Gene/Protein Disease Symptom Drug Enzyme Compound
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Query: EC:6.3.4.6 (urease)
 7,490 document(s) hit in 31,850,051 MEDLINE articles (0.00 seconds)

The urease proteins of the jack bean (Canavalia ensiformis) and Helicobacter pylori are similar in molecular mass when separated by non-denaturing gradient polyacrylamide gel electrophoresis, both having three main forms. The molecular mass of their major protein form is within the range 440-480 kDa with the other two lesser forms at 230-260 kDa and 660-740 kDa. These forms are all urease active; however, significant kinetic differences exist between the H. pylori and jack bean ureases. Jack bean urease has a single pH optimum at 7.4, whereas H. pylori urease has two pH optima of 4.6 and 8.2 in barbitone and phosphate buffers that were capable of spanning the pH range 3 to 10. The H. pylori Km was 0.6 mM at pH 4.6 and 1.0 mM at pH 8.2 in barbitone buffer, greater than 10.0 mM, and 1.1 mM respectively in phosphate buffer and also greater than 10.0 mM in Tris.HCl at pH 8.2. By comparison, the jack bean urease had a Km of 1.3 mM in Tris.HCl under our experimental conditions. The findings show that the urease activity of H. pylori was inhibited at the pH optimum of 4.6 in the phosphate buffer, but not in the barbitone buffer. This was shown to be due to competitive inhibition by the sodium and potassium ions in the phosphate buffer, not the phosphate ions as suggested earlier. Jack bean urease activity was similarly inhibited by phosphate buffer but again due to the effect of sodium and potassium ions.
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PMID:Kinetic properties of Helicobacter pylori urease compared with jack bean urease. 146 14

Urease was purified 112-fold to homogeneity from the microaerophilic human gastric bacterium, Helicobacter pylori. The urease isolation procedure included a water extraction step, size exclusion chromatography, and anion exchange chromatography. The purified enzyme exhibited a Km of 0.3 +/- 0.1 mM and a Vmax of 1,100 +/- 200 mumols of urea hydrolyzed/min/mg of protein at 22 degrees C in 31 mM Tris-HCl, pH 8.0. The isoelectric point was 5.99 +/- 0.03. Molecular mass estimated for the native enzyme was 380,000 +/- 30,000 daltons, whereas subunit values of 62,000 +/- 2,000 and 30,000 +/- 1,000 were determined. The partial amino-terminal sequence (17 residues) of the large subunit of H. pylori urease (Mr = 62,000) was 76% homologous with an internal sequence of the homohexameric jack bean urease subunit (Mr = 90,770; Takashima, K., Suga, T., and Mamiya, G. (1988) Eur. J. Biochem. 175, 151-165) and was 65% homologous with amino-terminal sequences of the large subunits of heteropolymeric ureases from Proteus mirabilis (Mr = 73,000) and from Klebsiella aerogenes (Mr = 72,000; Mobley, H. L. T., and Hausinger, R. P. (1989) Microbiol. Rev. 53, 85-108). The amino-terminal sequence (20 residues) of the small subunit of H. pylori urease (Mr = 30,000) was 65 and 60% homologous with the amino-terminal sequences of the subunit of jack bean urease and with the Mr = 11,000 subunit of P. mirabilis urease (Jones, B. D., and Mobley, H. L. T. (1989) J. Bacteriol. 171, 6414-6422), respectively. Thus, the urease of H. pylori shows similarities to ureases found in plants and other bacteria. When used as antigens in an enzyme-linked immunosorbent assay, neither purified urease nor an Mr = 54,000 protein that co-purified with urease by size exclusion chromatography was as effective as crude preparations of H. pylori proteins at distinguishing sera from persons known either to be infected with H. pylori or not.
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PMID:Purification and characterization of urease from Helicobacter pylori. 218 75

Apart from urine supersaturation with respect to struvite and calcium phosphate, crystal retention is considered to be necessary for the formation of infection stones. This study was performed to investigate the role of the mucous coat in rat bladders in the adhesion of sterile urease-induced crystals and to determine to what extent the adhesion was influenced by infection. Elimination of the mucous coat with 0.1 M HCl increased the adherence of crystals six times compared to that in bladders with an intact mucous coat. Infection with Proteus mirabilis, Escherichia coli, enterococci and Ureaplasma urealyticum increased the adherence six, five, four and two times, respectively. Injury to the mucous coat may thus be one mechanism by which microorganisms can contribute to the formation of infection stones in the urinary tract.
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PMID:Adherence of urease-induced crystals to rat bladder epithelium following acute infection with different uropathogenic microorganisms. 329 48

