asparagine synthetase | |||||||
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Identifiers | |||||||
Symbol | ASNS | ||||||
Alt. symbols | 11as, AsnS | ||||||
NCBI gene | 440 | ||||||
HGNC | 753 | ||||||
OMIM | 108370 | ||||||
RefSeq | NM_001673 | ||||||
UniProt | P08243 | ||||||
Other data | |||||||
EC number | 6.3.5.4 | ||||||
Locus | Chr. 7 q21-q31 | ||||||
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Asparagine synthetase (or aspartate-ammonia ligase) is a chiefly cytoplasmic enzyme that generates asparagine from aspartate. [1] This amidation reaction is similar to that promoted by glutamine synthetase. The enzyme is ubiquitous in its distribution in mammalian organs, but basal expression is relatively low in tissues other than the exocrine pancreas. [2]
Above average presence of asparagine synthetase in certain leukemia strains has been linked to be a significant contributing factor of chemotherapy resistance, particularly to the chemotherapy drug, L-asparaginase. [3]
Escherichia coli derived asparagine synthetase is a dimeric protein with each subunit folding into two distinct domains. [4] The N-terminal region consists of two layers of six-stranded antiparallel β-sheets between which is the active site responsible for the hydrolysis of glutamine. [4] The C-terminal domain consists of a five-stranded parallel β-sheet flanked on either side by α-helices. This domain is responsible for the binding of both Mg2+ATP and aspartate. [4] These two active sites are connected by a tunnel lined primarily with backbone atoms and hydrophobic, nonpolar amino acid residues. [4]
Structural characterization of asparagine synthetase from mammalian sources have been difficult due to the low abundance and instability of the enzyme during purification procedures. [5]
Using information from Escherichia coli derived asparagine synthetase, some basic mechanisms of the enzyme have been understood. [5] The N-terminal active site catalyzes glutamine hydrolysis to yield glutamate and ammonia. [5] The C-terminal active site catalyzes activation of the side-chain carboxylate of aspartate to form an electrophilic intermediate, β-aspartyl-AMP (βAspAMP) 1, and inorganic pyrophosphate (PPi). [5] The tunnel that links the two active sites allows for the passage of an ammonia molecule to act as a common intermediate to couple the two half-reactions carried out in the independent active sites of the enzyme. [5] Thus, after being released in, and channeled from, the glutaminase site, the ammonia molecule attacks the bound βAspAMP 1 to give asparagine and AMP via a tetrahedral intermediate. [5]
In plants, inorganic nitrogen is taken up from the environment in forms of nitrate or ammonium. [6] Assimilation of this nitrogen into asparagine for use in nitrogen recycling, transport, and storage is an essential process for plant development, making asparagine synthetase vital to maintaining these asparagine reserves. [6] Specific events in development which depend on asparagine synthetase are nitrogen mobilization in germinating seeds, nitrogen recycling and flow in vegetative cells in response to biotic and abiotic stresses, and nitrogen remobilization from source to sink organs. [6]
In mammals, asparagine synthetase expression has been found to be linked to cell growth, and its mRNA content is linked to changes in the cell cycle. [5] Hamster BHK ts11 cells produce an inactive asparagine synthetase enzyme, and this loss of asparagine synthetase activity directly led to cell cycle arrest in the cells as a consequence of a depletion of cellular asparagine. [5] Upregulation of asparagine synthetase mRNA was observed as well in these hamster cells. [5] Other experiments demonstrated that quiescent rat thyroid cells entering S phase as a result of thyroid-stimulating hormone treatment was matched with a concurrent increase in asparagine synthetase mRNA content. [5]
There seem to be two major groups of asparagine synthetase: [7] [6]
Cancerous cells exhibit rapid growth and cell division and subsequently have an increased nutritional need. [5] The particularly low-level expression of asparagine synthetase in primary acute lymphoblastic leukemia (ALL) and numerous ALL cell lines, as compared to that of normal cells, makes asparagine depletion an effective method of treatment due to the cells' unusual dependency on circulating serum asparagine as a necessary nutrition for growth. [2] [5] As a result, L-asparaginase is a common chemotherapy drug utilized in the treatment of ALL and may have applications in other asparagine synthetase negative cancers, such as lymphomas, due to its aspariginase activity to deplete serum asparagine. [9] This depletion of serum asparagine leads to a subsequent rapid efflux of cellular asparagine, which is immediately acted upon and destroyed by the L-asparaginase as well. [5] Due to the transient response from these susceptible cancers in reaction to the asparagine depletion, tumor growth is significantly inhibited due to nutritional deficiency. [5] [3]
