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Urease
Urease
Urease
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3D model of urease from Klebsiella aerogenes, two Ni2+-ions are shown as green spheres.[1]
Identifiers
EC no.3.5.1.5
CAS no.9002-13-5
Databases
IntEnzIntEnz view
BRENDABRENDA entry
ExPASyNiceZyme view
KEGGKEGG entry
MetaCycmetabolic pathway
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Ureases (EC 3.5.1.5), functionally, belong to the superfamily of amidohydrolases and phosphotriesterases.[2] Ureases are found in numerous Bacteria, Archaea, fungi, algae, plants, and some invertebrates. Ureases are nickel-containing metalloenzymes of high molecular weight.[3] Ureases are important in degrading avian faecal matter, which is rich in uric acid, the breakdown product of which is urea, which is then degraded by urease as described here.

These enzymes catalyze the hydrolysis of urea into carbon dioxide and ammonia:

(NH2)2CO + H2O urease→ CO2 + 2NH3

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.

History

[edit]

Urease 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]

Importance

[edit]

Urease is important because of its role in the nitrogen cycle as a key catalyst in the reaction converting urea to ammonium and CO2. Urease occurs as a soil enzyme, likely because soil microorganisms benefit from the nitrogen made available by urea degradation in the form of ammonium.[7]

Structure

[edit]

A 1984 study focusing on urease from jack bean found that the active site contains a pair of nickel centers.[8] In vitro activation also has been achieved with manganese and cobalt in place of nickel.[9] 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.[10]

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.[10]

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.[11] of repeating α-β subunits, each coupled pair of subunits has an active site, for a total of 12 active sites.[11] It plays an essential function for survival, neutralizing gastric acid by allowing urea to enter into periplasm via a proton-gated urea channel.[12] 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.[10]

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.[10]

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.[10] The three-chain organization is likely ancestral.[13]

Activity

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The kcat/Km of urease in the processing of urea is 1014 times greater than the rate of the uncatalyzed elimination reaction of urea.[5] There are many reasons for this observation in nature. The proximity of urea to active groups in the active site along with the correct orientation of urea allow hydrolysis to occur rapidly. Urea alone is very stable due to the resonance forms it can adopt. The stability of urea is understood to be due to its resonance energy, which has been estimated at 30–40 kcal/mol.[5] This is because the zwitterionic resonance forms all donate electrons to the carbonyl carbon making it less of an electrophile making it less reactive to nucleophilic attack.[5]

Active site

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The active site of ureases is located in the α (alpha) subunits. It is a bis-μ-hydroxo dimeric nickel center, with an interatomic distance of ~3.5 Å.[5] > The Ni(II) pair are weakly antiferromagnetically coupled.[14] X-ray absorption spectroscopy (XAS) studies of Canavalia ensiformis (jack bean), Klebsiella aerogenes and Sporosarcina pasteurii (formerly known as Bacillus pasteurii)[15] confirm 5–6 coordinate nickel ions with exclusively O/N ligation, including two imidazole ligands per nickel.[9] Urea substrate is proposed to displace aquo ligands.

Water molecules located towards the opening of the active site form a tetrahedral cluster that fills the cavity site through hydrogen bonds. Some amino acid residues are proposed to form mobile flap of the site, which gate for the substrate.[3] Cysteine residues are common in the flap region of the enzymes, which have been determined not to be essential in catalysis, although involved in positioning other key residues in the active site appropriately.[16] In Sporosarcina pasteurii urease, the flap was found in the open conformation, while its closed conformation is apparently needed for the reaction.[15]

When compared, the α subunits of Helicobacter pylori urease and other bacterial ureases align with the jack bean ureases.[16]

The binding of urea to the active site of urease has not been observed.[10]

Proposed mechanisms

[edit]

Blakeley/Zerner

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One mechanism for the catalysis of this reaction by urease was proposed by Blakely and Zerner.[17] It begins with a nucleophilic attack by the carbonyl oxygen of the urea molecule onto the 5-coordinate Ni (Ni-1). A weakly coordinated water ligand is displaced in its place. A lone pair of electrons from one of the nitrogen atoms on the Urea molecule creates a double bond with the central carbon, and the resulting NH2− of the coordinated substrate interacts with a nearby positively charged group. Blakeley and Zerner proposed this nearby group to be a Carboxylate ion, although deprotonated carboxylates are negatively charged.

A hydroxide ligand on the six coordinate Ni is deprotonated by a base. The carbonyl carbon is subsequently attacked by the electronegative oxygen. A pair of electrons from the nitrogen-carbon double bond returns to the nitrogen and neutralizes the charge on it, while the now 4-coordinate carbon assumes an intermediate tetrahedral orientation.

The breakdown of this intermediate is then helped by a sulfhydryl group of a cysteine located near the active site. A hydrogen bonds to one of the nitrogen atoms, breaking its bond with carbon, and releasing an NH3 molecule. Simultaneously, the bond between the oxygen and the 6-coordinate nickel is broken. This leaves a carbamate ion coordinated to the 5-coordinate Ni, which is then displaced by a water molecule, regenerating the enzyme.

The carbamate produced then spontaneously degrades to produce another ammonia and carbonic acid.[18]

Hausinger/Karplus

[edit]

The mechanism proposed by Hausinger and Karplus attempts to revise some of the issues apparent in the Blakely and Zerner pathway, and focuses on the positions of the side chains making up the urea-binding pocket.[5] From the crystal structures from K. aerogenes urease, it was argued that the general base used in the Blakely mechanism, His320, was too far away from the Ni2-bound water to deprotonate in order to form the attacking hydroxide moiety. In addition, the general acidic ligand required to protonate the urea nitrogen was not identified.[19] Hausinger and Karplus suggests a reverse protonation scheme, where a protonated form of the His320 ligand plays the role of the general acid and the Ni2-bound water is already in the deprotonated state.[5] The mechanism follows the same path, with the general base omitted (as there is no more need for it) and His320 donating its proton to form the ammonia molecule, which is then released from the enzyme. While the majority of the His320 ligands and bound water will not be in their active forms (protonated and deprotonated, respectively,) it was calculated that approximately 0.3% of total urease enzyme would be active at any one time.[5] While logically, this would imply that the enzyme is not very efficient, contrary to established knowledge, usage of the reverse protonation scheme provides an advantage in increased reactivity for the active form, balancing out the disadvantage.[5] Placing the His320 ligand as an essential component in the mechanism also takes into account the mobile flap region of the enzyme. As this histidine ligand is part of the mobile flap, binding of the urea substrate for catalysis closes this flap over the active site and with the addition of the hydrogen bonding pattern to urea from other ligands in the pocket, speaks to the selectivity of the urease enzyme for urea.[5]

