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)

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.
...
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

Some properties of carbamoyl-phosphate synthetase (ammonia) were studied in rat-liver mitochondria made selectively permeable by pretreatment with toluene. The Michaelis constants for NH3, MgATP and HCO-3 were 0.7, 1.2 and 2 mM respectively. N-Acetylglutamate activated the enzyme with a Ka of about 0.1 mM. At saturating concentrations of substrates and effectors the enzyme was inhibited by 50% by carbamoyl phosphate at a concentration of 13 mM. Binding of N-acetylglutamate to carbamoyl-phosphate synthetase required the presence of both free Mg2+ ions and MgATP, and was inhibited by Ca2+ ions and by N-carbamoylglutamate. The known activation of carbamoyl-phosphate synthetase by free Mg2+ is due to an increased affinity of the enzyme for N-acetylglutamate. Binding of N-acetylglutamate to carbamoyl-phosphate synthetase was a slow process: at N-acetylglutamate concentrations below 0.5 mM maximal binding was not completed within 30 min. The rate of binding increased with increasing N-acetylglutamate concentrations. Dissociation of N-acetylglutamate from the enzyme was relatively fast, with a half-time of about 5 min. Under all conditions studied there was a close relationship between carbamoyl-phosphate synthetase activity and the amount of N-acetylglutamate bound to the enzyme. The data are discussed in relation to the control of carbamoyl-phosphate synthetase in the intact hepatocyte.
...
PMID:Properties of carbamoyl-phosphate synthetase (ammonia) in rat-liver mitochondria made permeable with toluene. 688 64

The binding of N-acetyl-L-glutamate, the physiological allosteric activator, to rat liver carbamoyl-phosphate synthetase (ammonia) was studied by techniques of rate of dialysis and of ultracentrifugation in the Airfuge. There is one binding site for acetylglutamate per enzyme monomer (Mr 165 000). K+, Mg2+ (free) and ATP were required to demonstrate binding. The concentrations of ATP required indicate that binding of ATPA (the ATP molecule that yields Pi) is needed. HCO-3 was not essential, but it enhanced binding of acetylglutamate. Glycerol also favored binding. Plots of Kd values versus the reciprocal of free Mg2+ and ATP concentrations are linear and indicate that ATPA, K+ and Mg2+ bind before acetylglutamate. In the presence of these ligands and HCO-3, ammonia increased drastically the Kd value for acetylglutamate, whereas in absence of HCO-3 ammonia had little effect. This suggests that acetylglutamate dissociates with the products and explains the higher Km for acetylglutamate in the synthetase (overall) reaction than in the ATPase (partial) reaction. In the absence of ATP acetylglutamate was bound with high affinity if ADP and carbamoyl phosphate were present. ADP or carbamoyl phosphate alone did not promote substantial binding. Binding of acetylglutamate at low concentration was slow; it was accelerated at higher concentrations of the activator. Exchange of bound acetylglutamate with acetylglutamate in solution was fast. A scheme proposed earlier for allosteric activation of the enzyme [Rubio, V., Britton, H. G. and Grisolia, S. (1983) Eur. J. Biochem. (in preparation)] is refined to incorporate the new information. Binding of ATPA, K+ and Mg2+ and formation of 'active CO2' (the central complex) are greatly favored by acetylglutamate.
...
PMID:Binding of N-acetyl-L-glutamate to rat liver carbamoyl phosphate synthetase (ammonia). 688 68

1. When NH3 was added to isolated rat-liver mitochondria incubated with succinate and bicarbonate, oxidation of succinate was stimulated to a greater extent than could be accounted for by the net formation of carbamoyl phosphate. 2. Measurement of the rate of incorporation of [14C]bicarbonate into carbamoyl phosphate, after the mitochondria had been preincubated with NH3 and unlabelled bicarbonate, revealed that flux through carbamoyl-phosphate synthetase (ammonia) was much greater than the net formation of carbamoyl phosphate indicated. 3. It is concluded that part of the carbamoyl phosphate produced in the absence of ornithine is degraded. About 20% of the degradation can be accounted for by non-enzymatic reactions of carbamoyl phosphate outside the mitochondria. It is proposed that the remainder of the degradation of carbamoyl phosphate occurs by partial reversal of the reaction of carbamoyl-phosphate synthetase.
...
PMID:Activity of carbamoyl-phosphate synthetase (ammonia) in isolated rat-liver mitochondria: cycling of carbamoyl phosphate in the absence of ornithine. 708 32

