Carbamoyl phosphate synthetase catalyzes the ATP-dependent synthesis of

carbamoyl phosphate Carbamoyl phosphate is an anion of biochemical significance. In land-dwelling animals, it is an intermediary metabolite in nitrogen disposal through the urea cycle and the synthesis of pyrimidines. Its enzymatic counterpart, carbamoyl phosphate sy ...

from

glutamine Glutamine (symbol Gln or Q) is an α-amino acid that is used in the biosynthesis of proteins. Its side chain is similar to that of glutamic acid, except the carboxylic acid group is replaced by an amide. It is classified as a charge-neutral ...

() or ammonia () and bicarbonate. This enzyme catalyzes the reaction of ATP and

bicarbonate In inorganic chemistry, bicarbonate (IUPAC-recommended nomenclature: hydrogencarbonate) is an intermediate form in the deprotonation of carbonic acid. It is a polyatomic anion with the chemical formula . Bicarbonate serves a crucial biochem ...

to produce carboxy phosphate and ADP. Carboxy phosphate reacts with

ammonia Ammonia is an inorganic compound of nitrogen and hydrogen with the formula . A stable binary hydride, and the simplest pnictogen hydride, ammonia is a colourless gas with a distinct pungent smell. Biologically, it is a common nitrogenous wa ...

to give

carbamic acid Carbamic acid, which might also be called aminoformic acid or aminocarboxylic acid, is the chemical compound with the formula . It can be obtained by the reaction of ammonia and carbon dioxide at very low temperatures, which also yields an equ ...

. In turn, carbamic acid reacts with a second ATP to give

plus ADP. It represents the first committed step in pyrimidine and arginine biosynthesis in prokaryotes and eukaryotes, and in the

urea cycle The urea cycle (also known as the ornithine cycle) is a cycle of Biochemistry, biochemical reactions that produces urea (NH2)2CO from ammonia (NH3). Animals that use this cycle, mainly amphibians and mammals, are called ureotelic. The urea cycle ...

in most terrestrial

vertebrates Vertebrates () comprise all animal taxa within the subphylum Vertebrata () (chordates with backbones), including all mammals, birds, reptiles, amphibians, and fish. Vertebrates represent the overwhelming majority of the phylum Chordata, with c ...

. Most

prokaryote A prokaryote () is a single-celled organism that lacks a nucleus and other membrane-bound organelles. The word ''prokaryote'' comes from the Greek πρό (, 'before') and κάρυον (, 'nut' or 'kernel').Campbell, N. "Biology:Concepts & Conne ...

s carry one form of CPSase that participates in both arginine and pyrimidine biosynthesis, however certain

bacteria Bacteria (; singular: bacterium) are ubiquitous, mostly free-living organisms often consisting of one Cell (biology), biological cell. They constitute a large domain (biology), domain of prokaryotic microorganisms. Typically a few micrometr ...

can have separate forms. There are three different forms that serve very different functions: * Carbamoyl phosphate synthetase I (mitochondria,

) * Carbamoyl phosphate synthetase II (cytosol,

pyrimidine metabolism Pyrimidine biosynthesis occurs both in the body and through organic synthesis. ''De novo'' biosynthesis of pyrimidine ''De Novo'' biosynthesis of a pyrimidine is catalyzed by three gene products CAD, DHODH and UMPS. The first three enzymes of ...

). * Carbamoyl phosphate synthetase III (found in fish).

Mechanism

Carbamoyl phosphate synthase has three main steps in its mechanism and is, in essence, irreversible.Biochemistry, 3rd edition, J.M. Berg, J.L. Tymoczko, L. Stryer # Bicarbonate ion is phosphorylated with ATP to create carboxylphosphate. # The carboxylphosphate then reacts with ammonia to form carbamic acid, releasing inorganic phosphate. # A second molecule of ATP then phosphorylates carbamic acid, creating carbamoyl phosphate. The activity of the enzyme is known to be inhibited by both

Tris Tris, or tris(hydroxymethyl)aminomethane, or known during medical use as tromethamine or THAM, is an organic compound with the formula (HOCH2)3CNH2, one of the twenty Good's buffers. It is extensively used in biochemistry and molecular biology as ...

and

HEPES HEPES (4-(2-hydroxyethyl)-1-piperazineethanesulfonic acid) is a zwitterionic sulfonic acid buffering agent; one of the twenty Good's buffers. HEPES is widely used in cell culture, largely because it is better at maintaining physiological pH de ...

buffers.