Ciurli/Mangani

[edit]

The mechanism proposed by Ciurli and Mangani[20] is one of the more recent and currently accepted views of the mechanism of urease and is based primarily on the different roles of the two nickel ions in the active site.[15] One of which binds and activates urea, the other nickel ion binds and activates the nucleophilic water molecule.[15] With regards to this proposal, urea enters the active site cavity when the mobile 'flap' (which allows for the entrance of urea into the active site) is open. Stability of the binding of urea to the active site is achieved via a hydrogen-bonding network, orienting the substrate into the catalytic cavity.[15] Urea binds to the five-coordinated nickel (Ni1) with the carbonyl oxygen atom. It approaches the six-coordinated nickel (Ni2) with one of its amino groups and subsequently bridges the two nickel centers.[15] The binding of the urea carbonyl oxygen atom to Ni1 is stabilized through the protonation state of Hisα222 Nԑ. Additionally, the conformational change from the open to closed state of the mobile flap generates a rearrangement of Alaα222 carbonyl group in such a way that its oxygen atom points to Ni2.[15] The Alaα170 and Alaα366 are now oriented in a way that their carbonyl groups act as hydrogen-bond acceptors towards NH2 group of urea, thus aiding its binding to Ni2.[15] Urea is a very poor chelating ligand due to low Lewis base character of its NH2 groups. However the carbonyl oxygens of Alaα170 and Alaα366 enhance the basicity of the NH2 groups and allow for binding to Ni2.[15] Therefore, in this proposed mechanism, the positioning of urea in the active site is induced by the structural features of the active site residues which are positioned to act as hydrogen-bond donors in the vicinity of Ni1 and as acceptors in the vicinity of Ni2.[15] The main structural difference between the Ciurli/Mangani mechanism and the other two is that it incorporates a nitrogen, oxygen bridging urea that is attacked by a bridging hydroxide.[18]

Action in pathogenesis

[edit]

Bacterial ureases are often the mode of pathogenesis for many medical conditions. They are associated with hepatic encephalopathy / Hepatic coma, infection stones, and peptic ulceration.[21]

Infection stones

[edit]

Infection induced urinary stones are a mixture of struvite (MgNH4PO4•6H2O) and carbonate apatite [Ca10(PO4)6•CO3].[21] These polyvalent ions are soluble but become insoluble when ammonia is produced from microbial urease during urea hydrolysis, as this increases the surrounding environments pH from roughly 6.5 to 9.[21] The resultant alkalinization results in stone crystallization.[21] In humans the microbial urease, Proteus mirabilis, is the most common in infection induced urinary stones.[22]

Urease in hepatic encephalopathy / hepatic coma

[edit]

Studies have shown that Helicobacter pylori along with cirrhosis of the liver cause hepatic encephalopathy and hepatic coma.[23] Helicobacter pylori release microbial ureases into the stomach. The urease hydrolyzes urea to produce ammonia and carbonic acid. As the bacteria are localized to the stomach ammonia produced is readily taken up by the circulatory system from the gastric lumen.[23] This results in elevated ammonia levels in the blood, a condition known as hyperammonemia; eradication of Helicobacter pylori show marked decreases in ammonia levels.[23]

Urease in peptic ulcers

[edit]

Helicobacter pylori is also the cause of peptic ulcers with its manifestation in 55–68% reported cases.[24] This was confirmed by decreased ulcer bleeding and ulcer reoccurrence after eradication of the pathogen.[24] In the stomach there is an increase in pH of the mucosal lining as a result of urea hydrolysis, which prevents movement of hydrogen ions between gastric glands and gastric lumen.[21] In addition, the high ammonia concentrations have an effect on intercellular tight junctions increasing permeability and also disrupting the gastric mucous membrane of the stomach.[21][25]

Occurrence and applications in agriculture

[edit]

Urea is found naturally in the environment and is also artificially introduced, comprising more than half of all synthetic nitrogen fertilizers used globally.[26] Heavy use of urea is thought to promote eutrophication, despite the observation that urea is rapidly transformed by microbial ureases, and thus usually does not persist.[27] Environmental urease activity is often measured as an indicator of the health of microbial communities. In the absence of plants, urease activity in soil is generally attributed to heterotrophic microorganisms, although it has been demonstrated that some chemoautotrophic ammonium oxidizing bacteria are capable of growth on urea as a sole source of carbon, nitrogen, and energy.[28]

Inhibition in fertilizers

[edit]
Further information: Controlled release fertilizer and Ammonia volatilization from urea

The inhibition of urease is a significant goal in agriculture because the rapid breakdown of urea-based fertilizers is wasteful and environmentally damaging.[29] Phenyl phosphorodiamidate and N-(n-butyl)thiophosphoric triamide are two such inhibitors.[30]

Biomineralization

[edit]

By promoting the formation of calcium carbonate, ureases are potentially useful for biomineralization-inspired processes.[31] Notably, microbiologically induced formation of calcium carbonate can be used in making bioconcrete.[32]

Non-enzymatic action

[edit]

In addition to acting as an enzyme, some ureases (especially plant ones) have additional effects that persist even when the catalytic function is disabled. These include entomotoxicity, inhibition of fungi, neurotoxicity in mammals, promotion of endocytosis and inflammatory eicosanoid production in mammals, and induction of chemotaxis in bacteria. These activities may be part of a defense mechanism.[13]

Urease insect-toxicity was originally noted in canatoxin, an orthologous isoform of jack bean urease. Digestion of the peptide identified a 10-kDa portion most responsible for this effect, termed jaburetox. An analogous portion from the soybean urease is named soyuretox. Studies on insects show that the entire protein is toxic without needing any digestion, however. Nevertheless, the "uretox" peptides, being more concentrated in toxicity, show promise as biopesticides.[13]

As diagnostic test

[edit]
See also: Rapid urease test

Many gastrointestinal or urinary tract pathogens produce urease, enabling the detection of urease to be used as a diagnostic to detect presence of pathogens.

Urease-positive pathogens include:

Ligands

[edit]

Inhibitors

[edit]

A wide range of urease inhibitors of different structural families are known. Inhibition of urease is not only of interest to agriculture, but also to medicine as pathogens like H. pylori produce urease as a survival mechanism. Known structural classes of inhibitors include:[34][35]

  • Analogues of urea, the strongest being thioureas like 1-(4-chlorophenyl)-3-palmitoylthiourea.
  • Phosphoramidates, the most commonly used in agriculture (see above).
  • Hydroquinone and quinones. In medicine, the most interesting are quinolones, already a class of widely used antibiotics.
  • Some plant metabolites are also urease inhibitors, an example being allicin. These have potential both as environmentally-friendly fertilizer additives[36] and natural drugs.