Most somatic cells express sufficient basal amounts of asparagine synthetase to counteract this asparagine starvation and survive the effects of L-asparaginase. [2] [3] [5] In addition, these normal cells are able to upregulate their expression of asparagine synthetase in response to the asparagine depletion, further countering some of the toxic effects of the medication on normal cell activity, a desirable trait for chemotherapy drugs. [2] [3] [5]
However, the opposite effect is visible in cases of asparaginase resistant cancers. [3] In these resistant cancers, the effect of blood asparagine depletion through L-asparaginase instead leads to significant asparagine synthetase overexpression to compensate, effectively nullifying the effect of the chemotherapy drug. [3] For example, in mouse models, 24 hours after exposure to L-asparaginase, tumors resistant to the depletion responded with 5- to 19-fold increases in asparagine synthetase expression. [10] These resistant tumors also inherently express higher levels of asparagine synthetase activity, even without the application of L-asparaginase to drive further expression. [11] Similar trends are often seen in human studies as well, with high levels of asparagine synthetase activity being detected in asparaginase-resistant cases of treatment as compared with the negligible activity of susceptible cases. [12] As seen in in vitro studies of resistant human leukemia cell lines, even six weeks after the removal of asparagine depleting factors, the increased level of expression of asparagine synthetase failed to return to a basal state, instead remaining elevated and retaining continued drug resistance. [13]
While the mechanisms underlying the sustained over-expression of ASNS have not been reported in these studies, transcriptome profiling of two T-ALL patients that have relapsed after L-asparaginase treatment revealed a recurrent promoter swap with KMT2E leading to ASNS over-expression and L-asparaginase resistance. [14] It has been further demonstrated in mouse model systems that repeated subculturing of L-asparaginase sensitive tumor cells in sublethal concentrations of L-asparaginase could eventually make them resistant, a potential worry of lower dose chemotherapy treatments effectively encouraging resistant cell development. [15]
A correlation between L-asparaginase efficacy and asparagine synthetase protein levels in a number of human ovarian cell lines has been observed. [16] As mentioned above, this result confirmed similar observations in human leukemia cell lines. [16] Hence asparagine synthetase might be used as a biomarker in ovarian cancer screening and potential treatment. [16]
An epithelial to mesenchymal transition was mimicked in metastatic cells by adapting PC-3 prostate cancer cells from adherent to suspension culture and then examined to investigate changes in gene expression concurrent with this adaption to suspension. [17] It was found that the asparagine synthetase expression was sixfold greater in the suspension cells than in the adherent cells. [17] In xenografts from a human breast cancer cell line in an established metastatic mouse model, [2] [18] asparagine synthetase was elevated in circulating tumor cells isolated from the mouse blood compared with the parental cell line. [2] [18] When these circulating tumor cells were returned to an in vitro culture and exposed to hypoxia, they showed higher basal expression and greater induction of asparagine synthetase than their parental cell line. [2] [18] These circulating tumor cells were also found to have an increased capacity for colony formation in soft agar assays under hypoxic conditions and grew faster when reimplanted as xenografts. [2] [18] The increased prevalence of asparaginase synthetase in the metastatic cells suggests that its activity may be beneficial for circulating tumor cell survival. [2] [18]
Guinea pigs have some of the highest levels of naturally expressing asparagine synthetase due to the fact that their serum inherently containing detectable levels of L-asparaginase. [10]
Asparagine is an α-amino acid that is used in the biosynthesis of proteins. It contains an α-amino group, an α-carboxylic acid group, and a side chain carboxamide, classifying it as a polar, aliphatic amino acid. It is non-essential in humans, meaning the body can synthesize it. It is encoded by the codons AAU and AAC.
