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Arginase

How does the body get rid of toxic nitrogen?

To remove toxic nitrogen from the body a series of enzymes work together in the urea cycle to decompose amino acids and remove unwanted nitrogen. Arginase is the fith and final enzyme of this cycle that catalyzes the hydrolytic break down of the amino acid L-arginine into L-ornithine and urea. Arginase takes on two distinct forms: Class I and II, which refer to the catalytic activity of the enzyme. In addition, each class is encoded by a different gene. Arginase I (arg1) is found in the cytoplasm of liver cells functioning in the urea cycle, while arginase II (arg2) is found ubiquitously in the mitochondria of other tissues such as kidney, brain and small intestine. Arginase belongs to the enzymatic class hydrolase. Arginase functions by cleaving the amino acid arginine with water to produce urea. Arginase is known to have a high specific activity and functions to produce urea in appreciable amounts for excretion.^[1] In addition, this enzyme allows the regeneration of ornithine which is the final product of the urea cycle. In lieu of a deficiency in arginase, urea is not formed causing an increase in the amount of ammonia and arginine in the body.

Structure

Arginase is unique because of all the enzymes found in the urea cycle this enzyme has different isoforms. In the human body there are two isozymes with similar structural properties and differing immunological properties. Common among the differing isozymes is the requirement of two divalent cation cofactors for activity. . is the most common cofactor for activity but cobalt, nickel and iron have been reported to be suitable cofactors as well. The two required manganese II ions are located in two separate subunits and are separated by approximately 3.36-3.57 Å and bridged by oxygen and a solvent derived hydroxide ^[2] Many varying isoforms of arginase have produced different structural models based on the particular isoform and organism of origin. The most well studied arginase enzyme, arginase I isolated from rat liver, is a trimeric enzyme and each subunit is ordered in an alpha-beta-alpha structure. Each fold contains an eight-stranded, parallel β-sheet surrounded by alpha helices, with an approximate size 35kDa ^[3]. Fully active enzymes contain a binuclear metal cofactor center in a 15Å cleft that serves as the active site. One cofactor is held in place by terminal ligands His 101 and Asp 128, and bridged by Asp 124, Asp 132 and hydroxide ion ^[4]. The other cofactor held by terminal ligand residues His 101 and Asp 234 with bridging ligands Asp 232 and hydroxide ion ^[5].

Although arginase is present in different isoforms, there is a high level of conservation among the amino acids. For example, human type I and II arginases share a 58% sequence identity ^[6]. Overall, there are 20 amino acids that are highly conserved over the varying isoforms. Most of these conserved residues are key structural residues. Included in these conserved structural residues is gly 23, which begins the first α-helix of arginase^[7]. In addition to highly conserved nucleotides, there exist a few key invariant nucleotides as well. Among the most notable are Val 120, Ile 121, Val 233 and Ile 264 which are attributed to the packing of the hydrophobic interior of the protein. Roles of conserved residues in the arginase family^[8].

Mechanism

Arginine + H2O → Ornithine + Urea

The mechanism regarding arginase is highly important in the production of urea through the urea cycle. During the final step of the urea cycle, the amino acid arginine is present and needs to be cleaved in order for urea to be produced. Arginine is hydrolyzed and cleaved with the hydroxyl at the end of the amino group. We refer to this group as a guanidium end. The guanidium group is positively charged and has a high pKa. Therefore, it is willing to donate a proton since it has a hydrogen to give. First, the ligand will ionize the water to form hydroxide. The hydroxide now attacks the guanidine carbon and protons are transferred from the hydroxide to the substrate bridging. In the third and final step protons are transferred through a proton shuffle to create ornithine and urea. Afterwards, the urea is excreted through urine. Leaving the excess ornithine to be recycled through the final step in the cycle to react with citrulline and eliminate ammonia from the body. ^[9]

Medical Application

Arginine Deficiency

The urea cycle is responsible for producing urea to remove unwanted nitrogen from the body so that the body is able to maintain normal functions. Arginase is a key player in this process and functions to break down the amino acid arginine into ornithine while releasing urea. However, if the body is not able to dispose of the urea it can spell trouble for specific bodily functions. Similarly, an arginase deficiency can spell trouble as well. Studies have linked the deficiency of arginase to genetic inheritability and is thereby considered a genetic disorder. When argininase is deficient, it causes a build up in ammonia and arginine. An accumulation of high levels of ammonia is toxic to the body and can cause neurological effects.

Genetic Factors

This particular image shows how the gene for ARG1 is inherited.

How does the specific cell produce the enzyme?

Arginase is present in our liver cells due to the a specific gene called ARG1. The function of the ARG1 gene is primarily to provide instructions in order for liver cells to produce the enzyme arginase. Research has provided that this deficiency is caused by inheritance. Therefore, a deficiency in arginase production is due to a genetic mutation. New findings show 12 mutations have been accurately identified in the ARG1 gene. A mutated ARG1 gene may result in an arginase enzyme that is unstable, shorter than usual, oddly shaped, or may prevent the enzyme from being produced at all. More importantly the shape of the enzyme means alot as far as determining the chemical nature of argininase. This enzyme has a highly specific activity, a very specific active and will not function if any changes occur during the transcription and translation process of the gene. Changes in the gene have a result of an enzyme that is not able to correctly convert arginine into the products of ornithine and urea. If the body is unable to produce urea to remove the unwanted nitrogen, the body will accumulate excess nitrogen that can form toxic ammonia which may cause a variety of symptoms in the body.

Symptoms

Spastic tetraplegia

Progressive mental retardation

Seizures

Hyperactivity

Growth failure

Elevated blood ammonia level

Enlarged liver

Lack of appetite

Vomiting

Increased blood level of arginine

Muscle stiffness

Spasticity

Tremor

Balance problems

Coordination problems

Irritability

Treatments

1. Protein intake control – the amount of protein consumed is restricted for patients. A carefully monitored diet plan is necessary.

2. Intravenous Therapy - Ammonia sometimes accumulates at high levels which would be highly toxic. This treatment is rarely needed. however, if required, then the patient must go through hemodialysis to rid of ammonia levels.

3. Long-term therapy - this treatment requires an extended time for a patient to follow a low-protein diet. In addition, he/she may take oral sodium benzoate or sodium phenylbutyrate. However this is guided by a physician.

References

Ash, David Structure and Function of Arginases [http://www.ncbi.nlm.nih.gov/pubmed/15465781

Roche, Victoria Improving Pharmacy Students' Understanding and Long-term Retention of Acid-Base Chemistry [1]

Corporation, Worthington Biochemical. Arginase [2]

Perozich, John Roles of conserved residues in the arginase family [3]

Morris Jr., S.M. Recent Advances in Arginine Metabolism: Roles and Regulation of the Arginases. Br. J. Pharmacol. 2009, 157, 922-930.

Abumrad NN, Barbul A. The use of arginine in clinical practice. In: Cynober LA, editor. Metabolic and Therapeutic Aspects of Amino Acids in Clinical Nutrition. Boca Raton, FL: CRC Press; 2004. pp. 595–611