All the five enzymes of urea synthesis and the formation of urea in vitro can already be demonstrated in human liver as early as the 9th week of fetal development. At this stage the activity of carbamoyl phosphate synthetase is the highest, whereas that of ornithine carbamoyltransferase is the lowest as compared to those in the adult. The kinetic parameters of the urea cycle enzymes are the same in fetal liver as in adult liver, except that the Km values of ornithine carbamoyltransferase for L-ornithine are 3.5 mM and 0.42 mM in the fetus and in adult liver, respectively. Urea formation in vivo seems to begin in the second half of fetal life, and a gradual increase can be detected in the activity of the enzymes of urea synthesis. The activity of ornithine decarboxylase, the glutamine-dependent carbamoyl phosphate synthetase and aspartate carbamoyltransferase, however, changes in the opposite direction. The concentration of carbamoyl phosphate and aspartate remains constant, but that of ornithine gradually decreases during ontogenesis. The ornithine, carbamoylphosphate and aspartate pools are probably utilized in the polyamine, pyrimidine and urea syntheses at varying rates.
...
PMID:Urea cycle enzymes in human liver: ontogenesis and interaction with the synthesis of pyrimidines and polyamines. 708 58

The inhibition of aspartate carbamoyltransferase (ACTase) from rat Novikoff tumor by N-(phosphonacetyl)-L-aspartate (PALA) was studied in a substrate mixture permitting endogenous synthesis of carbamoyl phosphate. Among the components required for carbamoyl phosphate synthetase activity, ATP, Mg(C2H3O2)2 and KCl interfered with inhibition by PALA (with added carbamoyl phosphate). The inhibition was also decreased when the concentration of partially purified enzyme was increased. In the system dependent on carbamoyl phosphate synthetase, the 50% inhibitory concentration of PALA was lower than that in the same mixture plus 0.2 mM carbamoyl phosphate, but higher than in the usual simple assay mixture with 0.2 mM carbamoyl phosphate.
...
PMID:N-(Phosphonacetyl)-L-aspartate inhibition of the enzyme complex of pyrimidine biosynthesis. 715 Mar 57

The purification of mitochondrial carbamoyl phosphate synthetase I (carbon-dioxide: ammonia ligase (ADP-forming, carbamate-phosphorylating), EC 6.3.4.16) from small samples of human liver is described. The enzyme is composed of a single polypeptide of Mr 160 000 +/- 500 as shown by SDS-polyacrylamide gel electrophoresis in the presence of reducing agents. The synthetase migrates in polyacrylamide gradient gels in the absence of detergents at a rate corresponding to a Mr of 165 000. Estimates of the molecular weight of the native enzyme by gel filtration and density gradient centrifugation yield a value of 178 000. The results indicate that the enzyme exists predominantly as monomeres. Amino acids composition, isoelectric point, stability, Km values and the ability to catalyze partial reactions have been measured and compared with known properties of carbamoyl phosphate synthetases from other sources. From the available data a high degree of evolutionary conservation of the ammonia-dependent synthetase is suggested. This is also supported by the demonstration of extensive immunological cross-reactivity between the human and rat enzymes.
...
PMID:Carbamoyl phosphate synthetase I of human liver. Purification, some properties and immunological cross-reactivity with the rat liver enzyme. 724 16

Addition of ammonium ions to isolated rat hepatocytes stimulated the rate of synthesis of pyrimidines. Isolation and quantification of pyrimidine nucleotides orotic acid and the acid-hydrolyzed product of carbamoyl-aspartic acid by ion-exchange chromatography and high-pressure liquid chromatography show a marked stimulation in the incorporation of [14C]bicarbonate in incubations with added ammonium ions. The incorporation into total uridine nucleotides (sigma UMP) was increased twofold in the presence of 5 mM ammonium ion, and approximately eightyfold into orotic acid. There was a parallel increase in labelling of carbamoylaspartic acid from undetectable to a level similar to that of orotic acid. The specific activity of urea formed during the incubations did not change during incubations or in the presence of ammonium ions confirming that the change in labelling of pyrimidine was not due to a change in the specific activity of precursor. Despite the stimulation in incorporation of label into pyrimidines there was no increase in the hepatocyte content of sigma UMP, which was 11.5 mumol/g dry weight, although the orotic acid content increased from 0.09 mumol/g dry weight in the absence of added ammonium ions (but in the presence of 2 mM L-glutamine) to 8.6 mumol/g dry weight with 5 mM ammonium ion. The stimulated incorporation of [14C]bicarbonate in the presence of 5 mM and 10 mM ammonium ion was shown to be due to a stimulated synthesis of carbamoyl phosphate, since greater than 80% of label in the uracil ring was present at position C-2. Incubation of hepatocytes in basal medium (Eagles) containing 2.5% foetal calf serum and 20 mM bicarbonate showed that there was a significant stimulation of pyrimidine synthesis with 1 mM ammonium ion. The stimulatory effect of ammonium ions on incorporation of bicarbonate into pyrimidines was almost completely reversed by 5 mM L-ornithine and was partially reversed by 1 mM L-ornithine. Evidence for a contribution of the urea cycle carbamoyl phosphate synthetase to pyrimidine synthesis is discussed.
...
PMID:Effect of ammonium ion on pyrimidine synthesis de novo in isolated rat hepatocytes. 725 Jan 18