In molecular biology, biosynthesis is a multi-step, enzyme-catalyzed process where substrates are converted into more complex products in living organisms. In biosynthesis, simple compounds are modified, converted into other compounds, or joined to form macromolecules. This process often consists of metabolic pathways. Some of these biosynthetic pathways are located within a single cellular organelle, while others involve enzymes that are located within multiple cellular organelles. Examples of these biosynthetic pathways include the production of lipid membrane components and nucleotides. Biosynthesis is usually synonymous with anabolism.
Glutamine synthetase (GS) is an enzyme that plays an essential role in the metabolism of nitrogen by catalyzing the condensation of glutamate and ammonia to form glutamine:
Asparaginase is an enzyme that is used as a medication and in food manufacturing. As a medication, L-asparaginase is used to treat acute lymphoblastic leukemia (ALL) and lymphoblastic lymphoma (LBL). It is given by injection into a vein, muscle, or under the skin. A pegylated version is also available. In food manufacturing it is used to decrease acrylamide.
Argininosuccinate synthase or synthetase is an enzyme that catalyzes the synthesis of argininosuccinate from citrulline and aspartate. In humans, argininosuccinate synthase is encoded by the ASS gene located on chromosome 9.
Amino acid synthesis is the set of biochemical processes by which the amino acids are produced. The substrates for these processes are various compounds in the organism's diet or growth media. Not all organisms are able to synthesize all amino acids. For example, humans can synthesize 11 of the 20 standard amino acids. These 11 are called the non-essential amino acids).
Indoleamine-pyrrole 2,3-dioxygenase (IDO or INDO EC 1.13.11.52) is a heme-containing enzyme physiologically expressed in a number of tissues and cells, such as the small intestine, lungs, female genital tract or placenta. In humans is encoded by the IDO1 gene. IDO is involved in tryptophan metabolism. It is one of three enzymes that catalyze the first and rate-limiting step in the kynurenine pathway, the O2-dependent oxidation of L-tryptophan to N-formylkynurenine, the others being indolamine-2,3-dioxygenase 2 (IDO2) and tryptophan 2,3-dioxygenase (TDO). IDO is an important part of the immune system and plays a part in natural defense against various pathogens. It is produced by the cells in response to inflammation and has an immunosuppressive function because of its ability to limit T-cell function and engage mechanisms of immune tolerance. Emerging evidence suggests that IDO becomes activated during tumor development, helping malignant cells escape eradication by the immune system. Expression of IDO has been described in a number of types of cancer, such as acute myeloid leukemia, ovarian cancer or colorectal cancer. IDO is part of the malignant transformation process and plays a key role in suppressing the anti-tumor immune response in the body, so inhibiting it could increase the effect of chemotherapy as well as other immunotherapeutic protocols. Furthermore, there is data implicating a role for IDO1 in the modulation of vascular tone in conditions of inflammation via a novel pathway involving singlet oxygen.
CAD protein is a trifunctional multi-domain enzyme involved in the first three steps of pyrimidine biosynthesis. De-novo synthesis starts with cytosolic carbamoylphosphate synthetase II which uses glutamine, carbon dioxide and ATP. This enzyme is inhibited by uridine triphosphate.
CTP synthase is an enzyme involved in pyrimidine biosynthesis that interconverts UTP and CTP.
