scholarly journals Citrulline synthesis in rat tissues and liver content of carbamoyl phosphate and ornithine

1974 ◽  
Vol 138 (2) ◽  
pp. 225-232 ◽  
Author(s):  
Luisa Raijman
Keyword(s):  
Rat Liver ◽  
Soluble Fraction ◽  
Maximal Activity ◽  
Rat Tissues ◽  
Ornithine Carbamoyltransferase ◽  
Carbamoyl Phosphate Synthetase ◽  
Carbamoyl Phosphate ◽  
Citrulline Synthesis

Rat liver ornithine carbamoyltransferase appears to be located exclusively in the mitochondria; the activity that is found in the soluble fraction is indistinguishable from mitochondrial ornithine carbamoyltransferase by simple kinetic criteria, and seems to result from breakage of mitochondria during homogenization. Of several rat tissues studied, only the liver and the mucosa of small intestine contain significant amounts of ornithine carbamoyltransferase; the activity in intestinal mucosa is less than one thousandth of that in liver. Qualitatively, this distribution coincides with that of carbamoyl phosphate synthetase I and its cofactor, acetylglutamate. The rat liver contents of carbamoyl phosphate and ornithine were 0.1 and 0.15μmol/g wet wt. of tissue respectively. On the basis of these values, it is proposed that in vivo the ornithine carbamoyltransferase activity of liver may be much lower than its maximal activity in vitro might suggest.

scholarly journals Metabolic Enzymes from Psychrophilic Bacteria: Challenge of Adaptation to Low Temperatures in Ornithine Carbamoyltransferase from Moritellaabyssi

2003 ◽  
Vol 185 (7) ◽  
pp. 2161-2168 ◽  
Author(s):  
Ying Xu ◽  
Georges Feller ◽  
Charles Gerday ◽  
Nicolas Glansdorff
Keyword(s):  
Low Temperatures ◽  
Bacterial Species ◽  
Stable State ◽  
Maximal Activity ◽  
Metabolic Enzymes ◽  
Lower Sensitivity ◽  
Ornithine Carbamoyltransferase ◽  
Carbamoyl Phosphate ◽  
Enthalpy Changes

ABSTRACT The enzyme ornithine carbamoyltransferase (OTCase) of Moritella abyssi (OTCaseMab), a new, strictly psychrophilic and piezophilic bacterial species, was purified. OTCaseMab displays maximal activity at rather low temperatures (23 to 25°C) compared to other cold-active enzymes and is much less thermoresistant than its homologues from Escherichia coli or thermophilic procaryotes. In vitro the enzyme is in equilibrium between a trimeric state and a dodecameric, more stable state. The melting point and denaturation enthalpy changes for the two forms are considerably lower than the corresponding values for the dodecameric Pyrococcus furiosus OTCase and for a thermolabile trimeric mutant thereof. OTCaseMab displays higher Km values for ornithine and carbamoyl phosphate than mesophilic and thermophilic OTCases and is only weakly inhibited by the bisubstrate analogue δ-N-phosphonoacetyl-l-ornithine (PALO). OTCaseMab differs from other, nonpsychrophilic OTCases by substitutions in the most conserved motifs, which probably contribute to the comparatively high Km values and the lower sensitivity to PALO. The Km for ornithine, however, is substantially lower at low temperatures. A survey of the catalytic efficiencies (k cat/Km ) of OTCases adapted to different temperatures showed that OTCaseMab activity remains suboptimal at low temperature despite the 4.5-fold decrease in the Km value for ornithine observed when the temperature is brought from 20 to 5°C. OTCaseMab adaptation to cold indicates a trade-off between affinity and catalytic velocity, suggesting that optimization of key metabolic enzymes at low temperatures may be constrained by natural limits.

scholarly journals Altered enzyme activities and citrulline synthesis in liver mitochondria from ornithine carbamoyltransferase-deficient sparse-furash mice

