Gene/Protein Disease Symptom Drug Enzyme Compound
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Query: EC:6.3.5.5 (CPS)
 1,262 document(s) hit in 31,850,051 MEDLINE articles (0.00 seconds)

Carbamyl phosphate (CP) is synthesized in the liver by two separate enzymes, CPS I and CPS II. CPS I, an intramitochondrial enzyme involved in ureogenesis, has a relative activity of 500- to 1000-fold greater than CPS II, a cytoplasmic enzyme which initiates the sequence of reactions for pyrimidine biosynthesis. The contributions of NH4Cl (substrate for CPS I) ang glutamine (substrate for CPS II) as precursors for pyrimidine biosynthesis in isolated hepatocytes were compared by measuring their effect on uracil nucleotide pool size, the incorporation of NaH14CO3 into these pools, and the accumulation of orotic acid. Physiological concentrations of NH4Cl caused a marked stimulation of incorporation of radioactivity into uracil nucleotides (6-fold increase at 0.5 mM NH4Cl), and radioactive orotate appeared in both the cells and the medium. In contrast, glutamine (at concentrations up to 10 mM) had no effect on the incorporation of radioactivity into uracil nucleotides, and no orotic acid was detected. Uracil nucleotide pools were expanded up to 50% by low levels of NH4Cl, but there was no expansion of this pool in the presence of added glutamine. NH4Cl-driven pyrimidine de novo biosynthesis was insensitive to feedback inhibition by an expanded uracil nucleotide pool, to galactosamine treatment, and to acivicin treatment, indicating that NH+4 stimulated pyrimidine biosynthesis as a result of CP synthesis by mitochondrial CPS I. The consequence of intramitochondrially produced CP being available for pyrimidine biosynthesis is that the controlling step of this pathway (CPS II) is bypassed. The appearance of orotic acid following NH4Cl stimulation indicated that the rate-controlling step of hepatic de novo pyrimidine synthesis under these conditions was orotate phosphoribosyl transferase. These data indicate that, at physiological concentrations of NH+4, the majority of uracil nucleotides synthesized in isolated rat hepatocytes was derived from intramitochondrially generated CP. The effect of NH4Cl on the output of uridine by the isolated perfused rat liver was examined. In the presence of a single addition of 20 mM NH4Cl, the excretion of uridine was increased from 100-200 to 375 nmol h-1 g-1 liver and orotic acid was released into the circulating perfusate reaching a maximum of 2 microM (in 220 ml of perfusate) after 2 h. With 40 mM NH4Cl, uridine export was increased to 450 nmol h-1 g-1 and a maximum of 5 microM orotic acid was released into the perfusate after 2 h.(ABSTRACT TRUNCATED AT 400 WORDS)
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PMID:Influence of ammonium ions on hepatic de novo pyrimidine biosynthesis. 298 2

The inhibition of cytosolic carbamoyl-phosphate synthetase II by acivicin was used to study the role of the cytosolic carbamoyl phosphate pool as the exclusive substrate source for de novo pyrimidine synthesis in rat hepatocytes. De novo pyrimidine synthesis was stimulated: 1. by uridine triphosphate deficiency (incubation with D-galactosamine) leading to a stimulation of cytosolic carbamoyl phosphate synthesis, and 2. by accumulation and efflux of mitochondrial carbamoyl phosphate (incubation with ammonium ions and L-norvaline). The stimulated orotate formation from cytosolic carbamoyl phosphate in UTP depleted cells was completely blocked by acivicin. It was not influenced by an inhibition of mitochondrial carbamoyl phosphate synthesis mediated by 4-pentenoate, since mitochondrial carbamoyl phosphate did not participate in cytosolic pyrimidine synthesis even in the presence of ammonium ion concentrations maintaining physiological rates of urea synthesis. An excess of ammonium ions led to an artificial accumulation and efflux of mitochondrial carbamoyl phosphate, which could be avoided by 4-pentenoate. The non-regulated stimulation of pyrimidine synthesis from surplus mitochondrial carbamoyl phosphate was not inhibited by acivicin. Utilization of mitochondrial carbamoyl phosphate for de novo pyrimidine synthesis presumably does not occur under physiological conditions because mitochondrial CP efflux depends on the accumulation of this metabolite in the mitochondria under experimental or pathological circumstances. Acivicin inhibition of CPS II thus cannot be bypassed by mitochondrial CP. It is suitable as inhibitor of the physiological de novo pyrimidine synthesis.
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PMID:The glutamine analog acivicin as antipyrimidine. Studies on the interrelationship between pyrimidine and urea synthesis in liver. 383 21

