UREASE
Ureases (EC 3.5.1.5), functionally, belong to the superfamily of amidohydrolases and phosphotriesterases.[2]
Ureases are found in numerous bacteria, fungi, algae, plants, and some invertebrates, as well as in soils, as a soil enzyme. They are nickel-containing metalloenzymes of high molecular weight.[3]
These enzymes catalyze the hydrolysis of urea into carbon dioxide and ammonia:
(NH2)2CO + H2O urease→ CO2 + 2NH3
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The hydrolysis of urea occurs in two stages. In the first stage, ammonia and carbamic acid are produced. The carbamate spontaneously and rapidly hydrolyzes to ammonia and carbonic acid. Urease activity increases the pH of its environment as ammonia is produced, which is basic.
Its activity was first identified in 1876 by Frédéric Alphonse Musculus as a soluble ferment.[4] In 1926, James B. Sumner, showed that urease is a protein by examining its crystallized form.[5]
Sumner's work was the first demonstration that a protein can function as an enzyme and led eventually to the recognition that most enzymes are in fact proteins.
Urease was the first enzyme crystallized.
For this work, Sumner was awarded the Nobel prize in chemistry in 1946.[6]
The crystal structure of urease was first solved by P. A. Karplus in 1995.[5]
Structure
A 1984 study focusing on urease from jack bean found that the active site contains a pair of nickel centers.[7]
In vitro activation also has been achieved with manganese and cobalt in place of nickel.[8]
Lead salts are inhibiting.
The molecular weight is either 480 kDa or 545 kDa for jack-bean urease (calculated mass from the amino acid sequence). 840 amino acids per molecule, of which 90 are cysteine residues.[9]
The optimum pH is 7.4 and optimum temperature is 60 °C. Substrates include urea and hydroxyurea.
Bacterial ureases are composed of three distinct subunits, one large catalytic (α 60–76kDa) and two small (β 8–21 kDa, γ 6–14 kDa) commonly forming (αβγ)3 trimers stoichiometry with a 2-fold symmetric structure (note that the image above gives the structure of the asymmetric unit, one-third of the true biological assembly), they are cysteine-rich enzymes, resulting in the enzyme molar masses between 190 and 300kDa.[9]
An exceptional urease is obtained from Helicobacter sp.. These are composed of two subunits, α(26–31 kDa)-β(61–66 kDa). These subunits form a supramolecular (αβ)12 dodecameric complex.[10] of repeating α-β subunits, each coupled pair of subunits has an active site, for a total of 12 active sites.[10] It plays an essential function for survival, neutralizing gastric acid by allowing urea to enter into periplasm via a proton-gated urea channel.[11] The presence of urease is used in the diagnosis of Helicobacter species.
All bacterial ureases are solely cytoplasmic, except for those in Helicobacter pylori, which along with its cytoplasmic activity, has external activity with host cells. In contrast, all plant ureases are cytoplasmic.[9]
Fungal and plant ureases are made up of identical subunits (90 kDa each), most commonly assembled as trimers and hexamers. For example, jack bean urease has two structural and one catalytic subunit. The α subunit contains the active site, it is composed of 840 amino acids per molecule (90 cysteines), its molecular mass without Ni(II) ions amounting to 90.77 kDa. The mass of the hexamer with the 12 nickel ions is 545.34 kDa. Other examples of homohexameric structures of plant ureases are those of soybean, pigeon pea and cotton seeds enzymes.[9]
It is important to note, that although composed of different types of subunits, ureases from different sources extending from bacteria to plants and fungi exhibit high homology of amino acid sequences. The single plant urease chain is equivalent to a fused γ-β-α organization. The Helicobacter "α" is equivalent to a fusion of the normal bacterial γ-β subunits, while its "β" subunit is equivalent to the normal bacterial α.[9] The three-chain organization is likely ancestral.[12]