Carbamoyl phosphate synthetase synthesis

The two conditions can be distinguished by an increase in orotic add and uracil, which occurs in ornithine transcarbamoylase deficiency, but not in the defldency of carbamoyl phosphate synthetase. Orotic acid and uracil are intermediates in pyrimidine synthrais (see Chapter 18). This pathway is stimulated by the accumulation of carbamoyl phosphate, the substrate for ornithine transcarbamoylase in the urea cycle and for aspartate transcarbamoylase in pyrimidine synthesis. 

Pyrimidines are synthesized de novo in the cytoplasm from aspartate, COj, and glutamine as shown in Figure 1-18-2. Synthesis involves a cytoplasmic carbamoyl phosphate synthetase that differs from the mitochondrial enzyme with the same name used in the urea cycle. 

The carbamoyl phosphate synthetase (abbreviated to CPS-I) that is involved in the ornithine cycle differs from the enzyme that is involved in pyrimidine synthesis (carbamoyl phosphate synthetase-ll). The latter enzyme is cytosolic, requires glutamine for provision of nitrogen, rather than ammonia, and is regulated by different factors (Chapter 20). 

Carbamoyl phosphate synthetase formation in liver taken from tadpoles treated with thyroxine is enhanced by the addition of orotic acid, uracil or uridine (cytosine and adenosine had no effect). The synthesis of this enzyme is not affected by these pyrimidines in untreated animals. This indicates that there is a relative pyrimidine deficiency during thyroxine-induced metamorphosis [140]. 

MECHANISM FIGURE 18-11 Nitrogen-acquiring reactions in the synthesis of urea. The urea nitrogens are acquired in two reactions, each requiring ATP. (a) in the reaction catalyzed by carbamoyl phosphate synthetase 1, the first nitrogen enters from ammonia. The terminal phosphate groups of two molecules of ATP are used to form one molecule of carbamoyl phosphate. In other words, this reaction has two activa-... 

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. 

RGURE 18-13 Synthesis of /V-acetylglutamate and its activation of carbamoyl phosphate synthetase I. 

Carbamoyl phosphate synthetase 1 deficiency <0.5 Urea synthesis Carbamoyl phosphate synthetase 1 Lethargy convulsions early death... 

FIGURE 22-36 De novo synthesis of pyrimidine nucleotides biosynthesis of UTP and CTP via orotidylate. The pyrimidine is constructed from carbamoyl phosphate and aspartate. The ribose 5-phosphate is then added to the completed pyrimidine ring by orotate phosphori-bosyltransferase. The first step in this pathway (not shown here see Fig. 18-11a) is the synthesis of carbamoyl phosphate from C02 and NH), catalyzed in eukaryotes by carbamoyl phosphate synthetase II. 

The regulated step of this pathway in mammalian cells is the synthesis of carbamoyl phosphate from glutamine and C02, catalyzed by carbamoyl phosphate synthetase II (CPS U). CPS II is inhibited... 

Unlike in purine biosynthesis, the pyrimidine ring is synthesized before it is conjugated to PRPP. The first reaction is the conjugation of carbamoyl phosphate and aspartate to make N-carbamoylaspartate. The carbamoyl phosphate synthetase used in pyrimidine biosynthesis is located in the cytoplasm, in contrast to the carbamoyl phosphate used in urea synthesis, which is made in the mitochondrion. The enzyme that carries out the reaction is aspartate transcarbamoylase, an enzyme that is closely regulated. 

The first step in de novo pyrimidine biosynthesis is the synthesis of carbamoyl phosphate from bicarbonate and ammonia in a multistep process, requiring the cleavage of two molecules of ATP. This reaction is catalyzed by carbamoyl phosphate synthetase (CPS) (Section 23.4.1). Analysis of the structure of CPS reveals two homologous domains, each of which catalyzes an ATP-dependent step (Figure 25.3). 

The primary site of regulation is Carbamoyl Phosphate Synthetase II (glutamine) which is allosterically inhibited by UTP. Elevated PRPP increases the CPS-II activity to help control PRPP levels. Feedback inhibition (control) is provided by TDP inhibition of PRPP synthesis and UMP inhibition of OMP Decarboxylase. 

