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

Pseudomonas putida KT2440 metabolizes a wide range of carbon and nitrogen sources, including many amino acids. In this study, a sigma54-dependent two-component system that controls the uptake and metabolism of acidic amino acids was identified. The system (designated aau, for acidic amino acid utilization) involves a sensor histidine kinase, AauS, encoded by PP1067, and a response regulator, AauR, encoded by PP1066. aauR and aauS deletion mutants were unable to efficiently utilize aspartate (Asp), glutamate (Glu), and glutamine (Gln) as sole sources of carbon and nitrogen. Growth of the mutants was partially restored when the above-mentioned amino acids were supplemented with glucose or succinate as an additional carbon source. Uptake of Gln, Asp, and asparagine (Asn) by the aauR mutant was moderately reduced, while Glu uptake was severely impaired. In the absence of glucose, the aauR mutant even secreted Glu into the medium. Furthermore, disruption of aauR affected the activities of several key enzymes of Glu and Asp metabolism, leading to the intracellular accumulation of Glu and greatly reduced survival times under conditions of nitrogen starvation. By a proteomics approach, four major proteins were identified that are downregulated during growth of the aauR mutant on Glu. Two of these were identified as periplasmic glutaminase/asparaginase and the solute-binding protein of a Glu/Asp transporter. Transcriptional analysis of lacZ fusions containing the putative promoter regions of these genes confirmed that their expression is indeed affected by the aau system. Three further periplasmic solute-binding proteins were strongly expressed during growth of the aauR deletion mutant on Glu but downregulated during cultivation on glucose/NH4+. These systems may be involved in amino acid efflux.
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PMID:The AauR-AauS two-component system regulates uptake and metabolism of acidic amino acids in Pseudomonas putida. 1702 Dec 7

Asparaginase is a key component of the chemotherapy protocols used in the treatment of acute lymphoblastic leukemia (ALL). The current treatment protocols are remarkable in that childhood ALL cure rates are approaching 85%. As the name implies, asparaginase catalyzes the deamination of asparagine to aspartic acid. What is not generally realized is that asparaginase also catalyzes, essentially to the same extent, the removal of the amide nitrogen from glutamine to form glutamic acid. Glutamine is a required substrate for three enzymes involved in the de novo synthesis of purine nucleotides and two enzymes involved in the de novo synthesis of pyrimidine nucleotides. In this review, the specific roles of glutamine in the de novo synthesis of nucleotides are defined and an appropriate explanation for the cell cycle arrest and cytotoxicity induced in proliferating malignant lymphoblasts by asparaginase treatment is provided.
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PMID:Critical roles of glutamine as nitrogen donors in purine and pyrimidine nucleotide synthesis: asparaginase treatment in childhood acute lymphoblastic leukemia. 1709 64

AnsA is the cytoplasmic asparaginase from Escherichia coli involved in intracellular asparagine utilization. Analytical ultracentifugation and X-ray crystallography reveal that AnsA forms a tetrameric structure as a dimer of two intimate dimers. Kinetic analysis of the enzyme reveals that AnsA is positively cooperative, displaying a sigmoidal substrate dependence curve with an [S](0.5) of 1 mM L-asparagine and a Hill coefficient (n(H)) of 2.6. Binding of L-asparagine to an allosteric site was observed in the crystal structure concomitant with a reorganization of the quarternary structure, relative to the apo enzyme. The carboxyl group of the bound asparagine makes salt bridges and hydrogen bonds to Arg240, while the N(delta2) nitrogen interacts with Thr162. Mutation of Arg240 to Ala increases the [S](0.5) value to 5.9 mM, presumably by reducing the affinity of the site for L-asparagine, although the enzyme retains cooperativity. Mutation of Thr162 to Ala results in an active enzyme with no cooperativity. Transmission of the signal from the allosteric site to the active site appears to involve subtle interactions at the dimer-dimer interface and relocation of Gln118 into the vicinity of the active site to position the probable catalytic water molecule. These data define the structural basis for the cooperative regulation of the intracellular asparaginase that is required for proper functioning within the cell.
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PMID:Crystal structure and allosteric regulation of the cytoplasmic Escherichia coli L-asparaginase I. 1745 45

Natural Saccharomyces cerevisiae yeast strains exhibit very large genotypic and phenotypic diversity. However, the link between phenotype variation and genetic determinism is still difficult to identify, especially in wild populations. Using genome hybridization on DNA microarrays, it is now possible to identify single-feature polymorphisms among divergent yeast strains. This tool offers the possibility of applying quantitative genetics to wild yeast strains. In this instance, we studied the genetic basis for variations in acetic acid production using progeny derived from two strains from grape must isolates. The trait was quantified during alcoholic fermentation of the two strains and 108 segregants derived from their crossing. A genetic map of 2212 markers was generated using oligonucleotide microarrays, and a major quantitative trait locus (QTL) was mapped with high significance. Further investigations showed that this QTL was due to a nonsynonymous single-nucleotide polymorphism that targeted the catalytic core of asparaginase type I (ASP1) and abolished its activity. This QTL was only effective when asparagine was used as a major nitrogen source. Our results link nitrogen assimilation and CO(2) production rate to acetic acid production, as well as, on a broader scale, illustrating the specific problem of quantitative genetics when working with nonlaboratory microorganisms.
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PMID:Single QTL mapping and nucleotide-level resolution of a physiologic trait in wine Saccharomyces cerevisiae strains. 1753 82

