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)

Hyperornithinemia, hyperammonemia and homocitrullinuria (HHH disorder) is an inherited metabolic disorder which shows peculiar amino acid changes in the serum and urine. The primary defect is considered to be the transport of ornithine across the mitochondrial membrane, but there is no direct evidence for this so far. We have analyzed ornithine transport activities in the liver mitochondria from three patients with HHH disorder. In coupled liver mitochondria we demonstrated low activities of citrulline synthesis and low rates of ornithine uptake. However, there were no abnormalities in carbamoyl-phosphate synthetase activity, ornithine carbamoyltransferase activity, N-acetylglutamate levels or O2 uptake with succinate. We also performed a kinetic study of citrulline synthesis as a function of ornithine concentration. We found increased Km values for ornithine and varied Vmax values of citrulline synthesis, which suggested the presence of a mutant transport protein. From these results we conclude that the defect of hyperornithinemia, hyperammonemia and homocitrullinuria lies in the transport of ornithine across the mitochondrial membrane.
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PMID:Biochemical analysis of decreased ornithine transport activity in the liver mitochondria from patients with hyperornithinemia, hyperammonemia and homocitrullinuria. 333 77

A carbamoyl-phosphate synthase has been purified from mycelia of Phycomyces blakesleeanus NRRL 1555 (-). The molecular weight of the enzyme was estimated to be 188,000 by gel filtration. Polyacrylamide gel electrophoresis in the presence of sodium dodecyl sulfate showed that the enzyme consists of two unequal subunits with molecular weights of 130,000 and 55,000. The purified enzyme has been shown to be highly unstable. The carbamoyl-phosphate synthase from Phycomyces uses ammonia and not L-glutamine as a primary N donor and does not require activation by N-acetyl-L-glutamate, but it does require free Mg2+ for maximal activity. Kinetic studies showed a hyperbolic behavior with respect to ammonia (Km 6.34 mM), bicarbonate (Km 10.5 mM) and ATP.2 Mg2+ (Km 0.93 mM). The optimum pH of the enzyme activity was 7.4-7.8. The Phycomyces carbamoyl-phosphate synthase showed a transition temperature at 38.5 degrees C. It was completely indifferent to ornithine, cysteine, glycine, IMP, dithiothreitol, glycerol, UMP, UDP and UTP. The enzyme was inhibited by reaction with 5 mM N-ethylmaleimide.
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PMID:Carbamoyl-phosphate synthase in Phycomyces blakesleeanus. 339 22

Canaline and gabaculine, inhibitors of gamma-aminotransferases and thus of ornithine aminotransferase (E.C. 2.6.1.13), decreased the flow through ornithine carbamoyl transferase (E.C. 2.1.3.3) in isolated rat hepatocytes incubated with 10 mM NH4Cl and ornithine. The levels of acetylglutamate, an essential activator of carbamoyl phosphate synthetase (ammonia) (E.C. 6.3.4.16), were also decreased, suggesting that the inhibitors had also caused a decrease in the rate of carbamoyl phosphate synthesis. Under these conditions, ornithine appears to be a precursor of acetylglutamate, via ornithine aminotransferase, possibly as a consequence of glutamate synthesis. The influence of aminooxyacetate, an aminotransferase inhibitor, has also been examined.
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PMID:Effects of inhibition of ornithine aminotransferase or of general aminotransferases on urea and citrulline synthesis and on the levels of acetylglutamate in isolated rat hepatocytes. 339 32

Carbamoyl-phosphate synthetase from Escherichia coli is subject to allosteric activation by ornithine, allosteric inhibition by uridine 5'-phosphate (UMP), and reversible concentration-dependent self-association. Positive allosteric effectors, magnesium adenosine 5'-triphosphate (MgATP), K+, and inorganic phosphate facilitate association. The purpose of this study was to determine the state of association of carbamoyl-phosphate synthetase in the presence and absence of different substrates and effectors and to consider the basis for the observed effects of enzyme concentration on specific activity. Studies employing gel filtration chromatography have shown that when the concentration of carbamoyl-phosphate synthetase is low (less than 0.01 mg/mL), the enzyme exists as monomer under all conditions, including the presence of UMP in phosphate buffer and the presence of all substrates plus ornithine (conditions that support maximal catalytic activity). At higher enzyme concentrations (e.g., greater than 0.01 mg/mL) the specific activity increases with increasing enzyme concentration when MgATP is nonsaturating but is independent of enzyme concentration when MgATP is saturating or when ornithine is present with MgATP being either saturating or nonsaturating. These results indicate that the catalytic activity of this enzyme is not directly linked to oligomer formation. The theoretical properties and possible significance of a generalized model of enzyme association-dissociation in which the active monomeric form, in equilibrium with another monomeric form, is specifically subject to self-association but the different states of association have the same specific activity, are discussed.(ABSTRACT TRUNCATED AT 250 WORDS)
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PMID:Carbamoyl-phosphate synthetase: an example of effects on enzyme properties of shifting an equilibrium between active monomer and active oligomer. 353 81

