A. Steps of Urea Cycle: 1. Formation of Carbamoyl Phosphate: Condensation of ammonium ion with bicarbonate ion resulting in the formation of carbamoyl phosphate by the help of the enzyme carbamoyl phosphate synthase-I present in the liver mitochondria. It requires Mg2+ and a dicarboxylic acid i.e. N-acetyl glutamate. This step requires 2 ATPs. 2. Synthesis of Citrulline: Carbamoyl phosphate formed in the first step combines with ornithine resulting in the synthesis of citrulline aided by the enzyme citrulline synthase or ornithine transcarbamoylase. Citrulline is easily permeable to the mitochondrial membrane and hence it diffuses into the cytosol. 3. Synthesis of Argininosuccinate: In the cytosol, citrulline combines with the amino acid aspaate forming argininosuccinate catalysed by the enzyme argininosuccinate synthase. It requires ATP which is hydrolysed to AMP resulting in utilization of two high energy bonds. Mg2+ acts as cofactor. 4. Cleavage of Argininosuccinate: The enzyme argininosuccinase acts reversibly to cleave argininosuccinate into Arginine and fumarate. Fumarate enters the TCA cycle (the linkage between TCA and urea cycle is known as Krebs bi-cycle). 5. Cleavage of Arginine: Arginine is lysed into ornithine and urea under the influence of the enzyme arginase. Hence arginine is known as semi-essential amino acid i.e. though it is synthesized in the body it is not available for protein synthesis. Ornithine is regenerated in this step and the urea cycle completes by the formation of urea. Ornithine and lysine are potent inhibitors of the enzyme arginase. Arginase is also present in testis, renal tubules, mammary gland and skin in minute quantities. The intermediate amino acids formed in the urea cycle i.e. ornithine, citrulline and argininosuccinate are known as non-protein amino acids. Ref-Sathyanarayana 4/e p370