This Sandbox is Reserved from 30/08/2012, through 01/02/2013 for use in the course "Proteins and Molecular Mechanisms" taught by Robert B. Rose at the North Carolina State University, Raleigh, NC USA. This reservation includes Sandbox Reserved 636 through Sandbox Reserved 685.
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Carbamoyl Phosphate Synthetase or CPS is a synthetase. A synthetase (also called a ligase) is an enzyme that links together two molecules by using energy derived from the splitting off of pyrophosphate from Adenosine Triphosphate(ATP). Carbamoyl phosphate synthetase is a heterodimeric enzyme which catalyzes the ATP-dependent synthesis of carbamoyl phosphate from glutamine or ammonia and bicarbonate. [1] This enzyme catalyzes the reaction of ATP and bicarbonate to produce carbonyl phosphate and ADP. Carbonyl phosphate reacts with ammonia to give carbamate. Carbamate reacts with a second ATP to give carbamoyl phosphate plus ADP.
It represents the first committed step in pyrimidine and arginine biosynthesis in prokaryotes and eukaryotes, and in the urea cycle in most terrestrial vertebrates.[2] Most prokaryotes carry one form of CPSase that participates in both arginine and pyrimidine biosynthesis, however certain bacteria can have separate forms.
StructureStructure
The structure of CPS was solved using X-ray diffraction. Carbamoyl phosphate synthase is composed of small and large subunits.[2] There are three active sites in CPSase, one in the and two in the . The small subunit contains the glutamine binding site, which catalyses the hydrolysis of glutamine to glutamate and ammonia which is used by the large chain to synthesize carbamoyl phosphate. The small subunit has a 3-layer beta/beta/alpha structure, and it is thought to be mobile in most proteins that carry it, and in the C-terminal domain of the small subunit of CPSase has glutamine amidotransferase activity. The large subunit has two homologous carboxy phosphate domains, each of them have ATP-binding sites; however, the N-terminal carboxy phosphate domain catalyses the phosphorylation of biocarbonate, and the C-terminal domain catalyses the phosphorylation of the carbamate intermediate.[7]The large subunit in bacterial CPSase has four structural domains:
1)
2)
3)
4)
The carboxy phosphate domain found duplicated in the large subunit of CPSase is also present as a single copy in the biotin-dependent enzymes acetyl-CoA carboxylase, propionyl-CoA carboxylase, pyruvate carboxylase and urea carboxylase. [3]
CPSase contain two molecular tunnels, an ammonia tunnel, and a carbamate tunnel. These are inter-domain tunnels that connect the three active sites, and they function as conduits for the transport of unstable reaction intermediates between the active site. [9]
MechanismMechanism
Mechanism for the formation of Carbamoyl Phosphate
Mechanism
In the small subunit, glutamine is converted to glutamate and ammonia.In the large subunit, the bicarbonate ion is phosphorylated by ATP to form the carbonylphosphate. Next, ammonia reacts with the carbonylphosphate to form carbamate. Finally, another ATP phosphorylates carbamate to form carbamoyl phosphate!
Regulation
Purine and pyrimidine nucleotides are allosteric regulators of CPSI. Purines are allosteric activators of the enzyme and pyrimidines are inhibitors of CPSI. Ornithine is also an allosteric activator of CPSI. Increased levels of ammonia are also found to activathe the enzyme while decreased levels of ATP can inhibit enzyme activity. All of the allosteric binding sites can be found in the large subunit of CPSI.
ImplicationsImplications
Carbamoyl Phosphate Synthetase deficiency is an autosomal recessive disease. The gene for this enzyme is found on chromosome 2. It is a disorder in which a person who is homozygous recessive for the gene does not have the ability to process waste Nitrogen.This disease can also be caused by mutations in the gene that codes for the enzyme. These mutations can cause the enzyme to be misshapen, shorter than normal, or prevent the enzyme from being translated at all. The absence or inactivity of this enzyme causes an increase in the levels of free ammonia and can effect the Central Nervous System. If left untreated, the disease will cause brain damage throughout the lifetime of the patient and will eventually become fatal. In newborns, the disease is usually catastrophic and if not caught promptly, will quickly result in death. Treatment of this disease is usually a change in diet. The patient will need to decrease protein intake and increase glucose and lipid intake. Arginine, benzoate, and phenylacetate are usually administered as well. In patients with dangerously high levels of ammonia in the blood, hemodialysis is required.
Another disease also related to CPS is a disorder in which a genetic change has occured that causes asparagine to be substituted for threonine at position 1405. This polymorphism is believed to cause a decrease in the production of Nitric Oxide, a compound which affects circulation by dilating blood vessels. This compound reduces blood pressure.
