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

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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)

The reversed-phase high-performance liquid chromatography of three synthetic opioid peptides, 5leucine-enkephalin, 5methionine-enkephalin and [D-2alanine]-5methionine enkephalin, has been studied after their pre-column fluorogenic derivatization with naphthalene-2,3-dicarboxaldehyde in the presence of cyanide to the corresponding 1-cyanobenz[f]isoindole (CBI) derivatives. The chromatographic properties of the three synthetic CBI-peptides were characterized using three different stationary phases, ODS Hypersil, CPS Hypersil and Spherisorb Phenyl, eluted with mobile phases containing various concentrations of methanol, tetrahydrofuran or acetonitrile in 26 mM trifluoroacetic acid, adjusted to pH 3.5. The data obtained using single chromatographic columns were used to design a multi-dimensional system in which the three synthetic CBI-peptides of interest were transferred as a single fraction from one column to a second. The first column served to separate the peptides from the majority of the material in the samples, and the second column was used to separate the three CBI-peptides from each other. The best separation was achieved in which the first column was Spherisorb Phenyl and the second column was ODS Hypersil. Both columns were eluted with a mobile phase of 45% acetonitrile (v/v) in 26 mM trifluoroacetic acid (pH 3.5) at a flow-rate of 1.0 ml/min. The method has been applied to the determination of leucine- and methionine-enkephalin-like fluorescence in the striatum of the rat brain.
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PMID:Multi-dimensional high-performance liquid chromatography of opioid peptides following pre-column derivatization with naphthalene-2,3-dicarboxaldehyde in the presence of cyanide ion. Preliminary results on the determination of leucine- and methionine-enkephalin-like fluorescence in the striatum region of the rat brain. 259 17

RNA dot-blot, quantitative electron microscope immunocytochemistry, and electrophoretic immunoblotting techniques were employed to investigate the expression of carbamoyl-phosphate synthetase I (CPS) and ornithine carbamoyl transferase (OCT) genes in rat liver and intestinal mucosa. Comparing only those cell types in the two tissues which express these enzymes, we show that the concentration of CPS and OCT in hepatocyte mitochondria is 2.3-times and 1.2-times greater, respectively, than in intestinal epithelial cell mitochondria. As a percentage of total tissue protein, however, liver homogenates contain 10-20 times more CPS and 5-10 times more OCT than is found in intestinal mucosa. These relatively large differences in enzyme protein levels between the two tissues are not reflected by differences in their mRNA levels. As a percentage of total translational activity in vitro (based on incorporation of [35S]methionine), total liver mRNA directed synthesis of about twice as much precursor CPS (pCPS) and precursor OCT (pOCT) than did equivalent amounts of mRNA from intestinal mucosa. The ratio of pCPS and pOCT mRNA levels between the two tissues (2:1, liver:intestinal mucosa) was confirmed by dot-blot and Northern hybridizations employing specific cDNA probes. The sizes of the respective mRNAs were the same for the two tissues: about 6000 residues for pCPS mRNA and about 1700 residues for pOCT mRNA.
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PMID:Expression of nuclear genes encoding the urea cycle enzymes, carbamoyl-phosphate synthetase I and ornithine carbamoyl transferase, in rat liver and intestinal mucosa. 286 32

Acetyl-coenzyme A carboxylase (Ac-CoA carboxylase; EC 6.4.1.2) catalyzes the rate-limiting reaction in long-chain fatty acid biosynthesis. To investigate the mechanism of genetic control of expression of Ac-CoA carboxylase and the relationship between its structure and function, cDNA clones for Ac-CoA carboxylase were isolated. The complete coding sequence contains 7035 bases; it encodes a polypeptide chain of 2345 amino acids having a Mr of 265,220. The sequences of several CNBr peptides of Ac-CoA carboxylase were localized within the predicted protein sequence as were those peptides that contain the sites for phosphorylation. The deduced protein contains one putative site for biotinylation in the NH2-terminal half. The "conserved" biotinylation site peptide, Met-Lys-Met, is preceded by valine, whereas alanine is found in a similar position in all other known biotin-containing proteins. The primary sequences of Ac-CoA carboxylase and carbamoyl phosphate synthetase exhibit substantial identity.
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PMID:Structure of the coding sequence and primary amino acid sequence of acetyl-coenzyme A carboxylase. 290 Oct 88

