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Query: EC:3.5.1.1 (asparaginase)
 2,695 document(s) hit in 31,850,051 MEDLINE articles (0.00 seconds)

Yeast strains sigma1278b and Harden and Young, which synthesize only an internal constitutive form of L-asparaginase, do not grow on D-asparagine, as a sole source of nitrogen, and whole cell suspensions of these strains do not hydrolyze D-asparagine. Strains X2180-A2 and D273-10B, which possess an externally active form of asparaginase, are able to grow slowly on D-asparagine, and nitrogen-starved suspensions of these strains exhibit high activity toward the D-isomer. Nitrogen starvation of strain X218O-A2 results in coordinate increase of D- and L-asparaginase activity; the specific activity observed for the D-isomer is approximately 20% greater than that observed for the L-isomer. It was observed, in studies with cell extracts, that hydrolysis of D-asparagine occurred only with extracts from nitrogen-starved cells of strains that synthesize the external form of asparaginase. Furthermore, the activity of the extracts toward the D-isomer was always higher than that observed with the L-isomer. A 400-fold purified preparation of external asparaginase from Saccharomyces cerevisiae X218U-A2 hydrolyzed D-asparagine with an apparent Km of 0.23 mM and a Vmax of 38.7 mumol/min per mg of protein. D-Asparagine was a competitive inhibitor of L-asparagine hydrolysis and the Ki determined for this inhibition was approximately equal to its Km. These data suggest that D-asparagine is a good substrate for the external yeast asparaginase but is a poor substrate for the internal enzyme.
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PMID:Utilization of D-asparagine by Saccharomyces cerevisiae. 76 32

Acinetobacter glutaminase-asparaginase (AGA) and Escherichia coli asparaginase were compared for their effects on plasma and tissue levels of amino acids, ammonia, and glutamyl transferase activity in the mouse. Free asparagine was depleted similarly in plasma and tissues by both enzymes. AGA treatment produced partial depletion of glutamine concentrations in muscle, spleen, small intestine, and liver. Brain and kidney glutamine concentrations actually rose with treatment. Despite over 100-fold increase in plasma glutamate, only the kidney showed a substantial increase in free glutamate levels during AGA treatment. Glutamine biosynthesis measured by glutamyl transferase activity showed an appreciable increase only in the kidney. Ammonia levels in tissues and plasma rose 1.3- to 4.3-fold. In general, E. coli asparaginase treatment had much less effect on these measurements than did AGA. The changes in these levels are discussed in relation to sites of possible toxicity and antitumor effects.
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PMID:Effect of Acinetobacter glutaminase-asparaginase treatment on free amino acids in mouse tissues. 109 50

Acylation of L-asparaginase (L-asparagine amidohydrolase, EC 3.5.1.1) with complete retention of catalytic activity was achieved. Several parameters of the acylation method, based on the binding of palmitoyl residues to epsilon-NH2 groups of protein, were optimized. The correlation between the acylation degree of L-asparaginase and the retention of catalytic activity was established. For a palmitoyl chloride/protein molar ratio ranging from 50 to 900, a degree of modification of 10 to 30% and a retention of catalytic activity of 98 to 60% respectively, was observed. Hydrophobicity of 30% acylated protein was correlated with turbidity in water and octanol and was compared with the native protein. Acylated protein incorporated into liposomes, showed an increase in catalytic activity in intact form as compared to the native enzyme. By the introduction of a sequential acylation cycle, an improvement of the degree of modification with a maximal value at 50% was obtained. Total retention of catalytic activity was achieved by acylation in the presence of 8 mM L-asparagine in a reactional medium.
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PMID:Acylation of L-asparaginase with total retention of enzymatic activity. 212 8