Low molecular weight phosphoryl compounds, such as carbamoyl phosphate, 2,3-diphosphoglycerate and phytic acid protect, to different extents, mitochondrial and cytosolic proteins such as ornithine transcarbamoylase (OTC), carbamoyl phosphate synthetase (CPS), glutamate dehydrogenase (GDH) and glyceraldehyde 3-phosphate dehydrogenase (GAPDH), from proteolytic inactivation (rat liver lysosomal extracts, pronase, elastase). Given the wide variety and common occurrence of low molecular weight reagents such as typified here, it seems that this kind of inhibition may be important in the regulation of protein turnover. Regulation of intracellular proteolysis can also occur via the proteolytic systems. Immunocytochemical procedures for mitochondrial enzymes (CPS, GDH, OTC), show intracellular homogeneity, but intercellular heterogeneity in rat liver, compatible with a role of the autophagic-lysosomal system in degrading these proteins. However, degradation of short-lived proteins occurs by other mechanisms. Using centrifugation of cultured cells, we find that the Golgi apparatus takes part in the degradation of these proteins, probably by controlling the traffic of proteins or proteases to the degradation site.
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PMID:Regulatory mechanisms of intracellular proteolysis in mammalian cells. 355 76

The intramitochondrial localization of the urea cycle enzymes, carbamoyl phosphate synthetase and ornithine transcarbamoylase, has been examined by both in vitro and in situ studies. The following three lines of evidence are presented to establish that significant fractions of the rat liver enzymes are loosely associated with the inner mitochondrial membrane: 1) when the mitochondrion is fractionated, the enzymes partition between the matrix and membrane fractions in the absence of detergent and partition solely to the matrix in the presence of detergent; 2) the purified enzymes associate with purified inner membrane preparations; and, 3) protein A-gold electron microscopic immunocytochemical analysis of rat liver sections reveals a nonrandom arrangement of the enzyme, with the maximal enzyme density adjacent to the inner mitochondrial membrane. These findings serve as the basis for novel potential mechanisms for regulation of the activity of the enzymes and provide additional evidence for the extensive organization of the mitochondrial matrix. The membrane interaction might also serve as the organizing factor for a carbamoyl phosphate synthetase-ornithine transcarbamoylase or other multienzyme complex.
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PMID:The interaction of rat liver carbamoyl phosphate synthetase and ornithine transcarbamoylase with inner mitochondrial membranes. 368 Feb 20

Studies were conducted to determine whether rainbow trout fingerlings possess the ability to synthesize arginine via the urea cycle. Several urea cycle enzymes were detected in trout tissues. An experiment was conducted to determine whether the enzymes increase in response to starvation or in response to dietary protein level (0, 30, 40, 50% protein). Although some effects were observed, they did not appear to be consistent with the function of the urea cycle as a mechanism of detoxifying ammonia in the fish. The activities of kidney arginase and liver and muscle carbamoyl phosphate synthetase (CPS) were higher (P less than 0.05) when protein was omitted from the diet (P less than 0.05) than when it was present but were unaffected by protein level otherwise. The activities of liver arginase and kidney and muscle CPS and ornithine transcarbamoylase (OTC) were higher (P less than 0.05) in starved fish than in fish that received adequate levels of protein. Liver CPS and OTC were lower in starved fish than in fish fed 30% protein. L-[l-14C]ornithine hydrochloride and L-[carbamoyl-14C]citrulline, injected intraperitoneally, were incorporated into tissue arginine, a finding consistent with arginine biosynthesis via the urea cycle. When one-half of dietary arginine was replaced by equimolar amounts of glutamic acid, ornithine or citrulline, glutamic acid markedly reduced growth (P less than 0.05), whereas growth was depressed only slightly by ornithine (P less than 0.05) and not depressed by citrulline (P greater than 0.05). We conclude that trout have a urea cycle that provides for potential arginine biosynthesis.
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PMID:Urea cycle activity and arginine formation in rainbow trout (Salmo gairdneri) 376 Oct 21