Regulation of carbamoyl-phosphate synthetase I (CPS) synthesis by various hormones was compared in primary cultured hepatocytes from adult rat and in Reuber hepatoma H-35 by pulse labeling of the cells with [35S]methionine. CPS synthesis in hepatocytes was stimulated 8-fold and 5-fold by dexamethasone and glucagon respectively. CPS synthesis in hepatocytes was synergically (about 50-fold) stimulated by a combination of dexamethasone and glucagon. Less synergic stimulation was observed by combining dexamethasone with N6, O2'-dibutyryladenosine 3',5'-monophosphate (dibutyryl-cAMP) or with isoproterenol. The basal level of CPS synthesis in hepatoma cells was higher than that in hepatocytes. CPS synthesis in hepatoma cells was stimulated by dexamethasone and dibutyryl-cAMP but the extent was only 3-fold and 1.8-fold respectively. The synergic effect of combination of dexamethasone and dibutyryl-cAMP was not observed in hepatoma cells. Neither glucagon nor isoproterenol exhibited an appreciable effect on CPS synthesis in hepatoma cells. Insulin and epinephrine suppressed CPS synthesis both in hepatocytes and hepatoma cells. The effect of epinephrine was indicated to be through alpha-adrenergic receptors. The effects of insulin and epinephrine were additive on CPS synthesis both in hepatocytes and hepatoma cells.
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PMID:Hormonal regulation of carbamoyl-phosphate synthetase I synthesis in primary cultured hepatocytes and Reuber hepatoma H-35. Defective regulation in hepatoma cells. 304 Mar 99

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

Regulation of synthesis of carbamoyl-phosphate synthetase I by glucocorticoids, 8-bromoadenosine 3',5'-monophosphate (8-bromo-cAMP), and insulin was investigated in Reuber hepatoma H-35. By measuring the incorporation of [35S]methionine into carbamoyl-phosphate synthetase I and its precursor, we showed that dexamethasone stimulates the enzyme synthesis approximately fivefold. A detectable stimulation was observed at 1 nM of dexamethasone, half-maximal stimulation at 4 nM, and maximal stimulation above 40 nM. Corticosterone was more effective than dexamethasone both for the minimal concentration needed and for the extent of the stimulation. Hydrocortisone was less effective than dexamethasone. 8-Bromo-cAMP also stimulated the enzyme synthesis at a concentration of 3 mM. The effect of 8-bromo-cAMP was suggested to be additive to the effect of dexamethasone. Physiological concentrations of insulin strongly suppressed the stimulatory effect of dexamethasone on the enzyme synthesis but could not completely counteract the effect of dexamethasone. The half-maximal and maximal effects of insulin were observed at 0.5 nM and 5 nM, respectively. Insulin also counteracted the effect of 8-bromo-cAMP on the enzyme synthesis.
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PMID:Interaction between glucocorticoids, 8-bromoadenosine 3',5'-monophosphate, and insulin in regulation of synthesis of carbamoyl-phosphate synthetase I in Reuber hepatoma H-35. 389 22

In the course of studies on glutamine-dependent carbamyl phosphate synthetase from Aerobacter aerogenes, we purified another protein which was found to be glutamate synthase (EC 2.6.1.53). The enzyme, obtained in apparently homogeneous form (monomer molecular weight about 227,000; s(20,omega) = 17.6 S), was found to be a typical glutamine amidotransferase in that it exhibits glutaminase activity and can utilize ammonia in place of glutamine as a nitrogen donor. The enzyme also catalyzes at low rates the oxidative deamination of glutamate in the presence of TPN, and it exhibits TPNH oxidase activity. The enzyme is similar to the glutamate synthase found in Escherichia coli in that it is an iron-sulfide flavoprotein. Treatment of the enzyme with sodium dodecyl sulfate or potassium thiocyanate dissociates it into nonidentical subunits exhibiting molecular weights of about 175,000 and 51,500. The glutamine-dependent activity of the enzyme is inhibited by L-2-amino-4-oxo-5-chloropentanoic acid, but this chloroketone analog of glutamine does not affect the ammonia-dependent glutamate synthase activity. Studies with [(14)C]chloroketone show that the reagent binds to the heavy subunit only. Inhibition by the chloroketone and its binding to the heavy subunit are markedly reduced in the presence of L-glutamine. Sedimentation velocity studies carried out in potassium thiocyanate indicate that iron-sulfide and flavin sites are also located on the heavy subunit. While these studies show that glutamate synthase resembles other glutamine amidotransferases in certain of its catalytic properties, the findings indicate that the light subunit of this enzyme, in contrast to that of several other glutamine amidotransferases, does not function to bind glutamine. It is of interest that the enzyme exhibits an unusually high affinity for ammonia as compared to a number of other glutamine amidotransferases. Glutamate synthase is inhibited (competitively with respect to glutamine) by low concentrations of methionine sulfone, methionine sulfoximine, and methionine sulfoxide.
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PMID:Glutamine-binding subunit of glutamate synthase and partial reactions catalyzed by this glutamine amidotransferase. 453 Oct 4

