Glutamine synthetase mammalian

Covalent interconversion of enzymes is well established as a fundamental theme in metabolic regulation. The prototypic reversible interconverting systems include the sequence of phosphorylation/dephosphorylation steps in the activation of mammalian glycogen phosphorylase and pyruvate dehydrogenase as well as the nucleotidyla-tion/denucleotidylation using UTP and ATP in the bacterial glutamine synthetase cascade (see Fig. 1.). 

Elaborate cascades initiate the clotting of blood (Chapter 12) and the action of the protective complement system (Chapter 31). Cascades considered later in the book are involved in controlling transcription (Fig. 11-13) and in the regulation of mammalian pyruvate dehydrogenase (Eq. 17-9), 3-hydroxy-3-methyl-glutaryl-CoA reductase and eicosanoids (Chapter 21), and glutamine synthetase (Chapter 24). 

McDermott RH, Butler M (1993), Uptake of glutamate, not glutamine synthetase, regulates adaptation of mammalian cells to glutamine-free medium, J. Cell Sci. 104 51-58. 

Enzyme activity can be regulated by covalent modification or by noncovalent (allosteric) modification. A few enzymes can undergo both forms of modification (e.g., glycogen phosphorylase and glutamine synthetase). Some covalent chemical modifications are phosphorylation and dephosphorylation, acetylation and deacetylation, adeny-lylation and deadenylylation, uridylylation and deuridyly-lation, and methylation and demethylation. In mammalian systems, phosphorylation and dephosphorylation are most commonly used as means of metabolic control. Phosphorylation is catalyzed by protein kinases and occurs at specific seryl (or threonyl) residues and occasionally at tyrosyl residues these amino acid residues are not usually part of the catalytic site of the enzyme. Dephosphorylation is accomplished by phosphoprotein phosphatases ... 

Figure 4. Schematic representation of the metabolic fate of alanine in hepatocytes. Note that striking differences may exist between mammalian cell types on the one hand and individual amino acids on the other (see text). Solid and broken arrow lines refer to metabolic conversions and transport routes, respectively, and circles in membranes refer to specific transporters. Numbers refer to enzymes involved in alanine metabolism 1, alanine transaminase 2, pyruvate carboxylase 3, malate dehydrogenase 4, glutamate dehydrogenase 5, glutamine synthetase.

Use of the Glutamine Synthetase (CS) Expression System for the Rapid Development of Highly Productive Mammalian Cell Processes... 

Mammalian cell culture is becoming increasingly important for the production of high-volume biopharmaceutical proteins. This is driving improvements in process efficiency. This chapter provides examples of improvements in both the creation of cell lines and in cell culture optimization, focusing particularly on experience with the glutamine synthetase (GS) expression system. 

The mechanism of action of glutamine synthetase has been extensively studied, and both the bacterial and mammalian enzymes were reviewed in Volume X of this series (83, 84). Many experimental approaches have been applied to the question of the chemical activation of the y-carboxyl group of glutamate and its reaction with ammonia. All of these experiments supported the stepwise chemical mechanism outlined in reactions (24a) and (24b) (8J),... 

Glutamine synthetase catalyzes the ATP-dependent biosynthesis of glutamine according to equation (1). This enzyme plays a central role in nitrogen metabolism. In mammalian brain tissue, glutamine synthetase converts the neurotransmitter glutamate into glutamine. 

Glutamate dehydrogenase is one of three mammalian enzymes that can "fix" ammonia into organic molecules. The other two are glutamine synthetase and carbamoyl phosphate synthetase I. 

Wedler and Boyer (1972) were however unable to obtain any evidence for a sequential reaction mechanism for E. coli glutamine synthetase. Under their experimental conditions the enzyme failed to carry out any partial reaction unless all the substrates were present. They concluded that the reaction proceeds by a concerted mechanism with random substrate binding. Later work (Wedler and Horn, 1976) suggests that the mammalian enzyme... 

Phosphate inhibits angiosperm glutamine synthetase (Varner and Webster, 1955 O Neal and Joy, 1975) similarly to the mammalian enzyme (Meister, 1974), that is a competitive inhibition with respect to ATP. Glucosamine 6-phosphate which is inhibitory to the E. coli enzyme (Stadtman and Ginsburg, 1974) had no effect at 5 mM on the Lemna enzyme (Stewart and Rhodes, 1977). 

The production of therapeutic proteins in mammalian cells requires the expression of product-specific genes in the host cell line. The dihydrofolate reductase (DHFR) and the glutamine synthetase (GS) expression vector systems are most... 

To minimize the potential for mammalian or non pest toxicity the target should be present in plants, but not mammals. However, toxicity of xenobiotics is difficult to predict since it is often due to effects that are unrelated to inhibition of the target site. Most herbicidal inhibitors of glutamine synthetase (glufosinate) and acetyl-CoA carboxylase (diclofop and sethoxydim) have very low mammalian toxicity (2), yet both enzymes are found in animal tissues. 

Glutamine Synthetase The Major Mn(II) Enzyme in Mammalian Brain Frederick C. Wedler and Robert B. Denman... 

The following three characteristic mammalian liver and kidney enzymes are absent from muscle catalase, xanthine oxidase, and D-amino oxidase. The distribution of many other enzymes in mammals is limited to particular organs. Thus arginase occurs only in the liver, alkaline phosphatase in the intestinal mucosa, acid phosphatase in kidney, spleen, and prostate, 5-nucleotidase in the testis, and a-mannosidase in the epididymis (see Table 4.6). The blood is disproportionately rich in carbonic anhydrase, and the pancreas in ribonuclease. Glutamine synthetase, which condenses... 

The regulated step of this pathway in mammalian cells is the synthesis of carbamoyl phosphate from glutamine and C02, catalyzed by carbamoyl phosphate synthetase II (CPS U). CPS II is inhibited... 

Dihydroorotate dehydrogenase, the enzyme catalyzing the dehydrogenation of dihydroorotate to orotate (reaction 4 of the pathway Fig. 15-15), is located on the outer side of the inner mitochondrial membrane. This enzyme has FAD as a prosthetic group and in mammals electrons are passed to ubiquinone. The de novo pyrimidine pathway is thus compartmentalized dihydroorotate synthesized by trifunctional DHO synthetase in the cytosol must pass across the outer mitochondrial membrane to be oxidized to orotate, which in turn passes back to the cytosol to be a substrate for bifunctional UMP synthase. Mammalian cells contain two carbamoyl phosphate synthetases the glutamine-dependent enzyme (CPSase II) which is part of CAD, and an ammonia-dependent enzyme (CPSase /) which is found in the mitochondrial matrix, and which is used for urea and arginine biosynthesis. Under certain conditions (e.g., hyperammonemia), carbamoyl phosphate synthesized in the matrix by CPSase I may enter pyrimidine biosynthesis in the cytosol. 

T2. Tatibana, M., and Ito, K., Control of pyrimidine biosynthesis in mammalian tissues. 1. Partial purification and characterization of glutamine utilising carbamyl phosphate synthetase of mouse spleen and its tissue distribution. J. Biol. Chem. 244, 5403-5413 (1969). 

Mammalian cells contain two carbamoyl phosphate synthetases the glutamine-dependent enzyme (CPSase II) which is part of CAD and an ammonia-dependent enzyme (CPSase /) which is localized in the mitochondrial matrix and which is used in urea and arginine biosynthesis. Under certain conditions (e.g., hyperammonemia), carbamoyl phosphate synthesized in the matrix by CPSase I may enter pyrimidine biosynthesis in the cytoplasm. 

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