In studies on kinetics of thermoinactivation of glutaminase (asparaginase) from Ps. arantiaca BKMB-548 at 50 degrees and pH 7.0 in presence or in absence of L-glutamate the enzyme inactivation was found to obey the first order equation. Both the glutaminase and asparaginase activities decreased at a similar rate. L-Glutamate stabilized the enzyme due to direct interaction with its molecule. Stability of the complex formed was evaluated quantitatively. L-Glutamate reacted apparently with a specific site on the surface of the enzyme molecule; Kdiss was 0.42 +/- 0.03 mM at pH 7.0 and 50 degrees. No cooperative effect was found. L-Aspartate protected the enzyme completely; stabilizing effects of L-cysteine, L-serine and glycine were similar to the effect of L-glutamate (94%, 84%, 83% and 82%, respectively). At the same time, glutarate, succinate, alpha-ketobutyrate, alpha-ketoglutarate, gamma-aminobutyrate and N-benzoyl glutamate did not exhibit the stabilization effect. The data obtained suggest that the high stabilizing effect might exhibit only the substances containing simultaneously free alpha-NH2 and alpha-COOH groups in a molecule, whereas presence of COOH groups at beta--or gamma-carbon atoms was not essential for the stabilizing effect.
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PMID:[Thermostabilization of glutamin(asparagin)ase from Pseudomonas aurantica BKMB-548]. 402 28

Amino acid utilization was evaluated in seven children with acute lymphocytic leukemia treated with succinylated Acinetobacter glutaminase-asparaginase. All patients received food p.o. ad libitum and glucose-electrolyte solutions i.v.; four patients received an i.v. amino acid supplement (1.5 g/kg/day). Although all patients were in negative energy balance, there was a significant linear regression between nitrogen balance and nitrogen intake during Days 1 to 7 and Days 8 to 14 of the study. The slope of the regression line, reflecting exogenous nitrogen utilization, was not significantly different from that found in healthy young men ingesting adequate or subadequate energy intakes. The Y-intercept (-210 mg/kg/day) indicated an obligatory nitrogen loss that was much greater than normal. Most of the nitrogen loss was due to urinary excretion. Ammonia and urea accounted for 77 to 91% of the urine nitrogen. Urinary glutamate accounted for 4 to 10% of this loss. Urine protein excretion was abnormally high in each of the patients, ranging from 987 to 3440 mg/day. Urine excretion of N-acetyl-beta-glucosaminidase and beta 2-microglobulin was also abnormally high, despite normal blood urea nitrogen and serum creatinine, suggesting that these children had renal tubular dysfunction. The antileukemic effect of succinylated Acinetobacter glutaminase-asparaginase did not appear to be altered by amino acid supplementation. These data indicate that amino acid supplementation can improve nutritional status in patients treated with succinylated Acinetobacter glutaminase-asparaginase.
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PMID:Amino acid utilization and urine protein excretion in children treated with succinylated Acinetobacter glutaminase-asparaginase. 701 7

The activity profile of the periplasmic asparaginase of Saccharomyces cerevisiae was determined during cell growth in an ure2 mutant; in an ure2 transformed with a plasmid containing the gene URE2 and, for comparison, in the strain D273-10B. Cells were cultivated in media presenting variable quantitative and qualitative nitrogen availability and the enzyme activity was evaluated in fresh and in nitrogen-starved cells. Nitrogen affected the asparaginase II level in fresh and starved cells of all strains. In the best condition, enzyme was produced by the wild-type cells at the late log-phase in the glucose/ammonium medium with a carbon to nitrogen ratio 4.3:1. Upon starvation, the activity doubled. The overall profile of the transformed strain was similar to that of the wild-type strain. In the ure2 mutant, high-enzyme levels were observed during growth, as expected. However the activity level, upon starvation, in proline grown cells, increased sixfold, suggesting that in addition to the Ure2p-Gln3p system, another system regulates asparaginase II biosynthesis.
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PMID:L-asparaginase II of saccharomyces cerevisiae. Activity profile during growth using an ure2 mutant P40-3C and a P40-3C + URE2p strain. 1039 75

The carbon and nitrogen sources most suitable for L-asparaginase production by Enterobacter aerogenes were selected and their concentrations optimized in shake-flask cultures. Sodium citrate (1.0%) and diammonium hydrogen phosphate (0.16%) proved to be the best sources of carbon and nitrogen, respectively. Nitrogen catabolite repression of enzyme formation was absent in this bacterium. Cultivation in a reactor showed that the dissolved oxygen level is the limiting factor for L-asparaginase production by E. aerogenes. Glucose was found to be a repressor of enzyme synthesis. Asparagine was absent intracellularly when the L-asparaginase level was high. An increase in the extracellular alanine level when the dissolved oxygen remained low indicated a shift from aerobic to fermentative metabolism.
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PMID:Studies on nutritional and oxygen requirements for production of L-asparaginase by Enterobacter aerogenes. 1070 80