Extraction

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First isolated as a crystal in 1926 by Sumner, using acetone solvation and centrifuging.[37] Modern biochemistry has increased its demand for urease. Jack bean meal,[38] watermelon seeds,[39] and pea seeds[40] have all proven useful sources of urease.

See also

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References

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[edit]
Revisions and contributorsEdit on WikipediaRead on Wikipedia

Urease

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Urease (EC 3.5.1.5), also known as urea amidohydrolase, is a nickel-dependent metalloenzyme that catalyzes the of into and , with the spontaneously decomposing into a second molecule of and . This reaction, which proceeds at a rate accelerated by at least 10^14 times compared to the uncatalyzed process, plays a crucial role in and recycling across diverse organisms. Urease is widely distributed in prokaryotes, fungi, , and plants, but absent in animals, and its activity is essential for processes such as soil cycling and microbial . Structurally, urease is typically oligomeric, featuring a binuclear active site where two Ni(II) ions, separated by 3.5–3.7 , are coordinated by , aspartate, and a carbamylated residue to facilitate . The enzyme's quaternary assembly varies by source: bacterial ureases often form trimers ([αβγ]3) with three active sites, while ureases like that from jack bean (Canavalia ensiformis) are hexameric ((α3)2) with a molecular weight of approximately 540–590 kDa and subunits of 90–91 kDa. The catalytic mechanism involves a bridging from the cluster acting as a to attack urea's carbonyl carbon, forming a tetrahedral intermediate that collapses to release products, with a flexible "flap" domain regulating substrate access. First isolated and crystallized from jack bean in 1926 by James B. Sumner, who received the in 1946 for this achievement, urease represents one of the earliest enzymes to be purified and structurally characterized. The role of as an essential cofactor was established in 1975, resolving earlier debates about its metal dependency. Beyond , urease exhibits non-enzymatic functions, including insecticidal and fungitoxic activities in plants, as well as pro-inflammatory effects in bacterial infections. Urease holds significant biomedical and agricultural importance; for instance, in , it enables survival in the acidic gastric environment by neutralizing pH through , making it a target for anti-ulcer therapies. In agriculture, microbial ureases contribute to fertilizer efficiency but can lead to volatilization losses, prompting research into inhibitors. Recent structural studies, including high-resolution crystal structures from 2019–2020, continue to refine understanding of its mechanism and potential for inhibitor design.

History

Discovery and early studies

The isolation of urea from human urine in 1773 by French chemist Hilaire-Marin Rouelle marked the initial step toward understanding urea's role in biological processes, though the compound itself was not yet linked to enzymatic decomposition. Rouelle obtained urea crystals by evaporating urine and purifying the residue with alcohol, providing the first pure sample for further chemical analysis. In the late 18th century, Antoine François de Fourcroy and conducted key experiments on urine , recognizing in 1798 that the observed in urine resulted from the of . Their work on products, including the identification of as a primary urinary constituent in crystalline form around 1799, highlighted the instability of under certain conditions but did not yet attribute the process to a specific . These early 19th-century investigations laid the foundation for exploring urea's breakdown, though researchers initially attributed the reaction to spontaneous chemical or microbial activity rather than a dedicated catalyst. The enzymatic nature of urea hydrolysis remained unclear until the late , when confusion with non-enzymatic or bacterial mechanisms persisted in scientific discourse. In 1874, Frédéric Musculus isolated a ureolytic principle from putrid , demonstrating its activity independent of viable microbes and suggesting an enzymatic basis. This was formalized in 1890 when Henri Miquel proposed the name "urease" for the agent responsible. However, definitive proof came in the 1920s, culminating in 1926 when James B. Sumner isolated and crystallized urease from jack bean (Canavalia ensiformis) meal, confirming its proteinaceous nature and high catalytic purity. Sumner's breakthrough, which resolved lingering doubts about enzyme composition, earned him the 1946 , shared with John H. Northrop and Wendell M. Stanley for establishing that enzymes are proteins.

Isolation and biochemical characterization

In the , significant advancements were made in the isolation and purification of urease from jack meal, building on James B. Sumner's initial in 1926. Early methods involved extracting the from finely powdered, fat-free jack meal using a 31.6% (v/v) acetone-water solution, followed by filtration and allowing the extract to stand overnight at low temperature (around 4°C) to induce . was commonly employed as a step to concentrate the from aqueous extracts, enabling higher yields and purity before recrystallization. These techniques, refined through iterative experiments, yielded diamond-shaped crystals that confirmed urease's proteinaceous and facilitated biochemical studies. By the mid-20th century, detailed characterization of urease's oligomeric structure emerged, particularly for bacterial forms. Studies on enzymes from such as Sporosarcina pasteurii (formerly Bacillus pasteurii) revealed a native molecular weight of approximately 480–540 kDa, determined via ultracentrifugation and gel filtration chromatography. These bacterial ureases were found to assemble as hexamers, typically with a (αβγ)₃ subunit composition where the α subunit is catalytic (∼60–70 kDa), β is accessory (∼10–20 kDa), and γ is structural (∼10 kDa), contrasting with the homohexameric plant forms. A pivotal discovery in 1975 identified nickel as an essential cofactor for urease activity through atomic absorption spectroscopy and metal reconstitution experiments on jack bean urease. Dixon and colleagues demonstrated that the enzyme contains two nickel ions per active site, with apo-urease (nickel-depleted) showing negligible activity that was restored only upon nickel addition, establishing nickel's biological role in enzyme function. Early kinetic analyses in the mid-20th century confirmed that urease follows Michaelis-Menten kinetics for urea hydrolysis, with the Michaelis constant (Kₘ) for urea reported around 20–50 mM in bacterial systems like S. pasteurii for purified enzyme, reflecting the enzyme's adaptation to varying substrate concentrations in microbial environments. These studies, often conducted at neutral pH and 25–37°C using spectrophotometric assays for ammonia production, highlighted urease's high catalytic efficiency (k_cat/Kₘ ∼10⁶–10⁷ M⁻¹ s⁻¹) while underscoring substrate inhibition at concentrations above 100 mM.

