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

---

Query: EC:6.3.5.5 (CPS)
1,262 document(s) hit in 31,850,051 MEDLINE articles (0.00 seconds)

A combined genetic, biochemical, and immunological approach has clarified structural relationships involving the first three enzymes of de novo pyrimidine biosynthesis. A procedure involving antibody and protein A-Sepharose was used to isolate the enzymes carbamoyl-phosphate synthase [ATP:carbamate phosphotransferase (dephosphorylating, amido-transferring), EC 2.7.2.9], aspartate transcarbamoyltransferase (carbamoylphosphate:L-aspartate carbamoyltransferase, EC 2.1.3.2), and dihydro-orotase (L-5,6-dihydroorotate amidohydrolase, EC 3.5.2.3) from Chinese hamster ovary cell CHO-K1, the uridine-requiring auxotroph Urd(-)A, and selected Urd(-)A revertants. The enzymes of Urd(-)A and the Urd(-)A revertants were significantly altered in activity, native structure, and molecular weight from those of CHO-K1. The results presented permit the conclusion that (i) these three enzymes reside in a single multifunctional 220,000-dalton polypeptide; (ii) the aspartate transcarbamoyltransferase activity is located on a portion ( approximately 20,000 daltons) at one end of the polypeptide; (iii) this portion may also be required for monomers to aggregate into the multimeric from present in mammalian cells; (iv) the mutations in Urd(-)A and the Urd(-)A revertants lie in the structural gene for this multifunctional protein; and (v) increased sensitivity to proteases could account for the alterations in the structure of these enzymes in the mutants.
...
PMID:Alteration in structure of multifunctional protein from Chinese hamster ovary cells defective in pyrimidine biosynthesis. 3 10

Mutants resistant to 5-fluorouracil, 5-fluorouridine and 5-fluorodeoxyuridine have been selected in Aspergillus nidulans. Growth tests combined with genetic analysis showed that mutations conferring resistance to fluoropyrimidines could occur in at least seven genes. Three of these fulE, fulF and furA were concerned with either the uptake of pyrimidines or their conversion to uridine monophosphate. The other four genes did not affect these functions. Mutations in fulA probably confer resistance by lowering ornithine transcarbamoylase, thereby making the normally arginine-specific carbamoyl phosphate pool available for increased uracil synthesis. Mutations in fulD may make the arginine-specific carbamoyl phosphate synthetase insensitive to inhibition or repression by arginine, and so lead to increased carbamoyl phosphate pool sizes, and increased uracil synthesis. Both fulA and fulD mutants suppress pyrA mutants which lack the uracil-specific carbamoyl phosphate synthetase. Mutations in fulB and fulC do not suppress pyrA, and so may act more directly to increase uracil synthesis. The synthesis of aspartate carbamoyl transferase in fulB7 strains is not repressed by uracil. fulC mutants are closely linked to the pyrA, B, C, N region which codes for the first two enzymes of pyrimidine biosynthesis, and may result in these enzymes being less sensitive to inhibition by uracil.
...
PMID:Pyrimidine biosynthesis in Aspergillus nidulans. Isolation and characterisation of mutants resistant to fluoropyrimidines. 12 29

The purimidine-3 locus of Neurospora crassa specifies two enzyme activities, pyrimidine-specific carbamyl phosphate synthetase (CPSpyr) and aspartate transcarbamylase (ATC). ATC is translationally distal. CPSpyr, but not ATC, is subject to feedback inhibition by uridine triphosphate (UTP). To investigate the location of the feedback-specific region within the locus, inhibition of a number of pyr-3 alleles by UTP was investigated. All CPS+ ATC- polar alleles, revertants of CPS- ATC- polar alleles, and 5-fluorouracil-resistant mutants had normal UTP response. The location of the feedback-specific region is in or close to the CPS-specific region.
...
PMID:The location of the feedback-specific region with the pyrimidine-3 locus of Neurospora crassa. 18 23

The pyrimidine-3 locus of Neurospora crassa specifies a multienzyme complex comprising pyrimidine-specific carbamoyl phosphate synthase (CPSpyr) and aspartate carbamoyl transferase (ACT). It appears to be divided into a translationally proximal CPS-specific region and a distal ACT-specific region. Levels of complementation for ACT activity between pairs of four pyr-3 CPS+ ACT- mutants showed a range from 12% to 68% of the wild-type level of the enzyme. This is interpreted as interallelic complementation, contradicting certain earlier suggestion of two dissimilar ACT subunits. Proteolysis of an extract from a heterokaryon formed from two of the above CPS+ ACT- alleles (alpha and beta) did not lead to loss of ACT activity, but led to the formation of a fragment with ACT activity with a similar molecular weight (92,000 daltons) to that produced in extracts of wild type strain. The pyr-3 polar mutant 43-174 which is enzymatically CPS+ ACT- and which fails to complement with any other CPS+ ACT- alleles, thus suggesting its location towards the proximal end of the ACT region, has CPS activity associated with a form of 180,000 daltons molecular weight. These findings are used to contruct a model for structure of the native enzyme complex.
...
PMID:A possible model for the structure of the Neurospora carbamoyl phosphate synthase-aspartate carbamoyl transferase complex enzyme. 20 7