1989 ◽  
Vol 257 (1) ◽  
pp. 251-257 ◽  
Author(s):  
N S Cohen ◽  
C W Cheung ◽  
L Raijman
Keyword(s):  
Protein Interactions ◽  
Mitochondrial Protein ◽  
Normal Liver ◽  
Liver Mitochondria ◽  
Synthetase Activity ◽  
Single Mutation ◽  
Ornithine Carbamoyltransferase ◽  
Carbamoyl Phosphate Synthetase ◽  
Carbamoyl Phosphate ◽  
Citrulline Synthesis

Male mice carrying the spfash mutation have 5-10% of the normal activity of ornithine carbamoyltransferase, yet are only slightly hyperammonaemic and develop quite well. A study of liver mitochondria from normal and spfash males showed that they differ in important ways. (1) The spfash liver contains about 33% more mitochondrial protein per g than does normal liver. (2) The specific activities of carbamoyl-phosphate synthetase (ammonia) and glutamate dehydrogenase are about 15% lower than normal in mitochondria from spfash mice, whereas those of beta-hydroxybutyrate dehydrogenase and cytochrome oxidase are 22% higher and 30% lower respectively. (3) In the presence of 10 mM-ornithine and the substrates for carbamoyl phosphate synthesis, coupled and uncoupled mitochondria from spfash mice synthesize citrulline at unexpectedly high rates, about 25 and 44 nmol/min per mg respectively. Though these are somewhat lower than the corresponding rates obtained with normal mitochondria, the difference does not arise from the deficiency in ornithine carbamoyltransferase, but from the lower carbamoyl-phosphate synthetase activity of the mutant mitochondria. (4) At lower external [ornithine] (less than 2 mM), a smaller fraction of the carbamoyl phosphate synthesized is converted into citrulline in spfash than in normal mitochondria. These studies show that what appears to be a single mutation brings about major adaptations in the mitochondrial component of liver. In addition, they clarify the role of ornithine transport and of protein-protein interactions in citrulline synthesis in normal mitochondria.

scholarly journals Regulation of urea and citrulline synthesis under physiological conditions

1993 ◽  
Vol 292 (1) ◽  
pp. 241-247 ◽  
Author(s):  
G Beliveau Carey ◽  
C W Cheung ◽  
N S Cohen ◽  
S Brusilow ◽  
L Raijman
Keyword(s):  
Urea Synthesis ◽  
Substrate Availability ◽  
Carbamoyl Phosphate ◽  
Short Term ◽  
Two Factors ◽  
Moment Control ◽  
Citrulline Synthesis ◽  
The Relationship

Information on the regulation of urea synthesis in vivo was obtained by examining the relationship between ureagenesis in vivo, citrulline synthesis in vitro, and two factors currently hypothesized to exert short-term regulation of this pathway: the liver mitochondrial content of N-acetylglutamate (NAG) and substrate availability. Rats meal-fed for 4 h every day (4-20 schedule) or for 8 h every other day (8-40 schedule) were used. (1) The citrulline-synthesizing capacity of mitochondria from livers of rats on the 8-40 schedule exceeded the corresponding velocity of urea synthesis in vivo at all time points studied. (2) Mitochondrial NAG in these livers increased from 127 +/- 32 pmol/mg of protein at 0 h to 486 +/- 205 pmol/mg at 3 h after the start of a meal, and decreased thereafter, but the correlation between NAG content and the velocity of citrulline synthesis was not simple, suggesting that NAG is not the only determinant of the state of activation of carbamoyl phosphate synthase I. (3) In rats on the 4-20 schedule killed 1 h after the start of the meal, the liver content of ornithine, citrulline, arginine, glutamate, alanine and urea increased 2.1-12-fold with respect to the values at 0 h; glutamine decreased by 39%. (4) The combined findings indicate that in vivo, moment-to-moment control of the velocity of urea synthesis is exerted by substrate availability. (5) Digestion limits the supply of substrate to the liver, and prevents its ureagenic capacity from being overwhelmed following a protein-containing meal.