Biological Significance

Occurrence in organisms

Urease is a -dependent widely distributed across various biological kingdoms, including , , , fungi, and , but it is absent in mammals and higher animals, where degradation occurs primarily through microbial ureases in the gut or environment. In prokaryotes and certain eukaryotes, the exhibits evolutionary conservation, with metallation being a key feature maintained from bacterial and archaeal ancestors to and fungal forms, enabling in diverse ecological niches, including marine environments where archaeal ureases contribute to cycling. Among bacteria, urease is prevalent in numerous species, particularly those involved in environmental nitrogen cycling and pathogenesis, such as Helicobacter pylori, Proteus mirabilis, and Klebsiella pneumoniae, where it facilitates survival in urea-rich habitats. In plants, urease occurs prominently in legumes and seeds, including jack beans (Canavalia ensiformis), soybeans (Glycine max), and watermelon seeds (Citrullus lanatus), contributing to nitrogen mobilization during germination and growth. Urease is also found in fungi, exemplified by , which produces the enzyme for urea utilization in soil and decomposition. In algae, including blue-green algae () and species, urease supports in aquatic environments, though it is less studied compared to terrestrial organisms. While rare in animals beyond microbial symbionts, urease activity in the of certain vertebrates, such as hibernating frogs, derives from bacterial sources rather than host production.

Role in nitrogen metabolism

Urease catalyzes the hydrolysis of urea into and , releasing two molecules of that serve as a key source for assimilation into organic compounds. This is primarily incorporated into through the action of , which combines it with glutamate to form , facilitating recycling within cells. In organisms reliant on urea as a input, such as certain and plants, this process is crucial for maintaining and supporting growth in nitrogen-limited conditions. In , urease is essential for utilizing from or fertilizers, enabling these microbes to thrive in urea-rich environments and convert into bioavailable , thereby preventing loss through unhydrolyzed accumulation. Ureolytic , such as Sporosarcina pasteurii, dominate this process, accounting for a substantial portion of urease activity derived from microbial sources. In plants, particularly , urease plays a vital role in mobilizing stored during seed by hydrolyzing ureides—nitrogen-rich compounds accumulated in seeds—and arginine-derived , providing for early growth stages. Inhibition of urease in species like delays by up to 7–8 hours due to impaired release, underscoring its importance in this metabolic transition. Microbial ureases contribute significantly to the global by processing urea-based fertilizers, which represent the most widely used nitrogen input in , generating substantial that supports crop productivity and . This activity integrates into broader nitrogen transformations, enhancing nitrogen retention. The release of ammonia by urease elevates local in soils and microbial habitats, altering environmental conditions that influence the composition and activity of surrounding microbial communities. This shift can favor ureolytic while inhibiting others sensitive to , thereby shaping nitrogen-cycling dynamics in ecosystems.

Molecular Structure

Overall protein architecture

Urease enzymes exhibit a conserved structure characterized by a hexameric assembly, often denoted as (α₆), particularly in bacterial where the enzyme is composed of three distinct subunit types: a large catalytic α-subunit of approximately 60-76 kDa, along with smaller β- and γ-subunits forming the (αβγ)₃ . In and fungi, urease typically assembles as a homohexamer or homotrimer of identical ~90 kDa subunits, though variations exist such as trimeric forms in some . This oligomeric organization stabilizes the and facilitates cooperative interactions among active sites. The three-dimensional structure of urease was first elucidated in the 1990s through , with the enzyme resolved at 2.2 resolution, revealing a complex architecture comprising four domains per α-subunit: two α/β and two β-sheet domains that support the central (α/β)₈ barrel housing the . Subsequent structures, including the jack bean (Canavalia ensiformis) urease at 2.05 resolution in 2010, confirmed similar domain organization in plant forms, highlighting evolutionary conservation despite differences in subunit composition. A notable feature of the urease is the presence of flexible flaps, typically consisting of a motif, that cover the and regulate substrate access by undergoing conformational shifts. These mobile elements ensure controlled entry of while preventing premature of reaction intermediates. Sequence across urease enzymes from , , and fungi reveals conserved core domains essential for folding and assembly, with overall identity ranging from 20-50% between distant species, underscoring a common evolutionary origin. Recent cryo-EM studies in the 2020s have provided high-resolution insights (e.g., 2.0 Å for urease) into dynamic conformational changes within the oligomeric assembly, demonstrating flexibility in inter-subunit interfaces and flap regions that influence enzyme activation and stability. These findings complement earlier crystallographic data by capturing transient states relevant to physiological function. In 2025, a cryo-EM structure of urease at high resolution further revealed wide-open flap conformations in bacterial variants, supported by simulations showing enhanced flap mobility.

Active site and metal cofactors

The of urease harbors a bimetallic center composed of two Ni(II) ions, Ni1 and Ni2, which are bridged by the moiety of a post-translationally modified residue. In jack urease (Canavalia ensiformis), this bridging residue is Lys490, whose ε-amino group undergoes carbamylation to form the stabilizing . This modification occurs via a reaction involving CO₂ and , essential for active site maturation and cluster integrity. The coordination environment of the dinickel center involves key amino acid residues that position the metals for catalysis. Ni1 is ligated by the imidazole nitrogens of His519 (Nδ1) and His545 (Nε1), as well as one oxygen from the carbamylated Lys490 (Oδ1), resulting in a pseudotetrahedral geometry. Ni2, in contrast, coordinates to the imidazole nitrogens of His407 (Nε2) and His409 (Nε2), the carboxylate oxygen of Asp633 (Oδ1), and the second oxygen from Lys490 (Oδ2), adopting a distorted square-pyramidal arrangement with an apical ligand position often occupied by a water molecule or hydroxide. Alanine residues near the active site cleft contribute to the hydrophobic pocket surrounding the metals, though they do not directly ligate the nickels. This architecture is conserved across ureases, with the active site nestled within the α-subunit of the overall hexameric protein scaffold. Spectroscopic studies have corroborated the structural features of the dinickel center. (EPR) spectroscopy confirms the Ni(II) for both ions, consistent with their d⁸ electronic configuration and lack of antiferromagnetic . (EXAFS) analysis reveals a Ni-Ni distance of approximately 3.5 in jack bean urease, aligning with the bridged observed crystallographically and supporting the close proximity required for cooperative substrate binding. Recent structural studies, including high-resolution cryo-EM from , demonstrate that flexible flap regions (residues 312–355 in bacterial homologs and analogous segments in plant ureases) undergo opening-closing motions that widen the entrance channel, facilitating urea access to the buried dinickel center while maintaining overall stability. Such flap dynamics underscore the enzyme's adaptability without altering the core coordination.