1. Ammonia liberated continuously in large amounts in muscle, kidney and brain is used immediately for the synthesis of mainly glutamine because of the toxic effects of elevated ammonia concentrations. After glutamine hydrolysis in the liver ammonia serves as substrate for the urea biosynthesis. In ureotelic animals urea is the quantitatively most important product for the elimination of surplus nitrogen. 2. The rate of urea biosynthesis depends on the amount of surplus nitrogen and acts as regulatory factor for the nitrogen balance of the adult organism. 3. Urea cycle abnormalities in liver diseases or inborn enzymatic defects are important factors leading to hyperammonaemia in patients. 4. The hyperammonaemia induces an increase of the rate of hepatic pyrimidine nucleotide biosynthesis as a consequence of an ineffective feedback inhibition of the glutamine-dependent carbamoyl phosphate synthetase. 5. The distribution of ammonia between intra- and extracellular space and the amount of ammonium ions excreted in the urine depend on the pH value. An alkalosis induces an intracellular ammonia load and inhibits the urinary ammonium ion excretion, which is increased in acidosis as one mechanism of protein elimination. 6. The ammonia-induced inhibition of the citric acid cycle by an alpha-ketoglutarate deficiency is one important reason for the neurotoxicity of ammonia, which is the main point in the pathogenesis of hepatic coma.
...
PMID:[Biochemical and pathophysiological aspects of hyperammonaemia (author's transl)]. 31 94

Glutamine-dependent carbamoyl-phosphate synthetase was purified about 2100-fold from the cytosol of rat liver using 30% (v/v) dimethyl sulfoxide and 5% (w/v) glycerol as stabilizers. Throughout the purification, aspartate transcarbamylase and dihydroorotase, the second and third enzymes of pyrimidine biosynthesis, were copurified with the synthetase. These three enzymes sedimented as a single peak with a sedimentation coefficient of 27 S in sucrose gradients containing the stabilizers, indicating their existence as a multienzyme complex. The aggregation states of the complex were analyzed by sucrose gradient centrifugation under conditions approximating those used for enzymatic assay and correlated with the kinetic properties of the synthetase. In the presence of 10% glycerol and 10 mM MgATP(2-) at 18 degrees, the synthetase showed high activity and the three enzymes sedimented as a single peak with a coefficient of 25 S. The three enzymes also existed as a complex with the same coefficient when 50 muM PP-ribose-P was added in place of MgATP(2-), the sedimentation coefficient of the complex shifted to 28 S, indicating alteration in its molecular shape, rather than size. With 10% glycerol alone, the complex partially dissociated and the synthetase activity appeared in three peaks with coefficients of 26, 19, and 9 S (carbamoyl-phosphate synthetases (CPSase) a, b, and c, respectively). CPSases a, b, and c, thus obtained, were all sensitive to regulation by UTP and PP-ribose-P, but they differed MgATP(2-) (5.1, 4.8, AND 1.7 mM for CPSases a and b, and the enzyme within the original complex, respectively) and in their sensitivities to effectors. These results suggest that the aggregation may modify the catalytic and regulatory properties of the synthetase; Attempts to reassociate the components were unsuccessful.
...
PMID:Aggregation states and catalytic properties of the multienzyme complex catalyzing the initial steps of pyrimidine biosynthesis in rat liver. 114 71

Measurements have been made of the activities of the enzymes of the de novo and salvage pathways of pyrimidine synthesis (carbamoyl phosphate synthetase II (glutamine) (EC 6.3.5.5); dihydroorotate dehydrogenase (EC 1.3.99.11); the overall activity of Complex II (orotate phosphoribosyl pyrophosphate transferase (EC 2.4.2.10) and orotidine 5-phosphate decarboxylase (EC 4.1.1.23); uracil phosphoribosyltransferase (EC 2.4.2.9)) in the mammary gland of rats at different stages of the lactation cycle and the effects of diabetes on the activity of these enzymes in lactation have been studied. From a consideration of the changes in enzyme activities and the changes in the tissue concentration of phosphoribosyl pyrophosphate, an activator of the de novo pathway and substrate for both the de novo and salvage routes, it is concluded that the de novo pathway is the major route of pyrimidine synthesis in mammary tissue. Diabetes decreases the activity of the enzymes of the de novo pathway; the effects are particularly marked for Complex II. The present results on pyrimidine synthesis are compared to the pattern for purine synthesis previously published.
...
PMID:Pyrimidine nucleotide synthesis in the rat mammary gland: changes in the lactation cycle and effects of diabetes. 147 92

