Urea cycle activities

Metabolically, acetyl CoA that is generated is diverted to ketogenesis, and urea cycle activity is decreased, leading to hyperammonaemia associated with fasting hypoglycaemia, increased lactataemia associated with an increased lactate pyruvate ratio, and increased ketonaemia with ratio < 1. Data from a patient affected with a PC defect are presented in Table 1.4.12. 

These changes in demand for urea cycle activity are met over the long term by regulation of the rates of synthesis of the four urea cycle enzymes and carbamoyl phosphate synthetase I in the liver. All five enzymes are synthesized at higher rates in starving animals and in animals on veiy-high-protein diets than in well-fed animals eating primarily carbohydrates and fats. Animals on protein-free diets produce lower levels of urea cycle enzymes. 

Regulation of carbamoyl-phosphate synthase (ammonia) in liver in relation to urea cycle activity, A. J. Meijer,... 

Condensation of CO2, ammonia, and ATP to form carbamoyl phosphate is catalyzed by mitochondrial carbamoyl phosphate synthase I (reaction 1, Figure 29-9). A cytosolic form of this enzyme, carbamoyl phosphate synthase II, uses glutamine rather than ammonia as the nitrogen donor and functions in pyrimidine biosynthesis (see Chapter 34). Carbamoyl phosphate synthase I, the rate-hmiting enzyme of the urea cycle, is active only in the presence of its allosteric activator JV-acetylglutamate, which enhances the affinity of the synthase for ATP. Formation of carbamoyl phosphate requires 2 mol of ATP, one of which serves as a phosphate donor. Conversion of the second ATP to AMP and pyrophosphate, coupled to the hydrolysis of pyrophosphate to orthophosphate, provides the driving... 

The activity of carbamoyl phosphate synthase I is determined by A -acetylglutamate, whose steady-state level is dictated by its rate of synthesis from acetyl-CoA and glutamate and its rate of hydrolysis to acetate and glutamate. These reactions are catalyzed by A -acetylglu-tamate synthase and A -acetylglutamate hydrolase, respectively. Major changes in diet can increase the concentrations of individual urea cycle enzymes 10-fold to 20-fold. Starvation, for example, elevates enzyme levels, presumably to cope with the increased production... 

Gene therapy for rectification of defects in the enzymes of the urea cycle is an area of active investigation. Encouraging preliminary results have been obtained, for example, in animal models using an adenoviral vector to treat citrullinemia. 

Ornithine transcarbamylase deficiency. This is the most common of the urea cycle defects. Presentation is variable, ranging from a fulminant, fatal disorder of neonates to a schizophrenic-like illness in an otherwise healthy adult. Males characteristically fare more poorly than do females with this X-linked disorder because of random inactivation (lyonization) of the X chromosome. If inactivation affects primarily the X chromosome bearing the mutant OTC gene, then a more favorable outcome can be anticipated. Conversely, the unfavorably lyonized female has a more active disease. 

We begin this overview of manganese biochemistry with a brief account of its role in the detoxification of free radicals, before considering the function of a dinuclear Mn(II) active site in the important eukaryotic urea cycle enzyme arginase. We then pass in review a few microbial Mn-containing enzymes involved in intermediary metabolism, and conclude with the very exciting recent results on the structure and function of the catalytic manganese cluster involved in the photosynthetic oxidation of water. 

An increase in the protein content of the diet in rats increases the maximnm activities of all the enzymes of the cycle in the liver. It is assnmed that this represents increased amonnts of these enzymes in the liver (Table 10.4). Since a chronic increase in the protein in the diet in hnmans increases urea production over a long period and also a decrease in protein in the diet decreases urea production, it is assnmed that, as in the rat, this is due to changes in the concentrations and therefore activities of urea cycle enzymes. 

Table 10.4 Chronic effects of high and zero protein diets on maximum activities of urea cycle enzymes in the liver of the rat...

Note that total starvation of rats increases the activities of all the urea cycle enzymes. 

Ornithine is a metabolically quite active amino acid, and the important precursor of pyrrolidine nucleus, which is found in pyrrolizidine alkaloids. Ornithine itself is a non-protein amino acid formed mainly from L-glumate in plants, and synthesized from the urea cycle in animals as a result of the reaction catalyzed by enzymes in arginine. 