Catalytic Activity

Substrate hydrolysis reaction

Urease catalyzes the of in a highly that converts and into two molecules of and , as represented by the equation: (NH2)2CO+H2O→2NH3+CO2(NH_2)_2CO + H_2O \rightarrow 2 NH_3 + CO_2 This process is spontaneous under standard biological conditions, with a standard change (ΔG°_{298}) of approximately -14 kJ/mol, indicating thermodynamic favorability. The reaction proceeds via initial binding of to the , where the two ions coordinate the substrate, followed by the release of the first molecule and subsequent formation and release of ; the then undergoes non-enzymatic to yield the second and . The exhibits optimal activity in a range of 7 to 9, depending on the source organism, with jack bean urease peaking around 7.0 and some bacterial variants functioning effectively up to 9.0. optima vary similarly, typically between 37°C for mammalian-associated forms and 60°C for plant-derived urease, though the enzyme undergoes inactivation above 70°C across species. Kinetic parameters underscore urease's exceptional efficiency, with a (k_{cat}) reaching up to 10^4 s^{-1} for jack bean urease, positioning it among the fastest known enzymes; the for the catalyzed reaction is approximately 30 kJ/mol, a dramatic reduction from the uncatalyzed value of about 125 kJ/mol. In certain bacterial ureases, maturation and activity are influenced by accessory proteins such as UreG and UreE, which act as chaperones to facilitate ion insertion into the , enabling full catalytic competence.

Proposed mechanisms

The proposed mechanisms for urease have evolved with advances in and computational modeling, focusing on the roles of the dinuclear center and surrounding residues in facilitating urea . Early models emphasized the activation of by ions, while later proposals incorporated the bridging and substrate interactions observed in structures. In the , Blakeley, Dixon, and Zerner proposed a mechanism in which a nickel-coordinated molecule acts as the , attacking the carbonyl carbon of bound to the other nickel ion, with a carbamylated residue stabilizing the developing negative charge on the intermediate. This model, based on kinetic and spectroscopic studies of jack bean urease, highlighted the essential role of in polarizing the substrate but was later criticized for overlooking the bridging ligand between the nickel ions, which structural data revealed as crucial for . Building on initial structures in the 1990s and 2000s, Hausinger and Karplus refined the pathway to involve a bridging serving as the that attacks the carbonyl, forming a tetrahedral intermediate stabilized by the ions and nearby residues such as His219, which polarizes the carbonyl. studies of variants, including substitutions at Cys319, His320, and Asp363, supported this model by demonstrating impaired activity and altered dependence, consistent with the hydroxide's dual role as and general acid to protonate the departing . More recent structural insights from Ciurli and describe a substrate-assisted mechanism in which initially binds monodentately via its carbonyl oxygen to Ni1 in the , positioning one amino group near the bridging for nucleophilic attack and leading to carbamylation of a residue during intermediate formation. This pathway, elucidated through the crystal structure of the Sporosarcina pasteurii urease- complex (PDB: 6QDY), emphasizes the enzyme's flap domain in substrate positioning and is consistent with quantum mechanical calculations showing favorable energetics for the tetrahedral intermediate collapse to release and bound . Across these models, common elements include the initial binding of urea's carbonyl oxygen to Ni1, the release of the first molecule from the tetrahedral intermediate, and subsequent of the Ni1-bound to yield the second and , facilitated by the dinuclear center's ability to stabilize high-energy species. The features residues like and aspartate that assist in these steps. Recent / simulations integrating have further clarified proton shuttling during release, with Asp363 mediating transfer from the bridging to urea's amino group and His323 stabilizing the product state through hydrogen bonding.

Pathogenic Roles

In urinary tract infections

Urease produced by bacteria such as Proteus mirabilis and Klebsiella pneumoniae plays a central role in urinary tract infections (UTIs) by hydrolyzing urea in urine to ammonia and carbon dioxide, rapidly elevating urine pH to levels exceeding 9. This alkalization promotes the precipitation of magnesium ammonium phosphate (struvite, MgNH₄PO₄·6H₂O) and carbonate apatite (Ca₁₀(PO₄)₆·CO₃), forming infection stones that can develop into complex staghorn calculi. These branched stones often fill the renal pelvis and calyces, obstructing urine flow and leading to severe complications like hydronephrosis, recurrent infections, and pyelonephritis. Struvite stones account for approximately 15% of all urinary calculi in the United States and are particularly prevalent in patients with indwelling catheters or neurogenic bladders, where rates are elevated in long-term cases. Nearly all stones (over 90%) are associated with urease-producing pathogens, which colonize the urinary tract and perpetuate through formation on stones and catheters. The ammonia generated is directly toxic to uroepithelial cells, causing and exfoliation that exposes underlying tissue and facilitates deeper bacterial . This damage also enhances bacterial adhesion, particularly via type 1 fimbriae in pathogens like and species, promoting persistent colonization and chronic inflammation. Treatment of urease-associated UTIs is complicated by high rates of antibiotic resistance in producers like Proteus and Klebsiella, with multidrug-resistant strains reported in clinical isolates from stone formers. Complete stone removal via percutaneous nephrolithotomy is essential, but recurrence rates exceed 40% without addressing the underlying infection. Recent research has explored urease inhibitors, such as acetohydroxamic acid, which reduce stone growth in high-risk patients; a 2024 study on flavonoid fractions from selected plants demonstrated significant in vitro inhibition of Proteus vulgaris urease activity.

In gastrointestinal and hepatic diseases

Urease produced by plays a critical role in gastrointestinal diseases by enabling the bacterium to neutralize the acidic environment of the , facilitating its colonization of the . The enzyme hydrolyzes into and , creating a protective "ammonia cloud" around the that raises the local from approximately 2 to 5-7, allowing survival and adherence to epithelial cells despite the hostile gastric conditions. This acid neutralization is essential for initiating infection, leading to chronic and peptic ulcers as the bacteria burrow into the mucosal layer and provoke persistent inflammation. In , H. pylori urease not only supports colonization but also contributes to through immune-mediated . The enzyme acts as a potent , eliciting a strong humoral response including secretory IgA antibodies that target urease on the bacterial surface, though this often fails to clear the infection and instead promotes chronic mucosal damage. Urease-induced immune activation recruits inflammatory cells, exacerbating tissue injury and ulcer formation. Globally, H. pylori infects about 50% of the , with urease recognized as a key driving these outcomes; diagnostic tools like the exploit this activity by detecting labeled produced from urease-mediated in infected individuals. Beyond direct gastric effects, urease from gut bacteria contributes to hepatic diseases, particularly in patients with . In the colon, bacterial ureases hydrolyze to , which is absorbed into the bloodstream; impaired liver function in fails to detoxify this excess , leading to that crosses the blood-brain barrier and induces , manifesting as , , and other encephalopathic symptoms. This overproduction is a major pathogenic driver, with altered in cirrhotic patients amplifying urease activity and worsening outcomes. A phase III conducted in 2015 on an oral recombinant H. pylori , including urease B subunit formulations, demonstrated 71.8% protection efficacy in preventing infection in children at one year.