In enzymology, an asparaginyl-tRNA synthase (glutamine-hydrolysing) is an enzyme that catalyzes the chemical reaction
In enzymology, an aspartate—ammonia ligase (ADP-forming) (EC 6.3.1.4) is an enzyme that catalyzes the chemical reaction
In enzymology, an omega-amidase (EC 3.5.1.3) is an enzyme that catalyzes the chemical reaction
Folylpolyglutamate synthase, mitochondrial is an enzyme that in humans is encoded by the FPGS gene.
Glutaminolysis (glutamine + -lysis) is a series of biochemical reactions by which the amino acid glutamine is lysed to glutamate, aspartate, CO2, pyruvate, lactate, alanine and citrate.
6-Diazo-5-oxo-L-norleucine (DON) is a glutamine antagonist, which was isolated originally from Streptomyces in a sample of Peruvian soil. This diazo compound is biosynthesized from lysine by three enzymes in bacteria. It is one of the most famous non-proteinogenic amino acid and was characterized in 1956 by Henry W Dion et al., who suggested a possible use in cancer therapy. This antitumoral efficacy was confirmed in different animal models. DON was tested as chemotherapeutic agent in different clinical studies, but was never approved. In 2019, DON was shown to kill tumor cells while reversing disease symptoms and improve overall survival in late-stage experimental glioblastoma in mice, when combined with calorie-restricted ketogenic diet.
CTP synthase 1 is an enzyme that is encoded by the CTPS1 gene in humans. CTP synthase 1 is an enzyme in the de novo pyrimidine synthesis pathway that catalyses the conversion of uridine triphosphate (UTP) to cytidine triphosphate (CTP). CTP is a key building block for the production of DNA, RNA and some phospholipids.
The Purine Nucleotide Cycle is a metabolic pathway in protein metabolism requiring the amino acids aspartate and glutamate. The cycle is used to regulate the levels of adenine nucleotides, in which ammonia and fumarate are generated. AMP converts into IMP and the byproduct ammonia. IMP converts to S-AMP (adenylosuccinate), which then converts to AMP and the byproduct fumarate. The fumarate goes on to produce ATP (energy) via oxidative phosphorylation as it enters the Krebs cycle and then the electron transport chain. Lowenstein first described this pathway and outlined its importance in processes including amino acid catabolism and regulation of flux through glycolysis and the Krebs cycle.
Asparagine synthase (glutamine-hydrolysing) (EC 6.3.5.4, asparagine synthetase (glutamine-hydrolysing), glutamine-dependent asparagine synthetase, asparagine synthetase B, AS, AS-B) is an enzyme with systematic name L-aspartate:L-glutamine amido-ligase (AMP-forming). This enzyme catalyses the following chemical reaction
The glnALG operon is an operon that regulates the nitrogen content of a cell. It codes for the structural gene glnA the two regulatory genes glnL and glnG. glnA encodes glutamine synthetase, an enzyme which catalyzes the conversion of glutamate and ammonia to glutamine, thereby controlling the nitrogen level in the cell. glnG encodes NRI which regulates the expression of the glnALG operon at three promoters, which are glnAp1, glnAp2 located upstream of glnA) and glnLp. glnL encodes NRII which regulates the activity of NRI. No significant homology is found in Eukaryotes.
Fructose-asparagine (F-Asn) is a glycosylamine compound that is most notably used by Salmonella during Salmonella-mediated inflammation of the intestine. In addition to Salmonella, several other species of bacteria may utilize fructose-asparagine as a nutrient. The name of the genetic locus that encodes the uptake capability in Salmonella is fra. This fra locus has five genes: fraR, fraB a fructose-asparagine deglycase, fraD a sugar kinase, fraA a fructose-asparagine transporter, and fraE a L-asparaginase. Notably, mutations in fraB cause the buildup of the toxic intermediate 6-phosphofuctose-aspartate (6-P-F-Asp). The buildup of 6-P-F-Asp has a bacteriostatic effect on fraB mutant cells, making FraB a potential drug target.