Argininosuccinate synthetase

In the hepatic cytosol, citrulline reacts with aspartate to form argininosuccinate, catalyzed by argininosuccinate synthetase (AS Fig. 40-5 reaction 3). The mRNA for this enzyme is increased by starvation, corticosteroids or cyclic-AMP. Citrulline itself potently induces the mRNA. 

Z. Z., Morris, S. M., )r., Coinduction of nitric oxide synthase and argininosuccinate synthetase in a murine macrophage cell line. Implications for regulation of nitric oxide production,/. Biol. Chem. 269 (1994), p. 1257-12561... 

Condition Carbamoyl- phosphate synthetase Ornithine transcarbamoylase Argininosuccinate synthetase Argininosuccinate lyase Arginase... 

In view of the toxicity of ammonia, complete absence of any one of the enzymes of the cycle is fatal. Nonetheless, disorders of the cycle do occur, which are caused by a low activity of one of the enzymes or carbamoyl phosphate synthetase. In addition, defects in N-acetylglutamate synthase have been reported, but they are very rare. With the exception of ornithine transcarbamoylase, the deficiencies have an autosomal recessive mode of inheritance. The transcarbamoylase deficiency is inherited as an X-linked dominant trait, usually lethal in male patients. A deficiency of carbamoyl phosphate synthetase, ornithine transcarbamoylase or argininosuccinate synthetase results in accumulation and excretion of citrulline. A deficiency of argininosuccinate lyase results in the accumulation and excretion of argininosuccinate and arginine (Table 10.5). The abbreviations CPSD, OTCD, ASD, ALD and AD stand, respectively, for the deficiencies of these enzymes, where D stands for deficiency. 

OCTOPINE DEHYDROGENASE ARGININOSUCCINATE LYASE ARGININOSUCCINATE SYNTHETASE ARISTOLOCHENE SYNTHASE Arogenate dehydrogenase,... 

ARACHIDONYL-CoA SYNTHETASE ARGININOSUCCINATE SYNTHETASE ASPARAGINE SYNTHETASE... 

Argininosuccinate synthetase deficiency, which leads to citrullinemia. 

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 citrulline is then condensed with aspartate, the source of the second nitrogen atom in urea, by the enzyme argininosuccinate synthetase to form argininosuccinate. This reaction is driven by the hydrolysis of ATP to AMP and PP , with subsequent hydrolysis of the pyrophosphate. Thus both of the high-energy bonds in ATP are ultimately cleaved. 

Fig. 1. The urea cycle. The enzymes involved in this cycle are (1) carbamoyl phosphate synthetase (2) ornithine transcarbamoylase (3) argininosuccinate synthetase (4) arginosuccinase and (5) arginase.

Quantitation of plasma amino acids, especially of citrullinc, is the first step in determining the precise enzyme or transport protein defect in patients with a UCD. If the defect involves N-acetyglutamate synthetase (NAGS), CPSI, or OTC, then plasma citrulline concentration will be low. Marked hypercitrullinemia (>2000 pmol/I.) is seen in argininosuccinate synthetase (AS) deficiency, while moderate increases (>200 pmol/L, normal undetectable) are found in argininosuccinate lyase (AL) and citrin deficiencies. In AL deficiency, the presence of argininosuccinic acid and its anhydrides further distinguishes this disorder. 

Once citrulline is in the cytosol, argininosuccinic acid is formed by condensation of citrulline with aspartate. This is where the second nitrogen atom enters the cycle. Argininosuccinate synthetase, a homotetramer of a 46-kd polypeptide catalyzes the reversible reaction accompanied by hydrolysis of ATP to AMP and pyrophosphate. The subsequent hydrolysis of pyrophosphate shifts the equilibrium to the right and results in the consumption of two high-energy phosphate bonds. 

Arginine is synthesized from aspartate and ornithine during urea formation. Argininosuccinate synthetase and -lyase catalyze the condensation and cleavage reactions, respectively, that result in the formation of arginine (Sec. 15.5). 

Argininosuccinate synthetase (the second enzyme of the urea cycle) and the remaining two... 

Figure 6.10 The urea cycle. The enzymes of the urea cycle include, 1 carbamoyl phosphate synthetase-I, 2 ornithine transcarbamoylase, 3 argininosuccinate synthetase, 4 argininosuc-cinase, 5 arginase.

Classic citrullinaemia Argininosuccinate synthetase Episodic hyperammonaemia, vomiting, lethargy, ataxia, seizures, eventual coma. Treatment is with arginine administration to enhance citrulline excretion, also with sodium benzoate for ammonia detoxification. 

Citrulline is transported to the cytoplasm where it condenses with aspartate, the donor of the second amino group of urea. This synthesis of argininosuccinate, catalyzed by argininosuccinate synthetase, is driven by the cleavage of ATP into AMP and pyrophosphate and by the subsequent hydrolysis of pyrophosphate. 

The enzymes that catalyze these steps are homologous to argininosuccinate synthetase and argininosuccinase, respectively. Thus, four of the five enzymes in the urea cycle were adapted from enzymes taking part in nucleotide biosynthesis. The remaining enzyme, arginase, appears to be an ancient enzyme found in all domains of life. 

Therapeutic design. How would you treat an infant who is deficient in argininosuccinate synthetase Which molecules would carry nitrogen out of the body ... 

A defect in argininosuccinate synthetase coincides with markedly elevated citrulline values in the blood. Neither citrullinaemia nor a carbamylphosphate synthetase defect cause liver damage. By contrast, an argininosuc-cinase defect leads to microvesicular steatosis and megamitochondria with dilatation of the ER. 

In a 2-step reaction, catalyzed by cytosolic argininosuccinate synthetase, citrulline is converted to argininosuccinate. The reaction involves the addition of AMP (from ATP) to the amido carbonyl of citrulline, forming an activated intermediate on the enzyme surface (AMP-citrulline), and the subsequent addition of aspartate to form argininosuccinate. 

Kato, H Mi/utani-Funahashi, M., bhiosaka, S-, and Nakagawa, H. (1978). Circadian rhythms of urea formation and argininosuccinate synthetase activity in rat liver. /. Nutr, 108, 1071-1D77. 

---