Applications and Uses

In agriculture and soil science

Urease enzymes in , primarily produced by microorganisms, play a central role in dynamics by hydrolyzing applied fertilizers into and , which can lead to rapid volatilization losses ranging from 10% to 36% of the applied urea-, depending on conditions and application methods. This process not only reduces availability for crops but also contributes to environmental , as excess can be oxidized to , promoting leaching into and surface waters, where it exacerbates and water quality degradation. To mitigate these losses, urease inhibitors such as N-(n-butyl)thiophosphoric triamide (NBPT) are applied to urea-based fertilizers, temporarily blocking the enzyme's and slowing , which reduces volatilization by up to 53% and enhances use efficiency by approximately 25% through improved nitrogen recovery in crops like . In practical , this translates to higher crop yields and reduced inputs, with studies showing yield increases of around 6% across various when NBPT is used. Certain soil bacteria, including , can utilize as a source, but studies show variable effects on nodulation and growth in urea-supplemented conditions; high urea levels may inhibit . Soil urease activity also contributes to (N2O) emissions, a potent , as rapid urea breakdown increases substrate availability for nitrifying microbes; recent 2024 studies demonstrate that urease inhibitors can reduce these emissions by about 14%, thereby lowering overall greenhouse gas impacts from fertilized soils. Urease is widespread among microbes, including species like Bacillus sphaericus, which exhibit ureolytic activity and adapt to amendments by upregulating production in response to elevated substrate levels, thereby influencing local cycling and microbial community shifts in amended soils.

In diagnostics and

Urease plays a central role in non-invasive diagnostics for infections, which are associated with peptic ulcers and gastric diseases. In the , patients ingest a solution containing 13C- or 14C-labeled , which H. pylori urease hydrolyzes to produce labeled detectable in exhaled breath samples, typically within 10-30 minutes post-ingestion. This method achieves high sensitivity of approximately 96% and specificity of 93%, making it a preferred first-line diagnostic tool for confirming active infection without the need for . Another key diagnostic application is the , also known as the CLO (Campylobacter-like organism) test, which involves placing gastric samples obtained during into a medium containing and a . Urease from H. pylori in the tissue rapidly hydrolyzes , causing a color change from yellow to pink or red within minutes to hours, indicating and aiding in the diagnosis of ulcers. This test offers quick results with often exceeding 90%, though performance can vary based on bacterial load and site. In , urease is widely immobilized on surfaces or to develop biosensors for precise quantification in clinical settings, such as monitoring blood levels in patients undergoing dialysis or assessing function. These amperometric or potentiometric biosensors detect produced by urease-catalyzed hydrolysis, with limits of detection around 0.1 mM and linear ranges up to 15 mM, enabling real-time analysis in small sample volumes. Such devices improve dialysis efficiency by alerting to accumulation, reducing treatment times and complications. Urease also supports sustainable wastewater treatment through engineered microbial systems that express high levels of the enzyme to hydrolyze urea into ammonia, facilitating its recovery as a fertilizer precursor via processes like membrane separation or electrochemical capture. Bacteria such as Proteus vulgaris, naturally rich in urease, or genetically modified strains are integrated into bioreactors to treat urea-laden effluents from industrial or agricultural sources, achieving near-complete hydrolysis and ammonia yields over 90% under optimized conditions. This approach mitigates environmental nitrogen pollution while valorizing waste. Recent advances in have introduced urease-based nanobiosensors for real-time of in water bodies, leveraging nanostructures like nanobipyramids or paper-based lateral flow strips functionalized with immobilized urease for colorimetric or electrochemical detection. These portable devices achieve detection limits as low as 0.1 µM, enabling on-site assessment of urea pollution from fertilizers or effluents, which supports and management. Integration with pH-sensitive etching mechanisms enhances selectivity, marking a shift toward field-deployable tools for proactive environmental .

Inhibitors and Ligands

Types and mechanisms of inhibition

Urease inhibitors are classified based on their binding modes and interactions with the enzyme's , which features two ions essential for . Competitive inhibitors mimic the substrate and directly occupy the , preventing binding. Non-competitive inhibitors bind elsewhere but disrupt function, often by chelating metals. Allosteric inhibitors target regulatory regions like the flexible flap covering the , while metal-based inhibitors leverage coordination chemistry for high potency. Competitive inhibitors, such as hydroxyurea and phenylphosphorodiamidate, structurally resemble and bind directly to the centers in the , thereby blocking substrate access. Hydroxyurea acts as a competitive inhibitor by coordinating with the ions. Phenylphosphorodiamidate similarly functions as a slow-binding competitive inhibitor, forming a tight complex with the (Ki ≈ 94 pM for urease), effectively halting through direct metallocenter occupation. Non-competitive inhibitors, exemplified by acetohydroxamic acid, do not compete directly with but the ions, distorting the active site's geometry and impairing . Acetohydroxamic acid binds reversibly yet tightly to the nickel centers, leading to that reduces velocity independently of substrate concentration. This mechanism alters the coordination environment of the metals, with reported inhibition constants around 4.8 × 10^{-5} M for urease. Allosteric inhibitors modulate urease activity by binding to sites distant from the active center, such as the flap regions that control substrate entry. Flavonoids like target these mobile flap domains. Quercetin's inhibition involves non-competitive binding, with values around 11.2 μM for urease. Metal-based inhibitors, particularly gold(I) complexes developed in recent studies (up to 2022), exhibit exceptional potency through coordination with urease residues. These complexes, such as those featuring ligands, achieve values below 1 μM (e.g., 38 nM for select gold(I) variants) by forming covalent interactions that target and near the , indirectly disrupting coordination and function. Structure-activity relationships among urease inhibitors reveal that hydrogen bonding to key residues like histidine (e.g., His323) and aspartate (e.g., Asp363) significantly enhances binding affinity and inhibitory potency. Inhibitors with hydroxyl or carbonyl groups capable of forming such bonds show improved Ki values, as these interactions stabilize the enzyme-inhibitor complex and complement metal chelation or allosteric effects. For instance, sulfonamide derivatives leverage Asp hydrogen bonding to boost activity against bacterial ureases.