In the first reaction, glutamine reacts with C02 and 2 ATP to form carbamoyl phosphate. This reaction is analogous to the first reaction of the urea cycle. However, for pyrimidine synthesis, glutamine provides the nitrogen and the reaction occurs in the cytosol, where it is catalyzed by carbamoyl phosphate synthetase II, which is inhibited by UTP. 

C. Options A and B are true for purine but not pyrimidine biosynthesis. During pyrimidine synthesis, the entire aspartate molecule is incorporated into the ring. Glutamine is the substrate for carbamoyl phosphate synthetase II, the enzyme involved in pyrimidine biosynthesis. (NH4+ is the substrate for synthetase I used in urea synthesis.) Glydne supplies one nitrogen for purine synthesis. 

De novo pathway of uridine-5 -monophosphate (UMP) synthesis. Enzymes (1) carbamoyl phosphate synthetase II (2) aspartate transcarbamoylase (3) dihydroorotase (4) dihydroorotate dehydrogenase (5) orotate phosphoribosyltransferase (6) orotidine-5 -monophosphate decarboxylase (orotidylate decarboxyla.se). 

The urea cycle begins with the coupling of free NH3 with HC(.) A to form carbamoyl phosphate. Carbamoyl phosphate is a simple molecule, but its synthesis is complex. Carbamoyl phosphate synthetase catalyzes the required three steps. 

Note that NH, because it is a strong base, normally exists as NH4 in aqueous solution. However, carbamoyl phosphate synthetase uses only NH3 as a substrate. The reaction begins with the phosphorylation of HCO3 to form carboxyphosphate, which then reacts with NH3 to form carbamic acid. Finally, a second molecule of ATP phosphofylates carbamic acid to form carbamoyl phosphate. The structure and mechanism of the enzyme that catalyzes these reactions will be presented in Chapter 25. The consumption of two molecules of ATP makes this synthesis of carbamoyl phosphate essentially irreversible. The mammalian enzyme requires N-acetylglutamate for activity, as will be described shortly. 

The answer is c. (Murray, pp 375-401. Scriver, pp 2513-2570. Sack, pp 121-138. Wilson, pp 287-320.) The steps of pyrimicfine nucleotide biosynthesis are summarized in the figure below. The first step in pyrimidine synthesis is the formation of carbamoyl phosphate. The enzyme catalyzing this step, carbamoyl phosphate synthetase (1), is feedback-inhibited by UMP through allosteric effects on enzyme structure (not by competitive inhibition with its substrates). The enzyme of the second step, aspartate transcarbamoylase, is composed of catalytic and regulatory subunits. The regulatory subunit binds CTP or ATP TTP has no role in the feedback inhibition of pyrimidine synthesis. Decreased rather than increased activity of enzymes 1 and 2 would be produced by allosteric feedback inhibition. 

Urea synthesis, which occurs in hepatocytes, begins with the formation of carbamoyl phosphate in the matrix of mitochondria. The substrates for this reaction, catalyzed by carbamoyl phosphate synthetase I, are NH4 and HCO3. (The nitrogen source for carbamoyl phosphate synthetase II, the enzyme involved in pyrimidine synthesis, is glutamine.)... 

The major control of the urea cycle, other than the availability of nitrogen, is exerted at the first potentially irreversible step of urea synthesis, that is, carbamoyl phosphate synthetase. Carbamoyl phosphate... 

The control of urea synthesis is availability of nitrogen and activity of carbamoyl phosphate synthetase (CPSI). 

These substrates are different than those with the carbamoyl phosphate used for urea synthesis. The enzyme for the glutamine-dependent carbamoyl phosphate synthetase (CPS II) is in the cytosol, whereas that for urea synthesis (CPS I) is in the mitochondrion. The glutamine-dependent carbamoyl phosphate synthetase is present in most cells, whereas the mitochondrial carbamoyl phosphate synthetase is present primarily in the liver, kidney, and intestines (Fig. 20.6). 

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