Reuber hepatoma H-35 cells actively synthesize the urea cycle enzyme, carbamoyl-phosphate synthetase I. Treatment of H-35 cells with dexamethasone (0.14 microM), however, enhanced synthesis of the enzyme (as measured by incorporation of [35S]methionine) by 4-5-fold. Insulin (0.18 microM) completely inhibited dexamethasone-dependent stimulation of enzyme synthesis. In vitro translation and cDNA hybridization assays were employed to measure effects of dexamethasone plus or minus insulin on levels of mRNA encoding the biosynthetic precursor of carbamoyl-phosphate synthetase I (pCPS) in Reuber H-35 cells. Both measurements yielded similar results: dexamethasone increased pCPS mRNA levels by 4-5-fold and insulin suppressed this response, but only by 50%. Specific cDNA hybridization assays also demonstrated that Reuber H-35 cells, even after hormone treatments, contain only very low levels of albumin mRNA, and no detectable ornithine carbamoyl-transferase mRNA.
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PMID:Expression of carbamoyl-phosphate synthetase I mRNA in Reuber hepatoma H-35 cells. Regulation by glucocorticoid and insulin. 389 Sep 50

The effect of alloxan diabetes on citrulline formation from NH4Cl and bicarbonate was studied in rabbit liver mitochondria incubated with glutamate or succinate as respiratory substrate, as well as with exogenous ATP in the presence of uncoupler and oligomycin. In contrast to ornithine transcarbamoylase, the activity of carbamoyl-phosphate synthetase (ammonia) was higher in mitochondria from diabetic animals than in those from normal ones. In diabetic rabbits the rates of citrulline synthesis were stimulated under all conditions studied. In contrast, levels of N-acetylglutamate, an activator of carbamoyl-phosphate synthetase (ammonia), were significantly increased only in the presence of glutamate, while the highest rates of citrulline formation occurred in uncoupled mitochondria incubated with exogenous ATP as energy source. Treatment of animals with alloxan resulted in an increase of both the intramitochondrial ATP level and the rate of adenine nucleotide translocation across the mitochondrial membrane. The results indicate that the stimulation of citrulline formation in liver mitochondria of diabetic rabbits is mainly due to an increase in carbamoyl-phosphate synthetase (ammonia) activity and an elevation of content of intramitochondrial ATP, a substrate of this enzyme.
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PMID:The stimulatory effect of alloxan diabetes on citrulline formation in rabbit liver mitochondria. 397 23

Control of urea synthesis was studied in rat hepatocytes incubated with physiological mixtures of amino acids in which arginine was replaced by equimolar amounts of ornithine. The following observations were made. Intramitochondrial carbamoyl phosphate was always below 0.1 mM. Only when ornithine was absent and when, in addition, the concentration of amino acids was higher than four times their plasma concentration, intramitochondrial carbamoyl phosphate rose up to about 3 mM; under these conditions ammonia accumulated in the medium. The relationship between ornithine-cycle flux and the concentration of the cycle intermediates at varying amino acid concentration indicated that under near-physiological conditions the ornithine-cycle enzymes are far from being saturated with their subsidiaries. Moderate concentrations of norvaline had no effect on the rate of urea synthesis unless the cells were severely depleted of ornithine. Activation of carbamoyl-phosphate synthetase (ammonia) by addition of N-carbamoylglutamate only slightly stimulated urea production at all amino acid concentrations. However, in the presence of the activator the curve relating ornithine-cycle flux to the steady-state ammonia concentration was shifted to lower concentrations of ammonia. The intramitochondrial concentration of carbamoyl phosphate in rat liver in vivo was below 0.1 mM. This value is far below the concentration required for substantial inhibition of carbamoyl-phosphate synthetase. It is concluded that in vivo the function of activity changes in carbamoyl-phosphate synthetase, via the well-documented alterations in the intramitochondrial concentration of N-acetylglutamate, is to buffer the intrahepatic ammonia concentration rather than to affect urea production per se. At constant concentration of ammonia the rate of urea production is entirely controlled by the activity of carbamoyl-phosphate synthetase.
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PMID:Control of ureogenesis. 397 93


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