Reuber hepatoma H-35 was found to retain the activity of carbamoyl-phosphate synthetase I. The content of this enzyme in H-35 grown in Eagle's minimal essential medium was about half that in rat liver. The enzyme from H-35 was the same as that from rat liver in molecular weight estimated by SDS-polyacrylamide gel electrophoresis, specific enzyme activity, kinetic parameters for ATP and N-acetyl-L-glutamate, and immunological crossreactivity. The enzyme in H-35 was induced by dexamethasone (1.4-fold) but not by glucagon or dibutyryl cAMP. Incorporation of [35S] methionine into the enzyme indicated that the effect of dexamethasone was due to increased synthesis of the enzyme protein (2.1-fold). By labeling with [35S]methionine, the precursor and the mature forms of carbamoyl-phosphate synthetase I were observed in the post-mitochondrial and mitochondrial fractions, respectively. By chasing the labeled cells with unlabeled methionine and cycloheximide, it was observed that the rate of translocation of the precursor into mitochondria is not affected by dexamethasone.
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PMID:Induction of carbamoyl-phosphate synthetase I in Reuber hepatoma H-35 by dexamethasone. 630 26

The regulatory mechanism of the developmental increase of carbamoyl-phosphate synthetase I in fetal and neonatal rat liver was studied in vivo. The appearance and rapid increase of the enzyme in late fetal period were caused by de novo synthesis of the enzyme protein. The amount of the enzyme protein analyzed by SDS-polyacrylamide gel electrophoresis was proportional to the enzyme activity throughout the period of development. No indication was observed for preexisting protein which could be converted into the active protein. A novel system for the in vivo study of carbamoyl-phosphate synthetase I synthesis was developed. Hepatocytes, mechanically dispersed by repeated passage of the tissue through a pipet, incorporated [35S]methionine into the enzyme. Taking advantage of this system, the regulation of the enzyme synthesis was studied. In vivo synthesis of the enzyme was detected at 4 days before birth and rapidly increased until 1 day before birth. However, the enzyme synthesis was markedly repressed after birth, when the amount of carmamoyl-phosphate synthetase I itself reached the adult level. This result was in a clear contrast with the constant level of the translatable mRNA (Raymond, Y. and Shore, G.C. (1981) Biochim. Biophys. Acta 656, 111-119) and suggested that post-transcriptional regulation is important in addition to the level of mRNA for the regulation of the carbamoyl-phosphate synthetase I level.
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PMID:In vivo study of developmental changes in carbamoyl-phosphate synthetase I in rat liver. Repression of the enzyme synthesis immediately after birth. 683 Aug 3

The primary nitrogen metabolism of the N2-fixing root nodule symbiosis Alnus incana (L.)- Frankia was investigated by 31P and 15N nuclear magnetic resonance (NMR) spectroscopy. Perfusion of root nodules in a pulse-chase approach with 15N- or 14N-labeled NH4+ revealed the presence of the amino acids alanine (Ala), gamma-amino butyric acid, glutamine (Gln), glutamic acid (Glu), citrulline (Cit) and arginine (Arg). Labeling kinetics of the Gln amide-N and alpha-amino acids suggested that the glutamine synthetase (GS; EC 6.3.1.2)-glutamate synthase (GOGAT; EC 1.4.1.13) pathway was active. Inhibition of the GS-catalyzed reaction by methionine sulphoximine abolished incorporation of 15N. Cit was labeled in all three N positions but most rapidly in the omega position, consistent with carbamoyl phosphate as the precursor to which Gln could be the amino donor catalyzed by carbamoyl phosphate synthase (CPS; EC 6.3.5.5). Ala biosynthesis occurred consistent with a flux of N in the sequence Gln-Glu-Ala. 31P NMR spectroscopy in vivo and of extracts revealed several metabolites and was used in connection with the 15N pulse-chase experiment to assess general metabolic status. Stable concentrations of ATP and UDP-glucose during extended perfusions showed that the overall root nodule metabolism appeared undisturbed throughout the experiments. The metabolic pathways suggested by the NMR results were confirmed by high activities of the enzymes GS, NADH-GOGAT and ornithine carbamoyltransferase (OCT; EC 2.1.3.3). We conclude that the primary pathway of NH4+ assimilation in A. incana root nodules occurs through the GS-GOGAT pathway. Biosynthesis of Cit through GS-CPS-OCT is important and is a link between the first amino acid Gln and this final transport and storage form of nitrogen.
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PMID:Primary metabolism in N2-fixing Alnus incana-Frankia symbiotic root nodules studied with 15N and 31P nuclear magnetic resonance spectroscopy. 1517 12


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