Allosteric activation of CPS-I by iV-acetylglutamate, which is formed from acetyl CoA and glutamate, and indicates adequate availability of substrates for the urea cycle. 

Contraindications Active hepatic disease, urea cycle disorders... 

The carbamoyl phosphate, which functions as an activated carbamoyl group donor, now enters the urea cycle. The cycle has four enzymatic steps. First, carbamoyl phosphate donates its carbamoyl group to ornithine to form citrulline, with the release of Pj (Fig. 18-10, step ). Ornithine plays a role resembling that of oxaloacetate in the citric acid cycle, accepting material at each turn of the cycle. The reaction is catalyzed by ornithine transcarbamoylase, and the citrulline passes from the mitochondrion to the cytosol. 

As we noted in Chapter 16, the enzymes of many metabolic pathways are clustered (p. 605), with the product of one enzyme reaction being channeled directly to the next enzyme in the pathway. In the urea cycle, the mitochondrial and cytosolic enzymes appear to be clustered in this way. The citrulline transported out of the mitochondrion is not diluted into the general pool of metabolites in the cytosol but is passed directly to the active site of argininosuccinate synthetase. This channeling between enzymes continues for argininosuccinate, arginine, and ornithine. Only urea is released into the general cytosolic pool of metabolites. 

The activity of the urea cycle is regulated at the level of enzyme synthesis and by allosteric regulation of the enzyme that catalyzes the formation of carbamoyl phosphate. 

Transamination and the Urea Cycle Aspartate aminotransferase has the highest activity of all the mammalian liver aminotransferases. Why ... 

Fig. 3. Proposed reaction cycle for urease. For urea, R = —NH2. Step 1 urea is activated toward nucleophilic attack by O coordination to a nickel ion the =N+H2 is stabilized by interaction with a protein carboxylate. Step 2 nucleophilic attack by a hydroxide ion, coordinated to the second nickel, to form a tetrahedral intermediate. Step 3 breakdown of the tetrahedral intermediate to form a coordinated carbamate ion. Step 4 hydrolysis releases carbamate ion, the initial product of urease on urea. Reproduced, with permission, from Ref. 34.

An acceleration of protein turnover by thyroxine also has been shown, implying that the hormone may alter various processes by a specific effect on synthesis of certain key proteins Involved in enzymatic reactions, Thus, not only does thyroxine increase the rate of formation of new protein material, hut it also may be responsible for the transformation of non-en/.ymalically active protein Into protein with enzymatic activity. The hormone has also been shown to be capable of acceleration of the synthesis of urea cycle enzymes and probably is essential for the production of a... 

Carbamoyl phosphate synthetase, which is technically not a member of the urea cycle, catalyzes the condensation and activation of ammonia (from the oxidative deamination of glutamate by glutamate dehydrogenase Topic M2) and C02 (in the form of bicarbonate, HC03 ) to form carbamoyl phosphate. The hydrolysis of two ATP molecules makes this reaction essentially irreversible. 

The entry of activated ammonia into the urea cycle occurs by the ornithine transcarbamoylase reaction where the carbamoyl group is transferred to the side chain amino group of the non-protein amino acid, ornithine. Ornithine has five carbons its carbon chain therefore has the same length as that of arginine. The product of the ornithine transcarbamoylase reaction is the amino acid citrulline. 

Ornithine is required to maintain the urea cycle. Conversion of glutamate to ornithine via glutamate semialdehyde is therefore an anaplerotic reaction. Melatonin is a compound synthesized from serotonin. It has a function in the circadian rhythm activity and has nothing to do with intracellular protein degradation. 

An adolescent went into delirium after eating a high-protein meal. He had an extremely high blood ammonia level and excreted orotic acid and uracil in the urine. His blood also contained increased glutamine and lysine levels. Despite heroic symptomatic treatment, the patient expired after 2 weeks. His liver tissue showed normal urea cycle enzyme levels, except that for ornithine transcar-bamylase (OTC), whose activity was only 10% that of normal liver. The Km of OTC was normal. Liver carabmylphosphate levels were about 10 times normal. Theoretical computer simulations indicated that urea can be produced at normal rates when liver OTC levels are higher than 0.3% of normal. 

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