We have measured the 'core' mammalian carbamoyl-phosphate synthetase II (CPSII) activity, using NH4Cl as the nitrogen-donating substrate and trapping carbamoyl phosphate as urea through its reaction with ammonium ions. When ATP and magnesium ion concentrations are close to those found in the cell, the substrate saturation curves for ammonia and bicarbonate are hyperbolic, giving Km (NH3) values of 166 microM at high ATP concentrations and 26 microM at low ATP concentrations, while the Km (bicarbonate) is 1.4 mM at both ATP concentrations used. These values for the Km (NH3) are lower than previously reported for CPS II, and closer to the values for the mitochondrial counterpart. The Km for ammonia and bicarbonate are not altered by phosphorylation of the multienzyme polypeptide CAD, which contains the first three enzyme activities of pyrimidine biosynthesis. The CPS II activity is lower with an excess of either ATP or magnesium ions, causing the apparently sigmoid dependence of activity upon ATP concentration to be enhanced at low concentrations of free magnesium ions. The feedback inhibitor, UTP, acts by stabilising a state with a low affinity for magnesium ions and for ATP. In the presence of the activator, 5-phosphoribosyl diphosphate (PRibPP), the enzyme has a higher affinity for magnesium ions and thus the ATP dependence of the activity is hyperbolic. Phosphorylation of CAD similarly activates the CPS II enzyme by increasing the affinity for magnesium ions and by pushing the equilibrium away from the low-affinity UTP-stabilised state. Using our improved assay procedure, we observe a very large activation by PRibPP of carbamoylphosphate synthesis at low concentrations of magnesium ions, and we find that unlike UTP, the activator PRibPP is able to act on the phosphorylated enzyme.
...
PMID:Regulation of the mammalian carbamoyl-phosphate synthetase II by effectors and phosphorylation. Altered affinity for ATP and magnesium ions measured using the ammonia-dependent part reaction. 149 69

The CAD multidomain protein, which includes active sites of carbamyl phosphate synthetase II (CPS II, glutamine-dependent), aspartate transcarbamylase, and dihydroorotase, was immunostained in normal rat brains, the gliotic brains of myelin-deficient mutant rats, and brains from normal weanling hamsters. In each of these tissues CAD was observed in cells resembling astrocytes. In hamster brain, CAD immunofluorescence was also found in cells closely related to astrocytes, i.e., the Bergmann glia in cerebellum and the tanycytes surrounding the third ventricle. The astrocytic identity of the CAD-positive cells in rat brain was confirmed by double immunofluorescence staining with antibodies against glial fibrillary acidic protein (GFAP). The two enzymes carbonic anhydrase and glutamine synthetase occur in the cytoplasm of normal astrocytes in gray matter and of reactive astrocytes during gliosis. Products of each enzyme, i.e., bicarbonate and glutamine, are required for the CPS II reaction, which is the first step in the biosynthesis of pyrimidines. Therefore, the present results suggest roles for carbonic anhydrase and glutamine synthetase, as well as CAD, in pyrimidine biosynthesis in brain and a role for the astrocytes in the de novo synthesis of pyrimidines.
...
PMID:Localization of the multifunctional protein CAD in astrocytes of rodent brain. 167 39

1. Addition of concanavalin A to T-cell lymphocytes from rat cervical lymph nodes increases the activity of glutaminase within 1 h and those of carbamoyl-phosphate synthase II and aspartate transcarbamoylase within 3 h. There was a similar time course for the effects of concanavalin A on rates of glutamine utilization, which was increased within 1 h, and on pyrimidine nucleotide synthesis, which was increased by 40% at 2 h and by 100% at 3 h. 2. A delay in the addition of glutamine to the culture medium after addition of concanavalin A caused a decrease in [3H]thymidine incorporation only after 4-6 h. In the absence of glutamine, delay in addition of guanosine or inosine caused a decrease in [3H]thymidine incorporation only after 6-8 h after the addition of concanavalin A. 3. In contrast, a delay in addition of adenosine or uridine to the culture medium had an immediate effect (i.e. within 2 h) on the rate of incorporation of [3H]thymidine. It is suggested that adenosine and uridine have specific effects on proliferation via specific receptors for these nucleosides in the membrane.
...
PMID:The effect of time of addition of glutamine or nucleosides on proliferation of rat cervical lymph-node T-lymphocytes after stimulation by concanavalin A. 189 39


1 2 3 4 5 6 7 8 9 10 Next >>

---