The field study was conducted to evaluate the effect of municipal solid waste compost (MSWC) as a soil amendment on L-asparaginase (LA) and L-glutaminase (LG) activities. Experiments were conducted during the wet seasons of 1997, 1998 and 1999 on rice grown under a submerged condition, at the Agriculture Experimental Farm, Calcutta University at Baruipur, West Bengal, India. The treatments consisted of control, no input; MSWC, at 60 Kg N ha(- 1); well-decomposed cow manure (DCM), at 60 Kg N ha(- 1); MSWC (30 Kg N ha(- 1)) + Urea (U) (30 Kg N ha(- 1)); DCM (30 Kg N ha(- 1)) + U (30 Kg N ha(- 1)) and Fertilizer, (at 60:30:30 NPK kg ha(- 1)) through urea, single superphosphate and muriate of potash respectively). LA and LG activities alone and their ratio with organic-C (ratio index value, RIV), straw and grain yield were higher in DCM than MSWC-treated soils, due to higher amount of biogenic organic materials like water-soluble organic carbon, carbohydrate and mineralizable nitrogen in the former. The studied parameters were higher when urea was integrated with DCM or MSWC, compared to their single applications. The heavy metals in MSWC did not detrimentally influence the above-measured activities of soil. In the event of long term MSWC application, changes in soil quality parameters should be monitored regularly, since heavy metals once entering into soil persist over a long period.
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PMID:L-asparaginase and L-glutaminase activities in submerged rice soil amended with municipal solid waste compost and decomposed cow manure. 1756 68

Glutamine-free culture of Vero cells has previously been shown to cause higher cell yield and lower ammonia accumulation than that in glutamine-containing culture. Nitrogen metabolism of asparagine and glutamate as glutamine replacer was studied here using nuclear magnetic resonance (NMR) spectroscopy. (15)N-labelled glutamate or asparagine was added and their incorporation into nitrogenous metabolites was monitored by heteronuclear multiple bond coherence (HMBC) NMR spectroscopy. In cells incubated with L: -[(15)N]glutamate, the (15)N label was subsequently found in a number of metabolites including alanine, aspartate, proline, and an unidentified compound. No detectable (15)NH(+)(4) signal occurred, indicating that glutamate was utilized by transamination rather than by oxidative deamination. In cells incubated with L: -[2-(15)N]asparagine, the (15)N label was subsequently found in aspartate, the amine group of glutamate/glutamine, and in two unidentified compounds. Incubation of cells with L: -[4-(15)N]asparagine showed that the amide nitrogen of asparagine was predominantly transferred to glutamine amide. There was no detectable production of (15)NH(+)(4), showing that most of the asparagine amide was transaminated by asparagine synthetase rather than deaminated by asparaginase. Comparing with a glutamine-containing culture, the activities of phosphate-activated glutaminase (PAG), glutamate dehydrogenase (GDH) and alanine aminotransferase (ALT) decreased significantly and the activity of aspartate aminotransferase (AST) decreased slightly.
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PMID:Nitrogen metabolism of asparagine and glutamate in Vero cells studied by (1)H/ (15)N NMR spectroscopy. 1795 33

We describe an ATP-binding cassette (ABC) transporter in Pseudomonas putida KT2440 that mediates the uptake of glutamate and aspartate. The system (AatJMQP, for acidic amino acid transport) is encoded by an operon involving genes PP1071-PP1068. A deletion mutant with inactivated solute-binding protein (KTaatJ) failed to grow on Glu and Gln as sole sources of carbon and nitrogen, while a mutant lacking a functional nucleotide-binding domain (KTaatP) was able to adapt to growth on Glu after an extended lag phase. Uptake of Glu and Asp by either mutant was greatly impaired at both low and high amino acid concentrations. The purified solute-binding protein AatJ exhibited high affinity towards Glu and Asp (K(d)=0.4 and 1.3 muM, respectively), while Gln and Asn as well as dicarboxylates (succinate and fumarate) were bound with much lower affinity. We further show that the expression of AatJMQP is controlled by the sigma(54)-dependent two-component system AauRS. Binding of the response regulator AauR to the aat promoter was examined by gel mobility shift assays and DNase I footprinting. By in silico screening, the AauR-binding motif (the inverted repeat TTCGGNNNNCCGAA) was detected in further P. putida KT2440 genes with established or putative functions in acidic amino acid utilization, and also occurred in other pseudomonads. The products of these AauR-responsive genes include the H(+)/Glu symporter GltP, a periplasmic glutaminase/asparaginase, AnsB, and phosphoenolpyruvate synthase (PpsA), a key enzyme of gluconeogenesis in Gram-negative bacteria. Based on these findings, we propose that AauR is a central regulator of acidic amino acid uptake and metabolism in pseudomonads.
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PMID:Characterization of a Pseudomonas putida ABC transporter (AatJMQP) required for acidic amino acid uptake: biochemical properties and regulation by the Aau two-component system. 1831 26

