence of maternal energy intake on the development of gluconeogenesis was studied in the liver of the bovine fetus from Days 88 to 270 of gestation. Fetal liver activities (units per gram of tissue) of cytoplasmic GTP:oxalacetate carboxy-lyase (transphosphorylating) (PEPCK) and mitochondrial l-malate:NAD+ oxidoreductase (MDH) increased linearly with increasing gestational age. Fetal cytoplasmic MDH activities reached maternal levels by 120 days of gestation, and fetal mitochondrial pyruvate carboxylase approached maternal levels by 200 days of gestation. Fetal activities of mitochondrial and cytoplasmic propionyl-CoA:carbondioxide ligase (ADP-forming) (PCC) did not change with gestational age and were about 45 and 7%, respectively, of maternal levels. Fetal activities of mitochondrial and cytoplasmic l-aspartate: 2-oxoglutarate aminotransferase were both about 24% of the maternal activities throughout gestation. Maternal and fetal liver activities of d-fructose-1,6-diphosphate 1-phosphohydrolase (FDP) were similar and did not change with gestational age. Glucose synthesis from lactate by fetal liver slices in vitro was slightly lower and, from alanine and aspartate, was slightly higher than glucose synthesis by maternal liver slices. Restriction of maternal dietary energy intake did not significantly alter gluconeogenic-related enzyme activity in vitro in maternal or fetal liver or in the metabolism of aspartate, alanine, or lactate to glucose or CO2 by liver slices in vitro. A capacity for gluconeogenesis has been measured in the bovine fetus as early as 88 days of gestation. 相似文献
14.
Hajime Ohigashi Mitsuru Hirota Takanao Ohtsuka Koichi Koshimizu Hirota Fujiki Masami Suganuma 《Bioscience, biotechnology, and biochemistry》2013,77(10):2605-2608
Phosphoenolpyruvate (PEP) carboxylase purified from Brevibacterium flavum was specifically activated by fructose 1,6-bisphosphate (FBP). The other intermediates of sugar metabolism or their structural analogues did not influence the activity. FBP decreased the apparent Km for PEP but did not affect that for another substrate, bicarbonate, or the apparent maximum velocity for PEP. The dissociation constants for FBP from enzyme-FBP and enzyme-PEP-FBP complex were 63 and 32 μm, respectively, being almost equivalent to those for acetyl-CoA. Synergistic activation by FBP and acetyl-CoA was not observed with the B. flavum enzyme, unlike the Escherichia coli enzyme. FBP, like acetyl-CoA, was kinetically competitive with aspartate. With respect to another feedback inhibitor, 2-oxoglutarate, acetyl-CoA was non-competitive, whereas FBP was of mixed-type, i.e., FBP but not acetyl-CoA prevented 2-oxoglutarate from binding to the enzyme to a certain extent. Homotropic cooperativity was observed only with FBP but not with acetyl-CoA in the absence of inhibitors. Cooperativities of FBP and acetyl-CoA were increased by aspartate but not by 2-oxoglutarate. In the aspartate-overproducing mutant enzyme, the Michaelis constant for PEP was decreased, whereas the inhibitor constant for aspartate with or without simultaneous addition of 2-oxoglutarate and the activator constants for FBP and acetyl-CoA were increased. The decreased Michaelis constant for PEP was comparable to the apparent Km of the wild-type enzyme for PEP in the presence of the saturated concentration of FBP, and would result in a further decrease in the affinity of the mutant enzyme for aspartate. 相似文献
15.
A newly discovered enzyme in mammalian tissues, aspartate-4-decarboxylase (EC 4.1.1.12), catalyzes the exothermic conversion of aspartate to alanine and CO2. The occurrence of this enzyme poses at least two important questions. First, what is the purpose of such an enzyme in cell physiology? There are alternate ways to convert aspartate to alanine which are rapid and which conserve energy. Second, since the synthesis of aspartate is an energy-requiring process, how can the cell limit undue energy drain by this, seemingly pointless, beta-decarboxylation of aspartate? It is demonstrated that rat liver aspartate-4-decarboxylase is inhibited by acetyl-coenzyme A and stimulated by glutamate. These regulatory properties were predicted a priori. It was suggested that, in coordination with pyruvate carboxylase, aspartate-4-decarboxylase is important in regulating the metabolic fate of oxaloacetate and thus plays a role in determining the efficiency of carbohydrate metabolism. Furthermore, reciprocal regulation of rat liver pyruvate carboxylase and aspartate-4-decarboxylase would assure a limit on the extent of futile cycling that may occur between these enzymes. 相似文献
16.