Bacterial L-asparaginases, enzymes that catalyze the hydrolysis of L-asparagine to aspartic acid, have been used for over 30 years as therapeutic agents in the treatment of acute childhood lymphoblastic leukemia. Other substrates of asparaginases include L-glutamine, D-asparagine, and succinic acid monoamide. In this report, we present high-resolution crystal structures of the complexes of Erwinia chrysanthemi L-asparaginase (ErA) with the products of such reactions that also can serve as substrates, namely L-glutamic acid (L-Glu), D-aspartic acid (D-Asp), and succinic acid (Suc). Comparison of the four independent active sites within each complex indicates unique and specific binding of the ligand molecules; the mode of binding is also similar between complexes. The lack of the alpha-NH3(+) group in Suc, compared to L-Asp, does not affect the binding mode. The side chain of L-Glu, larger than that of L-Asp, causes several structural distortions in the ErA active side. The active site flexible loop (residues 15-33) does not exhibit stable conformation, resulting in suboptimal orientation of the nucleophile, Thr15. Additionally, the delta-COO(-) plane of L-Glu is approximately perpendicular to the plane of gamma-COO(-) in L-Asp bound to the asparaginase active site. Binding of D-Asp to the ErA active site is very distinctive compared to the other ligands, suggesting that the low activity of ErA against D-Asp could be mainly attributed to the low k(cat) value. A comparison of the amino acid sequence and the crystal structure of ErA with those of other bacterial L-asparaginases shows that the presence of two active-site residues, Glu63(ErA) and Ser254(ErA), may correlate with significant glutaminase activity, while their substitution by Gln and Asn, respectively, may lead to minimal L-glutaminase activity.
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PMID:Structural basis for the activity and substrate specificity of Erwinia chrysanthemi L-asparaginase. 1134 30

The localization of activity of immobilzed L-asparaginase by covalent binding was studied by X-ray microanalysis. Asparagine and MgCl2 served as substrate and capture agent respectively. Substrate was catalysed by immobilized L-asparaginase to produce NH3, and NH3 was captured by MgCl2 to form precipitate MgNH4PO4. Precipitae was deposited on active site of immobilized L-asparaginase. The results show that the macroporous resins of immobilized L-asparaginase has greater enzyme activity, while distribution of activated enzyme was uniform. Most of activated enzyme was immobilized on the macroporous resins. The optimum condition of localization of activity of immobilized L-asparaginase was studied.
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PMID:[X-ray microanalysis of the activity of immobilized L-asparaginase]. 1576 44

The fate of nitrogen originating from the amide group of asparagine in young pea leaves (Pisum sativum) has been studied by supplying [(15)N-amide]asparagine and its metabolic product, 2-hydroxysuccinamate (HSA) via the transpiration stream. Amide nitrogen from asparagine accumulated predominantly in the amide group of glutamine and HSA, and to a lesser extent in glutamate and a range of other amino acids. Treatment with 5-diazo,4-oxo-L-norvaline (DONV) a deamidase inhibitor, caused a decrease in transfer of label to glutamine-amide. Virtually no (15)N was detected in HSA of leaves supplied with asparagine and the transaminase inhibitor aminooxyacetate. When [(15)N]HSA was supplied to pea leaves, most of the label was also found in the amide group of glutamine and this transfer was blocked by the addition of methionine sulfoximine, which caused a large increase in NH(3) accumulation. DONV was not specific for asparaginase, and inhibited the deamidation of HSA, causing a decrease in transfer of (15)N into glutamine-amide, NH(3), and other amino acids. It is concluded from these results that use of the amide group of asparagine as a nitrogen source for young pea leaves involves deamidation of both asparagine and its transamination product HSA (possibly also oxosuccinamate). The amide group, released as ammonia, is then reassimilated via the glutamine synthetase/glutamate synthase system.
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PMID:Utilization of the amide groups of asparagine and 2-hydroxysuccinamic Acid by young pea leaves. 1666 59


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