Biological Chemistry - Berezov T. T., Korovkin B. F. 1998

Metabolism of Simple Proteins
Detoxification of Ammonia in the Body
Ornithine Urea Cycle

The primary mechanism for neutralizing ammonia in the body is urea Biosynthesis. Urea is excreted in the urine as the main end product of protein and, consequently, Amino acid METABOLISM. It accounts for up to 80–85% of total urinary nitrogen. The Liver is the primary and perhaps the sole site of urea synthesis. In 1932, H. Krebs and K. Henseleit first formulated the reaction equations for urea synthesis, representing them as a cycle known in literature as the Krebs Ornithine Urea Cycle. It is worth noting that this was the first cyclic metabolic system described in biochemistry, preceding H. Krebs's discovery of another metabolic pathway—The Tricarboxylic Acid Cycle (see above)—by nearly five years. Subsequent studies largely confirmed the cyclic nature of urea biosynthesis in the liver. Research by H. Cohen, S. Ratner, and co-workers helped clarify the intermediate steps and enzyme systems that catalyze urea formation.

Thus, the entire urea synthesis cycle can be outlined as follows. In the first step, the high-energy compound carbamoyl phosphate is synthesized—this is a metabolically active form of ammonia used as a Starting Material for the synthesis of pyrimidine NUCLEOTIDES (and, consequently, DNA and RNA) and Arginine (for Proteins and urea):

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To date, three distinct de novo pathways for carbamoyl phosphate synthesis, catalyzed by three different Enzymes, have been discovered. The first irreversible reaction is catalyzed by the regulatory enzyme ammonia-dependent carbamoyl phosphate synthetase (EC 6.3.4.16):

This reaction requires the expenditure of two ATP molecules, was discovered in the Cell/35.html">Mitochondria of liver Cells, and is primarily utilized for the synthesis of arginine and urea. N-acetylglutamate acts as an active allosteric stimulatory effector in this reaction.

The second, also irreversible, reaction is catalyzed by glutamine-dependent carbamoyl phosphate synthetase (EC 6.3.5.5):

This reaction was discovered in the Cytosol of animal cells and requires Mg²⁺ ions. It should be noted that, due to the inclusion of a hydrolytic step, this pathway is used primarily for the synthesis of pyrimidine nucleotides (see below). The enzyme is widely distributed in animal cells.

The third, reversible reaction is catalyzed by carbamate kinase (EC 2.7.2.2):

This reaction has been found in various microorganisms and is likely used for ATP resynthesis rather than carbamoyl phosphate synthesis.

In the second step of The urea cycle, carbamoyl phosphate and ornithine condense to form citrulline, a reaction catalyzed by ornithine transcarbamylase (EC 2.1.3.3).

In the next stage, citrulline is converted into arginine through two consecutive reactions. The first of these is an energy-requiring Condensation of citrulline and aspartic acid to form argininosuccinate, catalyzed by argininosuccinate synthetase. In the subsequent reaction, argininosuccinate is cleaved into arginine and fumarate by another enzyme, argininosuccinate lyase. In the final step, arginase cleaves arginine into urea and ornithine.

It must be emphasized that arginase is present in the liver of animals that excrete urea as the primary end product of Nitrogen metabolism. For instance, bird livers lack arginase because birds excrete uric acid instead of urea. Taking recent data into account, the ornithine urea cycle is illustrated in Fig. 12.5.

The overall reaction for urea synthesis, disregarding intermediates, can be represented as follows:

СO₂ + NH₃ + 3АТФ + 2Н₂O + Аспартат —> Мочевина + 2АДФ + АМФ +

+ Фумарат + 2Р_i+ РР_і.

This reaction is accompanied by a decrease in Free energy ($\Delta G^0 = -40$ kJ), meaning the process always proceeds in the direction of urea synthesis. It should be noted that urea synthesis is energetically costly for the Organism. The synthesis of a single urea molecule requires four high-energy phosphate groups: two ATP molecules are consumed in the synthesis of carbamoyl phosphate, and one is used to form argininosuccinic acid, during which ATP is cleaved into AMP and $\text{PP}_i$, which upon Hydrolysis also yields two $\text{P}_i$ molecules.

Fig. 12.5. The ornithine cycle of urea synthesis in the liver.

From the provided scheme of urea formation, it is easy to see that one of the nitrogen atoms in urea originates from free ammonia (via carbamoyl phosphate), while the second nitrogen atom is supplied by aspartate. Ammonia is generated primarily through the Glutamate dehydrogenase reaction. Three coupled reactions are involved in replenishing aspartate pools: first, fumarate adds Water under the action of fumarase to form malate, which is then oxidized by malate dehydrogenase to yield oxaloacetate; the latter regenerates aspartate via a Transamination reaction with glutamate.

Considering the available factual data on the mechanisms of ammonia detoxification in the body, the following Conclusions can be drawn. A portion of ammonia is utilized in the BIOSYNTHESIS OF AMINO Acids through the reductive amination of $\alpha$-keto acids via transamination. Ammonia is also bound during The biosynthesis of glutamine and asparagine. A small amount of ammonia is excreted in the urine as ammonium salts. A significant portion of Amino Acid Nitrogen is eliminated from the body as creatinine, which is derived from creatine and creatine phosphate. The largest share of ammonia goes toward the synthesis of urea, which is excreted in the urine as the principal end product of Protein metabolism in humans and animals. Estimates show that a healthy adult in nitrogen balance consumes and consequently excretes approximately 15 g of nitrogen per day. Of the excreted urinary nitrogen, urea accounts for about 85%, creatinine about 5%, ammonium salts 3%, uric acid 1%, and other forms roughly 6%.

Throughout evolution, living organisms have developed Different types of nitrogen metabolism. The ammonotelic type, in which ammonia is the primary end product of nitrogen metabolism, is characteristic mainly of fish. In the ureotelic type, urea is the main end product of protein metabolism; this type is typical of humans and animals. The uricotelic type is characteristic of birds and reptiles, with uric acid being the main end product of metabolism.