Tryptophan aminotransferase was purified from rat brain extracts. The purified enzyme had an isoelectric point at pH 6.2 and a pH optimum near 8.0. On electrophoresis the enzyme migrated to the anode. The enzyme was active with oxaloacetate or 2-oxoglutarate as amino acceptor but not with pyruvate, and utilized various L-amino acids as amino donors. With 2-oxoglutarate, the order of effectiveness of the L-amino acids was aspartate > 5-hydroxytryptophan > tryptophan > tyrosine > phenylalanine. Aminotransferase activity of the enzyme towards tryptophan was inhibited by L-glutamate. Sucrose density gradient centrifugation gave a molecular weight of approx. 55,000. The enzyme was present in both the cytosol and synaptosomal cytosol, but not in the mitochondria. The isoelectric focusing profile of tryptophan: oxaloacetate aminotransferase activity was identical with that of L-aspartate: 2-oxoglutarate aminotransferase (EC 2.6.1.1) activity, with both subcellular fractions. On the basis of these data, it is suggested that the enzyme is identical with the cytosol aspartate: 2-oxoglutarate aminotransferase. 相似文献
17.
Tsutomu Sugiura Yasuhide Ota Yasuji Minoda 《Bioscience, biotechnology, and biochemistry》2013,77(12):2479-2480
In accordance with the regulation by aspartate of phosphoenolpyrubate (PEP*) carboxylase, glutamate formation in Brevibacterium flavum, a glutamate-producing bacterium, was inhibited by the addition of aspartate. Furthermore, an increase in aspartate formation caused by a mutational decrease in citrate synthase specific activity was accompanied by a decrease in the total amount of glutamate and aspartate formed. However, a mutational decrease in glutamate dehydrogenase activity caused a decrease in the total amount without increasing the asparate formation but with accumulation of 2-oxoglutarate, suggesting that the feedback inhibition by the aspartate of PEP carboxylase was enhanced by 2-oxoglutarate. In fact, partially purified PEP carboxylase from this organism was found to be synergistically inhibited by aspartate and 2-oxoglutarate, citrate, cis-aconitase, or isocitrate. Among them, the effects of tricarboxylic acids were attributed to their non-specific chelating action with Mn2+, an activator of the enzyme. The synergistic action of 2-oxoglutarate was accompanied by a decrease in Hill coefficient for the aspartate of the enzyme. 相似文献
18.
A method for the purification of chicken liver soluble aspartate aminotransferase, lactate dehydrogenase free, is proposed. The preparation, which contained a mixture of the five molecular forms of the enzyme, showed a 120-fold increase in specific activity, with respect to the initial homogenates. Differences in Km(2-oxoglutarate) and saturating concentrations among solutions of purified enzyme and soluble fraction were due to the 2-oxoglutarate reductase activity of lactate dehydrogenase. 相似文献
19.
Selective permeability of rat liver mitochondria to purified aspartate aminotransferases in vitro. 总被引:4,自引:4,他引:0
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1. A method was devised to allow determination of intramitochondrial aspartate amino-transferase activity in suspensions of intact mitochondria. 2. Addition of purified rat liver mitochondrial aspartate aminotransferase to suspensions of rat liver mitochondria caused an apparent increase in the intramitochondrial enzyme activity. No increase was observed when the mitochondria were preincubated with the purified cytoplasmic isoenzyme. 3. These results suggest that mitochondrial aspartate aminotransferase, but not the cytoplasmic isoenzyme, is able to pass from solution into the matrix of intact rat liver mitochondria in vitro. 4. This system may provide a model for studies of the little-understood processes by which cytoplasmically synthesized components are incorporated into mitochondria in vivo. 相似文献
20.
Experimental diabetes causes mitochondrial loss and cytoplasmic enrichment of pyridoxal phosphate and aspartate aminotransferase activity 总被引:1,自引:0,他引:1
The streptozotocin diabetic rat was selected as a model to study how insulin deficiency alters vitamin B6 utilization by focusing on pyridoxal phosphate levels and aspartate aminotransferase activities in liver tissues. Diabetes of 15 weeks' duration lowered plasma pyridoxal phosphate levels by 84%. Normal plasma pyridoxal phosphate was 480 pmole/ml. Fractionation of liver into mitochondrial and extramitochondrial compartments demonstrated that diabetes caused a 43% diminution in mitochondrial pyridoxal phosphate per gram of liver. There was no cytoplasmic change in these diabetic rats. Mitochondrial aspartate aminotransferase activity was decreased 53% per gram of diabetic liver and cytoplasmic aspartate aminotransferase activity was elevated 3.4-fold. Damage to diabetic mitochondria during preparation procedures could not account for the rise in cytoplasmic aspartate aminotransferase activity. Electrophoresis showed that in the diabetic cytoplasm both cathodal and anodal forms of the enzyme were elevated. Speculations concerning mitochondrial loss and cytoplasmic gain of enzyme activity as well as those on the reduction of plasma pyridoxal phosphate in the diabetic rat are presented. 相似文献