Triple therapy has been recommended as the most effective treatment for Helicobacter pylori eradication. Despite achieving a comparatively high eradication result, however, around 10% of patients still fail to be cured. Omeprazole can enhance efficacy of single and double antibiotic protocols and is particularly effective when combined with clarithromycin and a nitroimidazole. This study examined the effect of combining triple therapy with omeprazole. A prospective, randomised, unblinded, single centre trial was carried out on consecutive patients with symptoms of dyspepsia and H pylori infection confirmed by rapid urease test, microbiological culture, and histological assessment. Patients were given a five times/day, 12 day course of colloidal bismuth subcitrate chewable tablets (108 mg), tetracycline HCl (250 mg), and metronidazole (200 mg) with either 20 mg omeprazole twice daily (triple therapy+omeprazole) or 40 mg famotidine (triple therapy+famotidine) at night. Compliance and side effects were determined using a standard questionnaire form. One hundred and twenty five of 165 triple therapy+omeprazole patients and 124 of 171 triple therapy+famotidine patients returned for rebiopsy four weeks after completion of treatment. Significantly more triple therapy+omeprazole patients achieved eradication 122 of 125 (97.6%) as assessed by negative urease test, culture, and histological assessment, when compared with 110 of 124 (89%) triple therapy+famotidine patients (p = 0.006; chi 2). There were 30 triple therapy+omeprazole (24%) and 26 triple therapy+famotidine (21%) patients with de novo metronidazole resistant H pylori included in the study. Side effects were mild and infrequent and were comparable in both groups, although pain in duodenal ulcer, gastric ulcer, and oesophagitis patients seemed to subside earlier in those taking omeprazole. Compliance (>95% of drugs taken) was achieved by 98% of patients of both groups. A 12 days regimen of triple therapy with omeprazole is more effective in achieving H pylori eradication than is triple therapy plus famotidine. Use of 20 mg omeprazole twice daily rather than 40 mg famotidine with a 12 day, low dose triple therapy enhances eradication to over 97% whether the H pylori is metronidazole sensitive or resistant.
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PMID:Omeprazole enhances efficacy of triple therapy in eradicating Helicobacter pylori. 748 31

The effects of urea-urease-ammonia on the rat gastric mucosa were examined and compared with those of NH4OH and NH4Cl. The mucosal application of urea with urease produced a reduction in potential difference (PD) in a dose-related manner for urea, and a significant drop was observed by > 0.1% urea in the presence of 100 units urease. Such PD reduction was also observed when the mucosa was exposed to either NH4OH (> 0.03%) or NH4Cl (> 1%); delta PD (20 mV) caused by 0.3% NH4OH and 3% NH4Cl was equivalent to that induced by 0.5% urea+urease (100 units). The combined oral administration of urea (approximately 6%) and urease (100 units) did not induce any macroscopic damage in the gastric mucosa. NH4Cl given orally had no or little effect on the mucosa at any dose levels even at 10%, while NH4OH given orally caused hemorrhagic lesions in the mucosa at the dose of > 0.3%. In contrast, both urea+urease and NH4Cl given prior to HCl/ethanol protected the gastric mucosa against damage in a dose-related manner, and a significant effect was obtained by urea at > 0.5% and by NH4Cl at > 1%. NH4OH was also effective in reducing the severity of HCl/ethanol-induced gastric lesions at lower dose (0.3%). The protective effect of urea+urease was attenuated significantly by prior administration of indomethacin or coadministration of hydroxyurea, while that of NH4Cl or NH4OH was mitigated by indomethacin.(ABSTRACT TRUNCATED AT 250 WORDS)
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PMID:Irritant and protective action of urea-urease ammonia in rat gastric mucosa. Different effects of ammonia and ammonium ion. 785 Nov 89

We investigated the action of the ammonia produced by Helicobacter pylori urease on the cultured cells. The urease was purified from supernatant fluid of sonicated cell of H. pylori cultured on blood agar for 2 days at 37 degrees C under microaerophilic condition. Purification was carried out by DEAE-Sepharose chromatography, Phenyl-Sepharose chromatography, Sephacryl S-200 SF chromatography and fast protein liquid chromatography on Mono-Q. Vero, HeLa and Intestin 407 cells with or without the addition of 30 mM urea were exposed to the purified urease. Those cells showed cytotoxic effects within 80 minutes after addition of purified urease in the presence of urea. The ammonia production was observed on tissue culture medium within 10 minutes, and the ammonia concentration ranged from 5.56 mg/ml to 7.3 mg/ml and pH in the medium was over pH 9.0. No such effect was observed on the cells exposed to urease without urea. Ammonia water added to Vero cells showed the same cytotoxic effect within 70 minutes on the production of ammonia and raised the pH. However, when the cells were exposed to the ammonia water pre-neutralized to a given pH 7-8 using 1 N HCl cytotoxic effect was not observed. It was concluded that the cytotoxic effect of H. pylori urease was dependent on ammonia generated by hydrolysis of urea.
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PMID:[Cytotoxic effect of ammonia produced by Helicobacter pylori urease on the cultural cells]. 840 83