The interrelationship between the two carbamoyl phosphate pools in intact hepatocytes and intact liver was studied with respect to de novo pyrimidine synthesis by use of selective inhibitors of the mitochondrial and the cytosolic carbamoyl-phosphate synthetase. Inhibition of mitochondrial carbamoyl phosphate synthesis by 4-pentenoate was without effect on galactosamine-stimulated pyrimidine synthesis. Conditions favouring mitochondrial carbamoyl phosphate accumulation, like excess ammonium ions or L-norvaline, led to an increase in pyrimidine synthesis bypassing the feedback inhibition of cytosolic carbamoyl-phosphate synthetase by UTP. A stimulation of pyrimidine synthesis was not observed when the carbamoyl phosphate accumulation was due to aspartate deficiency in the presence of aminooxyacetate. The full response of pyrimidine synthesis to excess ammonium ions was restored, even in the presence of aminooxyacetate, when aspartate was substituted. This is explained by an inhibition of aspartate carbamoyltransferase flux [in view of the Km (aspartate = 0.7 mmol/l) of this enzyme] resulting from a 90% decrease in aspartate tissue levels. Acivicin, the inhibitor of cytosolic carbamoyl-phosphate synthetase, completely abolished the galactosamine-induced stimulation of pyrimidine synthesis, but was without effect on the stimulation of pyrimidine synthesis by ammonium ions and L-norvaline. It is concluded that experimental changes in mitochondrial carbamoyl phosphate content exert effects on de novo pyrimidine synthesis; however, it is considered unlikely that relevant amounts of mitochondrial carbamoyl phosphate are used for pyrimidine synthesis under physiological conditions. In addition the data point to a potential regulatory role of aspartate in hepatic pyrimidine synthesis.
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PMID:Hepatic carbamoyl phosphate metabolism. Role of cytosolic and mitochondrial carbamoyl phosphate in de novo pyrimidine synthesis. 401 77

The glutamine antagonist acivicin, L-(alpha S, 5S)-alpha-amino-3-chloro-4,5-dihydro-5-isoxazoleacetic acid, strongly reduced CTP and GTP contents in AS-30D rat hepatoma cells in suspension. UTP only dropped to 63% of the respective control after 4 hr; however, by combining acivicin with the uridylate-trapping sugar analogue D-galactosamine, a synergistic decrease in UTP contents to 7% of control was induced. Incorporation of 14CO2 into purine and pyrimidine nucleotides followed by radio-high performance liquid chromatography showed marked inhibition of purine and pyrimidine biosynthesis de novo; the latter was reduced to 35% of control. The inhibitory potency of acivicin on glutamine-dependent carbamoyl-phosphate synthetase and consequently on de novo uracil nucleotide formation was also reflected by the complete suppression of the D-galactosamine-induced rise in total uridylate. Induction of UTP deficiency by interference with the first and rate-limiting step in pyrimidine biosynthesis de novo together with a trapping of uridylate by D-galactosamine may provide a promising approach to the chemotherapy of hepatocellular carcinoma.
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PMID:Combined action of acivicin and D-galactosamine on pyrimidine nucleotide metabolism in hepatoma cells. 688 63

Chemical composition of lipopolysaccharide (LPS) isolated from an effective (97) and ineffective (87) strains of R. l. viciae has been determined. LPS preparations from the two strains contained: glucose, galactose, mannose, fucose, arabinose, heptose, glucosamine, galactosamine, quinovosamine, and 3-N-methyl-3,6-dideoxyhexose, as well as glucuronic, galacturonic and 3-deoxyoctulosonic acid. The following fatty acids were identified: 3-OH 14:0, 3-OH 15:0, 3-OH 16:0, 3-OH 18:0 and 27-OH 28:0. The ratio of 3-OH 14:0 to other major fatty acids in LPS 87 was higher that in LPS 97. SDS/PAGE profiles of LPS indicated that, in lipopolysaccharides, relative content of S form LPS I to that of lower molecular mass (LPS II) was much higher in the effective strain 97 than in 87. All types of polysaccharides exo-, capsular-, lipo, (EPS, CPS, LPS, respectively) examined possessed the ability to bind faba bean lectin. The degree of affinity of the host lectin to LPS 87 was half that to LPS 97. Fatty acids (FA) composition from bacteroids and peribacteroid membrane (PBM) was determined. Palmitic, stearic and hexadecenoic acids were common components found in both strains. There was a high content of unsaturated fatty acids in bacteroids as well as in PBM lipids. The unsaturation index in the PBM formed by strain 87 was lower than in the case of strain 97. Higher ratio of 16:0 to 18:1 fatty acids was characteristic for PMB of the ineffective strain.
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PMID:Chemical characterization of effective and ineffective strains of Rhizobium leguminosarum bv. viciae. 1082 71

Small leucine-rich proteoglycans, such as biglycan, and their side chain sulfated glycosaminoglycans (GAGs), have been suggested to be involved in bone formation and mineralization processes. The present study was designed to investigate whether chondroitin sulfate (CS), one of the GAG, and its oversulfated structures coupled with bone morphogenetic protein-4 (BMP-4) alter the differentiation and subsequent mineralization of MC3T3-E1 osteoblastic cells. CS-E, one of the oversulfated CS structure, enhanced cell growth, alkaline phosphatase (ALP) activity, collagen deposition, and mineralization whereas heparin enhanced only ALP activity and mineralization. As well as CS-E, CS-H, and CPS also enhanced the mineralization of the cells. CS-E enhanced the mineralization of the cells by interacting with protein in the conditioned medium. CS-E induced mineralization was significantly inhibited by an antibody against BMP-4. The addition of exogenous BMP-4 further increased the capacity of CS-E to enhance mineralization. Fluorescence correlation spectroscopy method using fluoresceinamine-labeled GAG revealed that the oversulfated GAGs have a high affinity for BMP-4. The disaccharide analysis of the cells indicated that MC3T3-E1 cells are capable of producing oversulfated structures of CS by themselves. The lack of CS from the cells after chondroitinase treatment resulted in the inhibition of mineralization. These results in the present study indicate that oversulfated CS, which possesses 4,6-disulfates in N-acetyl-galactosamine, binds to BMP-4 and promotes osteoblast differentiation and subsequent mineralization.
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PMID:Oversulfated chondroitin sulfate-E binds to BMP-4 and enhances osteoblast differentiation. 1872 Mar 84