The L-glutamine- and N-acetyl-L-glutamate-dependent carbamoyl phosphate synthetase III present in liver of spiny dogfish (Squalus acanthias) has been purified to a high state of purity. The purified enzyme has a Mr congruent to 160,000 and is subject to self-association which is facilitated by the presence of MgATP, L-glutamine, and N-acetyl-L-glutamate. The enzyme exhibits hysteretic properties. The time course of the reaction is characterized by a lag or a burst in activity, depending upon preincubation conditions, which can last up to 30 min. The lag period can be eliminated by preincubating the enzyme at 26 degrees C (but not at 4 degrees C) in the presence of the above three ligands. Mg2+ in excess of that required to complex ATP as MgATP and N-acetyl-L-glutamate are both required for full activity. The requirement of K+ for activity can be replaced by NH4+, but not by Na+. Ammonia can act as a substrate in place of L-glutamine, but the maximal rate is much less than that which can be obtained with L-glutamine. The glutamine-dependent activity is inhibited by ammonia. Apparent Km values under optimal conditions for N-acetyl-L-glutamate, L-glutamine, MgATP, bicarbonate, and ammonia are 0.013 mM, 0.16 mM, 0.35 mM, 1.7 mM, and 2 mM, respectively. The apparent Km for N-acetyl-L-glutamate decreases when the concentration of L-glutamine increases, and vice versa. The apparent Km values for these two ligands are increased when urea is present at normal physiological concentrations (0.4 M), and the activity of the enzyme is significantly affected by changes in urea concentration. Compounds known to act as allosteric effectors on other glutamine-dependent carbamoyl phosphate synthetases had little or no effect on this enzyme. The properties of the enzyme are consistent with the view that the function of this carbamoyl phosphate synthetase III is related to the synthesis of urea which is retained in these species as a mechanism for osmoregulation.
...
PMID:Purification and properties of the glutamine- and N-acetyl-L-glutamate-dependent carbamoyl phosphate synthetase from liver of Squalus acanthias. 729 55

Carbamoyl phosphate synthetase from Escherichia coli catalyzes the synthesis of carbamoyl phosphate from bicarbonate, ammonia, and two molecules of MgATP. The enzyme is composed of two nonidentical subunits. The small subunit catalyzes the hydrolysis of glutamine to glutamate and ammonia. The large subunit catalyzes the formation of carbamoyl phosphate and has the binding sites for bicarbonate, ammonia, MgATP, and the allosteric ligands IMP, UMP, and ornithine. The allosteric ligands are believed to bind to the extreme C-terminal portion of the large subunit. Truncation mutants were constructed to investigate the allosteric binding domain. Stop codons were introduced at various locations along the carB gene in order to delete amino acids from the carboxy-terminal end of the large subunit. Removal of 14-119 amino acids from the carboxy-terminal end of the large subunit resulted in significant decreases in all of the enzymatic activities catalyzed by the enzyme. A 40-fold decrease in the glutamine-dependent ATPase activity was observed for the delta 14 truncation. Similar losses in activity were also observed for the delta 50, delta 65, delta 91, and delta 119 mutant proteins. However, formation of carbamoyl phosphate was detected even after the deletion of 119 amino acids from the carboxy-terminal end of the large subunit. No allosteric effects were observed for UMP with either the delta 91 or delta 119 truncation mutants, but alterations in the catalytic activity were observed in the presence of ornithine even after the removal of the last 119 amino acids from the large subunit of CPS. Six conserved amino acids within the allosteric domain were mutated.(ABSTRACT TRUNCATED AT 250 WORDS)
...
PMID:Regulatory changes in the control of carbamoyl phosphate synthetase induced by truncation and mutagenesis of the allosteric binding domain. 757 87


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

---