RESUMEN
Ureases (EC 3.5.1.5) are nickel-dependent metalloenzymes that catalyze the hydrolysis of urea to ammonia and carbon dioxide. Produced by plants, fungi and bacteria, but not by animals, ureases share significant homology and similar mechanisms of catalysis, although differing in quaternary structures. While fungal and plant ureases are homo-oligomeric proteins of 90 kDa subunits, bacterial ureases are multimers of two (e.g. Helicobacter pylori) or three subunit complexes. It has been proposed that in plants these enzymes are involved in nitrogen bioavailability and in protection against pathogens. Previous studies by our group have shown that plant ureases, but not a bacterial (Bacillus pasteurii) urease, display insecticidal activity. Herein we demonstrate that (Glycine max) embryo-specific soybean urease, jackbean (Canavalia ensiformis) major urease and a recombinant H. pylori urease impair growth of selected phytopathogenic fungi at sub-micromolar concentrations. This antifungal property of ureases is not affected by treatment of the proteins with an irreversible inhibitor of the ureolytic activity. Scanning electron microscopy of urease-treated fungi suggests plasmolysis and cell wall injuries. Altogether, our data indicate that ureases probably contribute to the plant arsenal of defense compounds against predators and phytopathogens and that the urease defense mechanism is independent of ammonia release from urea.
Asunto(s)
Antifúngicos/farmacología , Canavalia/enzimología , Glycine max/enzimología , Helicobacter pylori/enzimología , Ureasa/farmacología , Secuencia de Aminoácidos , Proteínas Bacterianas/metabolismo , Proteínas Bacterianas/farmacología , Relación Dosis-Respuesta a Droga , Hongos/efectos de los fármacos , Hongos/ultraestructura , Datos de Secuencia Molecular , Proteínas de Plantas/metabolismo , Proteínas de Plantas/farmacología , Proteínas Recombinantes , Factores de Tiempo , Ureasa/química , Ureasa/metabolismoRESUMEN
Ureases (EC 3.5.1.5) are highly homologous enzymes found in plants, bacteria and fungi. Canatoxin, an isoform Canavalia ensiformis urease, has several biological properties unrelated to its ureolytic activity, like platelet-aggregating and pro-inflammatory effects. Here, we describe that Bacillus pasteurii urease (BPU) also induces aggregation of rabbit platelets, similar to the canatoxin-induced effect (ED(50) 0.4 and 0.015 mg/mL, respectively). BPU induced-aggregation was blocked in platelets pretreated with dexamethasone and esculetin, a phospholipase A(2) and a lipoxygenase inhibitor, respectively, while platelets treated with indomethacin, a cyclooxygenase inhibitor, showed increased response to BPU. Methoxyverapamil (Ca(2+) channel blocker) and AMP (ADP antagonist) abrogated urease-induced aggregation, whereas the PAF-acether antagonist Web2170 had no effect. We concluded that platelet aggregation induced by BPU is mediated by lipoxygenase-derived eicosanoids and secretion of ADP from the platelets through a calcium-dependent mechanism. Potential relevance of these findings for bacterium-plant interactions and pathogenesis of bacterial infections are discussed.
Asunto(s)
Bacillus/enzimología , Plaquetas/fisiología , Fabaceae/enzimología , Proteínas de Plantas/administración & dosificación , Agregación Plaquetaria/fisiología , Ureasa/administración & dosificación , Animales , Plaquetas/efectos de los fármacos , Células Cultivadas , Relación Dosis-Respuesta a Droga , Agregación Plaquetaria/efectos de los fármacos , ConejosRESUMEN
Ureases are enzymes from plants, fungi and bacteria that catalyze the hydrolysis of urea to form ammonia and carbon dioxide. While fungal and plant ureases are homo-oligomers of 90-kDa subunits, bacterial ureases are multimers of two or three subunit complexes. We showed that some isoforms of jack bean urease, canatoxin and the classical urease, bind to glycoconjugates and induce platelet aggregation. Canatoxin also promotes release of histamine from mast cells, insulin from pancreatic cells and neurotransmitters from brain synaptosomes. In vivo it induces rat paw edema and neutrophil chemotaxis. These effects are independent of ureolytic activity and require activation of eicosanoid metabolism and calcium channels. Helicobacter pylori, a Gram-negative bacterium that colonizes the human stomach mucosa, causes gastric ulcers and cancer by a mechanism that is not understood. H. pylori produces factors that damage gastric epithelial cells, such as the vacuolating cytotoxin VacA, the cytotoxin-associated protein CagA, and a urease (up to 10% of bacterial protein) that neutralizes the acidic medium permitting its survival in the stomach. H. pylori whole cells or extracts of its water-soluble proteins promote inflammation, activate neutrophils and induce the release of cytokines. In this paper we review data from the literature suggesting that H. pylori urease displays many of the biological activities observed for jack bean ureases and show that bacterial ureases have a secretagogue effect modulated by eicosanoid metabolites through lipoxygenase pathways. These findings could be relevant to the elucidation of the role of urease in the pathogenesis of the gastrointestinal disease caused by H. pylori.
Asunto(s)
Canavalia/enzimología , Eicosanoides/metabolismo , Infecciones por Helicobacter/microbiología , Helicobacter pylori/enzimología , Proteínas de Plantas/biosíntesis , Toxinas Biológicas/biosíntesis , Ureasa/biosíntesis , Animales , Relación Dosis-Respuesta a Droga , Enfermedades Duodenales/metabolismo , Enfermedades Duodenales/microbiología , Infecciones por Helicobacter/metabolismo , Helicobacter pylori/patogenicidad , Humanos , Proteínas de Plantas/farmacología , Gastropatías/metabolismo , Gastropatías/microbiología , Toxinas Biológicas/farmacologíaRESUMEN
Ureases are enzymes from plants, fungi and bacteria that catalyze the hydrolysis of urea to form ammonia and carbon dioxide. While fungal and plant ureases are homo-oligomers of 90-kDa subunits, bacterial ureases are multimers of two or three subunit complexes. We showed that some isoforms of jack bean urease, canatoxin and the classical urease, bind to glycoconjugates and induce platelet aggregation. Canatoxin also promotes release of histamine from mast cells, insulin from pancreatic cells and neurotransmitters from brain synaptosomes. In vivo it induces rat paw edema and neutrophil chemotaxis. These effects are independent of ureolytic activity and require activation of eicosanoid metabolism and calcium channels. Helicobacter pylori, a Gram-negative bacterium that colonizes the human stomach mucosa, causes gastric ulcers and cancer by a mechanism that is not understood. H. pylori produces factors that damage gastric epithelial cells, such as the vacuolating cytotoxin VacA, the cytotoxin-associated protein CagA, and a urease (up to 10 percent of bacterial protein) that neutralizes the acidic medium permitting its survival in the stomach. H. pylori whole cells or extracts of its water-soluble proteins promote inflammation, activate neutrophils and induce the release of cytokines. In this paper we review data from the literature suggesting that H. pylori urease displays many of the biological activities observed for jack bean ureases and show that bacterial ureases have a secretagogue effect modulated by eicosanoid metabolites through lipoxygenase pathways. These findings could be relevant to the elucidation of the role of urease in the pathogenesis of the gastrointestinal disease caused by H. pylori.