Therapeutic and industrial implications

In medicine, acetohydroxamic acid serves as a key urease inhibitor for treating stones associated with urea-splitting bacterial infections, functioning by reducing urinary levels to prevent stone growth. Clinical trials have demonstrated its efficacy, with stone growth occurring in only 17% of treated patients compared to 46% in groups over 12-24 months, corresponding to a of approximately 60-70%. However, side effects such as (manifesting as tremulousness in up to 28% of patients) and phlebothrombosis often necessitate dose adjustments or discontinuation in 50% of cases. Urease inhibitors also play a role in anti-Helicobacter pylori therapies, where they act as adjuvants to antibiotics like clarithromycin, enhancing bacterial susceptibility by disrupting acid neutralization and biofilm formation. This approach is particularly valuable amid rising clarithromycin resistance, as urease targeting reduces the pathogen's survival in acidic environments. Industrially, urease inhibitors such as N-(n-butyl) thiophosphoric triamide (NBPT) are incorporated into slow-release urea fertilizers to mitigate nitrogen loss through ammonia volatilization, achieving reductions of up to 78% in emissions compared to untreated urea. This enhances nitrogen use efficiency by 6% on average across crops, minimizing environmental pollution and fertilizer costs. Conversely, the enzyme urease itself is harnessed in biomineralization processes for cement production, where it catalyzes urea hydrolysis to precipitate calcium carbonate, strengthening self-healing concrete. Recent preclinical studies (as of 2024) have explored new hydroxamic acid derivatives as potent urease inhibitors for H. pylori, showing promise for improved anti-ulcer therapies. A key challenge in inhibitor development is bacterial resistance, often arising from ureG mutations that impair nickel ion insertion into the urease , thereby evading maturation-targeted therapies.

Extraction and Non-Enzymatic Actions

Purification methods

The purification of urease from natural sources has traditionally relied on classical techniques starting with extraction from jack bean (Canavalia ensiformis) meal. A widely adopted protocol involves initial ammonium sulfate precipitation at 35-45% saturation to fractionate proteins, followed by anion-exchange chromatography on DEAE-Sepharose columns equilibrated with phosphate buffer (pH 7.0). This approach achieves substantial enrichment, with specific activities reaching up to 3000 U/mg protein after combining steps, representing approximately 400-500-fold purification from crude extracts. Alternative classical methods, such as one-step affinity chromatography using epoxy-activated Sepharose 6B linked to urea, have been employed on jack bean extracts, yielding approximately 80% recovery with a specific activity of 500 U/mg and electrophoretic homogeneity. Recombinant production of urease, often from bacterial sources like , facilitates higher yields and scalability through expression in . The urease gene is typically cloned into vectors like pH6HTN His6HaloTag T7 for N-terminal fusion, followed by induction with IPTG in BL21 strains. Purification proceeds via Ni-NTA under native conditions, eluting with gradients, often combined with size-exclusion FPLC for polish. This yields over 200 mg/L culture of purified protein with 98% purity, as confirmed by showing a predominant 45 kDa band. Advanced protocols integrate (FPLC) and (HPLC) for ultra-high purity suitable for . For instance, post-affinity FPLC on Superdex columns refines recombinant urease to >95% purity, enabling and spectroscopic analyses of coordination. Recent implementations, such as ion-exchange HPLC followed by gel filtration, have similarly attained near-homogeneous preparations from bacterial lysates, with purification folds exceeding 100 and recoveries above 45%. Recent advances in recombinant urease production include optimized co-expression of accessory proteins (UreD, UreE, UreF, UreG) to facilitate nickel insertion, improving activation yields for H. pylori urease in E. coli systems as of 2019.

Non-catalytic functions

In Helicobacter pylori, urease functions as an adhesin beyond its enzymatic role, binding to host Lewis^b antigens on gastric epithelial cells in an acid-dependent manner, which facilitates bacterial colonization of the stomach mucosa. This adherence is mediated by specific interactions between the urease protein and diverse Lewis antigens, including Lewis^b, as demonstrated through binding assays with biotinylated urease and immobilized glycoconjugates. Structural analyses reveal that surface-exposed alpha-helices on the urease contribute to this recognition, enhancing the pathogen's ability to persist in the acidic gastric environment. Urease also exhibits moonlighting activities in immune modulation, particularly in H. pylori, where the B subunit binds to the invariant chain CD74 on gastric epithelial cells. This interaction triggers activation and subsequent release of the proinflammatory IL-8, promoting recruitment and chronic ; notably, this effect persists even with enzymatically inactive or heat-denatured urease, confirming its independence from catalytic hydrolysis. Such non-catalytic signaling contributes to the pathogen's evasion of host defenses and persistence during infection.

