Nitrogen Excretion and the Urea Cycle:- Urea Is Produced from Ammonia in Five Enzymatic Steps
The urea cycle begins inside liver mitochondria, but three of the subsequent steps take place in the cytosol; the cycle thus spans two cellular compartments (Fig. 18–10). The first amino group to enter the urea cycle is derived from ammonia in the mitochondrial matrix—NH4+ arising by the pathways described above.
The liver also receives some ammonia via the portal vein from the intestine, from the bacterial oxidation of amino acids. Whatever its source, the NH4 generated in liver mitochondria is immediately used, together with CO2 (as HCO3-) produced by mitochondrial respiration, to form carbamoyl phosphate in the matrix (Fig. 18–11a; see also Fig. 18–10). This ATP-dependent reaction is catalyzed by carbamoyl phosphate synthetase I, a regulatory enzyme (see below). The mitochondrial form of the enzyme is distinct from the cytosolic (II) form, which has a separate function in pyrimidine biosyn the sis (Chapter 22). 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 Pi (Fig. 18–10, step 1). 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. The second amino group now enters from aspartate (generated in mitochondria by transamination and trans ported into the cytosol) by a condensation reaction between the amino group of aspartate and the ureido (carbonyl) group of citrulline, forming argininosuccinate (step 2 in Fig. 18–10). This cytosolic reaction, cat alyzed by argininosuccinate synthetase, requires ATP and proceeds through a citrullyl-AMP intermediate (Fig. 18–11b). The argininosuccinate is then cleaved by argininosuccinase (step 3 in Fig. 18–10) to form free arginine and fumarate, the latter entering mitochondria to join the pool of citric acid cycle intermediates. This is the only reversible step in the urea cycle. In the last reaction of the urea cycle (step 4 ), the cytosolic en zyme arginase cleaves arginine to yield urea and ornithine. Ornithine is transported into the mitochondrion to initiate another round of the urea cycle. 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 di rectly 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.
FIGURE 18–10 (facing page)Urea cycle and reactions that feed amino groups into the cycle. The enzymes catalyzing these reactions (named in the text) are distributed between the mitochondrial matrix and the cytosol. One amino group enters the urea cycle as carbamoyl phosphate, formed in the matrix; the other enters as aspartate, formed in the matrix by transamination of oxaloacetate and glutamate, cat alyzed by aspartate aminotransferase. The urea cycle consists of four steps. 1Formation of citrulline from ornithine and carbamoyl phosphate (entry of the first amino group); the citrulline passes into the cy tosol. 2Formation of argininosuccinate through a citrullyl-AMP intermediate (entry of the second amino group). 3Formation of arginine from argininosuccinate; this reaction releases fumarate, which enters the citric acid cycle. 4Formation of urea; this reaction also regenerates, ornithine. The pathways by which NH4+ arrives in the mitochondrial matrix of hepatocytes were discussed in Section 18.1.
MECHANISM FIGURE 18–11 Nitrogen-acquiring reactions in the syn thesis of urea. The urea nitrogens are acquired in two reactions, each requiring ATP. (a) In the reaction catalyzed by carbamoyl phosphate synthetase I, 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 activation steps (1and 3). Carbamoyl Phosphate Synthetase I Mechanism (b)In the reaction catalyzed by argininosuccinate synthetase, the second nitrogen enters from aspartate. The ureido oxygen of citrulline is activated by the addition of AMP in step 1 ; this sets up the addition of aspartate in step 2 , with AMP (including the ureido oxygen) as the leaving group.