scholarly journals Organization of a multifunctional protein in pyrimidine biosynthesis. A domain hypersensitive to proteolysis

1984 ◽  
Vol 217 (2) ◽  
pp. 435-440 ◽  
Author(s):  
P C Rumsby ◽  
P C Campbell ◽  
L A Niswander ◽  
J N Davidson
Keyword(s):  
Pyrimidine Biosynthesis ◽  
Carbamoyl Phosphate Synthetase ◽  
Carbamoyl Phosphate ◽  
Multifunctional Protein ◽  
Aggregation Site ◽  
Aspartate Carbamoyltransferase ◽  
Low Concentrations ◽  
Proteinase A

When the multifunctional protein that catalyses the first three steps of pyrimidine biosynthesis in hamster cells is treated with staphylococcal V8 proteinase, a single cleavage takes place. The activities of carbamoyl-phosphate synthetase (EC 6.3.5.5), aspartate carbamoyltransferase (EC 2.1.3.2) and dihydro-orotase (EC 3.5.2.3) and the allosteric inhibition by UTP are unaffected. One fragment, of Mr 182000, has the first and third enzyme activities, whereas the other fragment, of Mr 42000, has aspartate carbamoyltransferase activity and an aggregation site. A similar small fragment is observed in protein digested with low concentrations of trypsin. A similar large fragment is seen after digestion with trypsin and as the predominating form of this protein in certain mutants defective in pyrimidine biosynthesis. These results indicate that a region located adjacent to the aspartate carbamoyltransferase domain is hypersensitive to proteinase action in vitro and may also be sensitive to proteolysis in vivo.

scholarly journals Effect of cortisol on the thiol content of rat liver nuclear proteins

1972 ◽  
Vol 126 (5) ◽  
pp. 1171-1179 ◽  
Author(s):  
D. Doenecke ◽  
M. Beato ◽  
L. F. Congote ◽  
C. E. Sekeris
Keyword(s):  
Rat Liver ◽  
Rna Synthesis ◽  
Soluble Fraction ◽  
Polyacrylamide Gel Electrophoresis ◽  
Nuclear Proteins ◽  
Carboxymethyl Lysine ◽  
Thiol Content ◽  
Liver Nuclei

Administration of cortisol to normal or adrenalectomized rats leads within 15–30min to an increased thiol content of nuclear proteins, measured by the incorporation of iodo[3H]-acetate or N-[14C]ethylmaleimide or by colorimetric methods. The same effect is observed after incubation of isolated rat liver nuclei with corticosteroids. The increased thiol content of the nuclear proteins shows the same time-dependence as the stimulation of RNA synthesis by corticosteroids observed in vivo and in vitro. Amino acid analysis of the carboxymethylated proteins reveals that in the experiments in vivo most of the label is present as carboxymethylcysteine with small amounts of carboxymethyl-lysine and carboxymethylhistidine, whereas in the experiments in vitro more carboxymethyl-lysine and carboxymethylhistidine than carboxymethylcysteine are found. The increase in the content of thiol groups is due to cleavage of the disulphide bridges between the nuclear proteins. Polyacrylamide-gel electrophoresis of the acid-soluble fraction reveals that most of the iodo[3H]acetate label is incorporated into a non-histone fraction with a molecular weight of approx. 45000 whereas in the acid-insoluble fractions many protein bands are labelled.

Effect of X-irradiation on gene expression of rat liver chemokines: in vivo and in vitro studies

2008 ◽  
Vol 46 (01) ◽  
Author(s):  
F Moriconi ◽  
H Christiansen ◽  
H Christiansen ◽  
N Sheikh ◽  
J Dudas ◽  
...  
Keyword(s):  
Gene Expression ◽  
Rat Liver ◽  
In Vitro Studies ◽  
X Irradiation
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