This report represents the conclusions of a Joint FAO/WHO Expert Committee convened to evaluate the safety of various food additives, including flavouring agents, with a view to recommending acceptable daily intakes (ADIs) and to preparing specifications for identity and purity. The first part of the report contains a general discussion of the principles governing the toxicological evaluation and assessment of intake of food additives (in particular, flavouring agents). A summary follows of the Committee's evaluations of technical, toxicological and intake data for certain food additives (asparaginase from Aspergillus niger expressed in A. niger, calcium lignosulfonate (40-65), ethyl lauroyl arginate, paprika extract, phospholipase C expressed in Pichia pastoris, phytosterols, phytostanols and their esters, polydimethylsiloxane, steviol glycosides and sulfites [assessment of dietary exposure]) and 10 groups of related flavouring agents (aliphatic branched-chain saturated and unsaturated alcohols, aldehydes, acids and related esters; aliphatic linear alpha,beta-unsaturated aldehydes, acids and related alcohols, acetals and esters; aliphatic secondary alcohols, ketones and related esters; alkoxy-substituted allylbenzenes present in foods and essential oils and used as flavouring agents; esters of aliphatic acyclic primary alcohols with aliphatic linear saturated carboxylic acids; furan-substituted aliphatic hydrocarbons, alcohols, aldehydes, ketones, carboxylic acids and related esters, sulfides, disulfides and ethers; miscellaneous nitrogen-containing substances; monocyclic and bicyclic secondary alcohols, ketones and related esters; hydroxy- and alkoxy-substituted benzyl derivatives; and substances structurally related to menthol). Specifications for the following food additives were revised: canthaxanthin; carob bean gum and carob bean gum (clarified); chlorophyllin copper complexes, sodium and potassium salts; Fast Green FCF; guar gum and guar gum (clarified); iron oxides; isomalt; monomagnesium phosphate; Patent Blue V; Sunset Yellow FCF; and trisodium diphosphate. Re-evaluation of flavouring agents for which estimated intake was based on anticipated poundage data was carried out for 2-isopropyl- N,2,3-trimethylbutyramide (No. 1595) and L-monomenthyl glutarate (No. 1414). Annexed to the report are tables summarizing the Committee's recommendations for intakes and toxicological evaluations of the food additives considered.
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PMID:Evaluation of certain food additives. 2011 97

In addition to their classical roles as carbon or nitrogen sources, amino acids can be used for bacterial virulence, colonization, or stress resistance. We found that original deamidase-transport systems impact colonization by Helicobacter pylori, a human pathogen associated with gastric pathologies, including adenocarcinoma. We demonstrated that l-asparaginase (Hp-AnsB) and gamma-glutamyltranspeptidase (Hp-gammaGT) are highly active periplasmic deamidases in H. pylori, producing ammonia and aspartate or glutamate from asparagine and glutamine, respectively. Hp-GltS was identified as a sole and specialized transporter for glutamate, while aspartate was exclusively imported by Hp-DcuA. Uptake of Gln and Asn strictly relies on indirect pathways following prior periplasmic deamidation into Glu and Asp. Hence, in H. pylori, the coupled action of periplasmic deamidases with their respective transporters enables the acquisition of Glu and Asp from Gln and Asn, respectively. These systems were active at neutral rather than acidic pH, suggesting their function near the host epithelial cells. We showed that Hp-DcuA, the fourth component of these novel deamidase-transport systems, was as crucial as Hp-gammaGT, Hp-AnsB, and Hp-GltS for animal model colonization. In conclusion, the pH-regulated coupled amino acid deamidase-uptake system represents an original optimized system that is essential for in vivo colonization of the stomach environment by H. pylori. We propose a model in which these two nonredundant systems participate in H. pylori virulence by depleting gastric or immune cells from protective amino acids such as Gln and producing toxic ammonia close to the host cells.
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PMID:Coupled amino acid deamidase-transport systems essential for Helicobacter pylori colonization. 2036 42

Cultural factors affecting the production of L-asparaginase by Streptomyces tendae isolated from laterite soil samples of Guntur region were investigated on glycerol-asparagine-salts (modified ISP-5) broth. Optimal yields of L-asparaginase were recorded in the culture medium with the initial pH 7.0 incubated at 30 degrees C for 72 h. The strain utilized sucrose (2%) and yeast (2%) extract as carbon and nitrogen sources for L-asparaginase production. The productivity of L-asparaginase was slightly enhanced when the strain was treated with cell-disrupting agents like EDTA. The crude enzyme was purified to homogeneity by ammonium sulfate precipitation, Sephadex G-100 and CM-Sephadex G-50 gel filtration. By employing sodium dodecyl sulfate-polyacrylamide gel electrophoresis, the molecular weight of the enzyme was recorded as 97.4 kDa. This is the first report on production and purification of L-asparaginase from S. tendae.
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PMID:Optimization and purification of L-asparaginase produced by Streptomyces tendae TK-VL_333. 2073 24


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