Structure

Carbamoyl

phosphate In chemistry, a phosphate is an anion, salt, functional group or ester derived from a phosphoric acid. It most commonly means orthophosphate, a derivative of orthophosphoric acid . The phosphate or orthophosphate ion is derived from phosph ...

synthase (CPSase) is a

heterodimeric In biochemistry, a protein dimer is a macromolecular complex formed by two protein monomers, or single proteins, which are usually non-covalently bound. Many macromolecules, such as proteins or nucleic acids, form dimers. The word ''dimer'' has ...

enzyme Enzymes () are proteins that act as biological catalysts by accelerating chemical reactions. The molecules upon which enzymes may act are called substrates, and the enzyme converts the substrates into different molecules known as products ...

composed of a small and a large subunit (with the exception of CPSase III, which is composed of a single polypeptide that may have arisen from gene fusion of the glutaminase and

synthetase In biochemistry, a ligase is an enzyme that can catalyze the joining (ligation) of two large molecules by forming a new chemical bond. This is typically via hydrolysis of a small pendant chemical group on one of the larger molecules or the enz ...

domains). CPSase has three active sites, one in the small subunit and two in the large subunit. The small subunit contains the

binding site In biochemistry and molecular biology, a binding site is a region on a macromolecule such as a protein that binds to another molecule with specificity. The binding partner of the macromolecule is often referred to as a ligand. Ligands may includ ...

and catalyses the

hydrolysis Hydrolysis (; ) is any chemical reaction in which a molecule of water breaks one or more chemical bonds. The term is used broadly for substitution, elimination, and solvation reactions in which water is the nucleophile. Biological hydrolys ...

of glutamine to glutamate and

, which is in turn used by the large chain to synthesize carbamoyl phosphate. The small subunit has a 3-layer beta/beta/alpha structure, and is thought to be mobile in most

protein Proteins are large biomolecules and macromolecules that comprise one or more long chains of amino acid residues. Proteins perform a vast array of functions within organisms, including catalysing metabolic reactions, DNA replication, res ...

s that carry it. The C-terminal domain of the small subunit of CPSase has glutamine amidotransferase activity. The large subunit has two homologous carboxy phosphate domains, both of which have ATP-binding sites; however, the N-terminal carboxy phosphate domain

catalyse Catalysis () is the process of increasing the rate of a chemical reaction by adding a substance known as a catalyst (). Catalysts are not consumed in the reaction and remain unchanged after it. If the reaction is rapid and the catalyst recyc ...

s the phosphorylation of biocarbonate, while the C-terminal domain catalyses the phosphorylation of the carbamate intermediate. 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 (ACC),

propionyl-CoA carboxylase Propionyl-CoA carboxylase (, PCC) catalyses the carboxylation reaction of propionyl-CoA in the mitochondrial matrix. PCC has been classified both as a ligase and a lyase. The enzyme is biotin-dependent. The product of the reaction is (S)-methylmal ...

(PCCase),

pyruvate carboxylase Pyruvate carboxylase (PC) encoded by the gene PC is an enzyme () of the ligase class that catalyzes (depending on the species) the physiologically irreversible carboxylation of pyruvate to form oxaloacetate (OAA). Image:Pyruvic-acid-2D-sk ...

(PC) and urea carboxylase. The large subunit in bacterial CPSase has four

structural A structure is an arrangement and organization of interrelated elements in a material object or system, or the object or system so organized. Material structures include man-made objects such as buildings and machines and natural objects such ...

domains: the carboxy phosphate domain 1, the oligomerisation domain, the carbamoyl phosphate domain 2 and the allosteric domain. CPSase heterodimers from ''

Escherichia coli ''Escherichia coli'' (),Wells, J. C. (2000) Longman Pronunciation Dictionary. Harlow ngland Pearson Education Ltd. also known as ''E. coli'' (), is a Gram-negative, facultative anaerobic, rod-shaped, coliform bacterium of the genus '' Esc ...

'' contain two molecular tunnels: an ammonia tunnel and a carbamate tunnel. These inter-domain tunnels connect the three distinct active sites, and function as conduits for the

transport Transport (in British English), or transportation (in American English), is the intentional movement of humans, animals, and goods from one location to another. Modes of transport include air, land ( rail and road), water, cable, pipelin ...

of unstable reaction intermediates (ammonia and carbamate) between successive active sites. The catalytic mechanism of CPSase involves the

diffusion Diffusion is the net movement of anything (for example, atoms, ions, molecules, energy) generally from a region of higher concentration to a region of lower concentration. Diffusion is driven by a gradient in Gibbs free energy or chemica ...

of carbamate through the interior of the enzyme from the site of synthesis within the N-terminal domain of the large subunit to the site of phosphorylation within the C-terminal domain.

References

External links

GeneReviews/NCBI/NIH/UW entry on Urea Cycle Disorders Overview
{{Portal bar, Biology, border=no Urea cycle Protein domains EC 6.3.4