References

  1. https://pmc.ncbi.nlm.nih.gov/articles/PMC6077230/
  2. https://link.springer.com/article/10.1007/s00775-020-01808-w
  3. https://www.science.org/doi/10.1126/sciadv.adf7790
  4. https://www.sciencedirect.com/science/article/pii/S2090123218300651
  5. https://www.jbc.org/article/S0021-9258%2820%2969970-7/fulltext
  6. https://www.sciencedirect.com/science/article/abs/pii/S1381117709000150
  7. https://www.nobelprize.org/prizes/chemistry/1946/sumner/facts/
  8. https://pubs.acs.org/doi/10.1021/ja00847a045
  9. https://pmc.ncbi.nlm.nih.gov/articles/PMC10467343/
  10. https://pmc.ncbi.nlm.nih.gov/articles/PMC12001548/
  11. https://pmc.ncbi.nlm.nih.gov/articles/PMC7184782/
  12. https://academic.oup.com/femsre/article/27/2-3/239/614619
  13. https://www.pnas.org/doi/10.1073/pnas.1201914109
  14. https://pmc.ncbi.nlm.nih.gov/articles/PMC4263730/
  15. https://pmc.ncbi.nlm.nih.gov/articles/PMC9630907/
  16. https://www.sciencedirect.com/science/article/pii/S0016706123001817
  17. https://www.sciencedirect.com/science/article/abs/pii/S0141813024076074
  18. https://www.sciencedirect.com/science/article/abs/pii/S1878818119303111
  19. https://pubmed.ncbi.nlm.nih.gov/5931453/
  20. https://pubmed.ncbi.nlm.nih.gov/27034039/
  21. https://pmc.ncbi.nlm.nih.gov/articles/PMC5966601/
  22. https://link.springer.com/article/10.1007/s11157-018-9466-1
  23. https://www.intechopen.com/chapters/81535
  24. https://journals.asm.org/doi/pdf/10.1128/aem.42.1.89-96.1981
  25. https://pmc.ncbi.nlm.nih.gov/articles/PMC7433671/
  26. https://pmc.ncbi.nlm.nih.gov/articles/PMC6981326/
  27. http://www.international-agrophysics.org/pdf-139714-67527?filename=Effectiveness%2520of%2520the%2520use.pdf
  28. https://pmc.ncbi.nlm.nih.gov/articles/PMC6077125/
  29. https://www.sciencedirect.com/topics/biochemistry-genetics-and-molecular-biology/urease
  30. https://www.science.org/doi/10.1126/science.7754395
  31. https://pubmed.ncbi.nlm.nih.gov/20471401/
  32. https://www.nature.com/articles/s41467-020-18870-2
  33. https://www.ncbi.nlm.nih.gov/books/NBK2417/
  34. https://www.rcsb.org/structure/9IT2
  35. https://doi.org/10.1016/j.jmb.2010.05.009
  36. https://pubmed.ncbi.nlm.nih.gov/6838612/
  37. http://exonjournal.com/index.php/about/article/download/57/56
  38. https://www.sigmaaldrich.com/deepweb/assets/sigmaaldrich/product/documents/382/636/u1500dat.pdf
  39. https://cdnsciencepub.com/doi/10.1139/cjss-2020-0044
  40. https://www.rcsb.org/structure/6QDY
  41. https://pubs.acs.org/doi/10.1021/acs.jpcb.1c10200
  42. https://www.ncbi.nlm.nih.gov/books/NBK568783/
  43. https://emedicine.medscape.com/article/439127-overview
  44. https://pmc.ncbi.nlm.nih.gov/articles/PMC5905542/
  45. https://pmc.ncbi.nlm.nih.gov/articles/PMC4457377/
  46. https://pmc.ncbi.nlm.nih.gov/articles/PMC2223845/
  47. https://journals.lww.com/jdra/fulltext/2024/09050/inhibition_of_proteus_vulgaris_and_its_associate.4.aspx
  48. https://www.researchgate.net/publication/387283210_Inhibition_of_Proteus_vulgaris_and_its_associate_urease_by_flavonoid_fraction_of_selected_plants_used_as_spices_in_India
  49. https://pmc.ncbi.nlm.nih.gov/articles/PMC135060/
  50. https://www.biorxiv.org/content/10.1101/045328v1.full-text
  51. https://pubmed.ncbi.nlm.nih.gov/8730260/
  52. https://pmc.ncbi.nlm.nih.gov/articles/PMC5851894/
  53. https://pmc.ncbi.nlm.nih.gov/articles/PMC95630/
  54. https://pubmed.ncbi.nlm.nih.gov/1341068/
  55. https://www.ncbi.nlm.nih.gov/books/NBK534233/
  56. https://pubmed.ncbi.nlm.nih.gov/11218379/
  57. https://www.ncbi.nlm.nih.gov/books/NBK542286/
  58. https://pmc.ncbi.nlm.nih.gov/articles/PMC4442863/
  59. https://pmc.ncbi.nlm.nih.gov/articles/PMC10272964/
  60. https://pmc.ncbi.nlm.nih.gov/articles/PMC9050925/
  61. https://pubmed.ncbi.nlm.nih.gov/26142048/
  62. https://www.nature.com/articles/srep43853
  63. https://link.springer.com/article/10.1007/s44279-024-00060-z
  64. https://www.sciencedirect.com/science/article/pii/S2090123218300638
  65. https://pubmed.ncbi.nlm.nih.gov/30094079/
  66. https://pmc.ncbi.nlm.nih.gov/articles/PMC3209853/
  67. https://www.frontiersin.org/journals/sustainable-food-systems/articles/10.3389/fsufs.2024.1460994/full
  68. https://www.frontiersin.org/journals/microbiology/articles/10.3389/fmicb.2018.00634/full
  69. https://onlinelibrary.wiley.com/doi/10.1155/2020/6483803
  70. https://pmc.ncbi.nlm.nih.gov/articles/PMC4306177/
  71. https://www.sciencedirect.com/topics/medicine-and-dentistry/rapid-urease-test
  72. https://ietresearch.onlinelibrary.wiley.com/doi/full/10.1049/nbt2.12050
  73. https://www.sciencedirect.com/science/article/abs/pii/0956566391870338
  74. https://www.researchgate.net/publication/354817298_A_Study_on_the_Hydrolysis_of_Urea_Contained_in_Wastewater_and_Continuous_Recovery_of_Ammonia_by_an_Enzymatic_Membrane_Reactor
  75. https://www.nature.com/articles/s41378-025-00931-5
  76. https://pmc.ncbi.nlm.nih.gov/articles/PMC12562505/
  77. https://pubmed.ncbi.nlm.nih.gov/1864988/
  78. https://pubmed.ncbi.nlm.nih.gov/2674118/
  79. https://www.sciencedirect.com/topics/medicine-and-dentistry/acetohydroxamic-acid
  80. https://www.journalofdairyscience.org/article/S0022-0302%2881%2982624-0/pdf
  81. https://pubmed.ncbi.nlm.nih.gov/23067328/
  82. https://pmc.ncbi.nlm.nih.gov/articles/PMC11003495/
  83. https://arabjchem.org/sulfonamide-derivatives-targeting-urease-structural-diversity-and-therapeutic-potential/
  84. https://ajps.journals.ekb.eg/article_187826_5fc714b93e7b4a05788393e39bb1e7c6.pdf
  85. https://pubmed.ncbi.nlm.nih.gov/6472365/
  86. https://www.karger.com/Article/Pdf/471707
  87. https://pubs.acs.org/doi/10.1021/acsomega.5c02725
  88. https://www.nature.com/articles/s41598-023-50061-z
  89. https://www.nature.com/articles/s44296-023-00004-6
  90. https://pubs.rsc.org/en/content/articlelanding/2024/md/d4md00378k
  91. https://journals.plos.org/plosbiology/article?id=10.1371/journal.pbio.2003887
  92. https://mro.massey.ac.nz/bitstream/10179/3016/1/02_whole.pdf
  93. https://pubmed.ncbi.nlm.nih.gov/3148323/
  94. https://pmc.ncbi.nlm.nih.gov/articles/PMC8838460/
  95. https://www.mdpi.com/2304-6740/7/7/85
  96. https://www.gastrojournal.org/article/S0016-5085(00)71015-8/fulltext
  97. https://pmc.ncbi.nlm.nih.gov/articles/PMC1360328/
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