Protein aggregation is believed to be due to conformers that expose hydrophobic clusters that promote protein association. Such conformers can be detected using a fluorescent probe like 8-anilino 1-naphthalenesulfonic acid (ANSA). Here we show that urease exposed to 1.0 M guanidine-hydrogen chloride has a higher affinity for ANSA that native or denatured urease. The binding occurs over a narrow range of denaturant concentration, well below the concentration required to induce denaturation. The impact of ANSA on urease aggregation was further studied by fluorescence, light-scattering, and activity measurements. We found that ANSA modifies urease aggregates and can provide partial protection against inactivation arising from thermally induced aggregation. It seems that the well-known susceptibility of urease to aggregation is due to an intermediate that can be populated in the absence of denaturation. Such a rationale would explain why folding stability of urease is a poor indicator of long-term stabilization by various media.
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PMID:Detection of an unfolding intermediate in alpha-urease with enhanced affinity for ANSA. 919 74

Bestatin, an aminopeptidase inhibitor, permits the degradation of cellular proteins to di- and tripeptides but interferes with the further breakdown of these peptides to amino acids. We propose to measure instant rates of protein degradation in skeletal muscles of intact mice by the accumulation of bestatin-induced intermediates. Muscle protein was labeled by injection of L-[guanidino-14C]arginine; 3 days later, maximum accumulation of intermediates was measured in abdominal wall muscles 10 min after the intravenous injection of 5 mg of bestatin. The peptides were partially purified and hydrolyzed in 6 N HCl, and the radioactivity in peptide-derived arginine was determined, after conversion to 14CO2 by treatment with arginase and urease. The measurement of bestatin-induced intermediates provides a unique tool for studying acute changes in muscle protein turnover in live mice. We observed a 62% increase in muscle protein breakdown after a 16-h fast, which was reversed by refeeding for 3.5 h, and a 38% increase after 3 days of protein depletion.
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PMID:Measurement of muscle protein degradation in live mice by accumulation of bestatin-induced peptides. 943 31

Hexagonal crystals of urease from Bacillus pasteurii have been obtained by vapour diffusion at 293 K in 20 mM Tris-HCl, neutral pH, containing 50 mM Na2SO3. Isomorphous crystals of urease inhibited with beta-mercaptoethanol were also obtained by including 4 mM of the inhibitor in the enzyme solution. Crystals of the native and inhibited enzyme diffract respectively to 2.00 A (96.7% completeness) and to 1.65 A (98.7% completeness) using synchrotron X-ray cryogenic (100 K) conditions. The space group is P6322 for both forms, and the unit-cell parameters are a = b = 131.36, c = 189. 76 A for native urease and a = b = 131.34, c = 190.01 A for inhibited urease. Under the same conditions, single crystals of B. pasteurii urease inhibited with acetohydroxamic acid, cisteamine, and phenylphosphorodiamidate were also obtained.
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PMID:Crystallization and preliminary high-resolution X-ray diffraction analysis of native and beta-mercaptoethanol-inhibited urease from Bacillus pasteurii. 976 12

Most current non-invasive tests for Helicobacter pylori depend on the conversion of labelled (13C or 14C) urea to labelled carbon dioxide (13CO2 or 14CO2) and ammonium (NH4+) by the enzyme urease, with the labelled CO2 detected in exhaled air. Despite suggestions going back over a number of years, the alternative possibility of using NH4+ (in the form of gaseous ammonia [NH3]) as the test parameter has received little or no attention. However, this approach is now being explored using a chemiresistive sensor detecting sub-parts per million concentrations of NH3. An in vitro 'glass stomach' (containing various volumes of hydrochloric acid [HCl] and ammonium chloride [NH4Cl]) was used to evaluate the means of increasing 'gastric' pH to that of the NH4+-->NH3 transition that occurs significantly at pH 9.24. This 'stomach' also was used to study mechanisms by which NH3 may be expelled in a pulse (as a surrogate belch), either by the in situ production of CO2 or through an exogenous source. On the basis of the protocols developed, H. pylori-negative subjects were tested before and after ingestion of 10 mg NH4Cl (as a surrogate for bacteria-produced NH4,), and H. pylori-positive subjects were tested without taking urea or NH4Cl. 'Intragastric' pH in the in vitro 'glass stomach' could be increased above pH 9.24 by adding a mixture of 15-30 mL magnesium hydroxide mixture (or the proprietary equivalent) and 50 mL water, and the resulting NH3 expelled by adding 100 mL CO2-saturated cold water (sparkling water). In vivo, NH3 levels in the oral cavity of H. pylori-negative subjects were increased after ingestion of 10 mg NH4Cl; however, levels in the oral cavity of a small number of H. pylori-positive subjects were two- to threefold higher after magnesium hydroxide and sparkling water. On the basis of in vitro studies, an in vivo protocol was developed to increase gastric pH above that required for the NH4+-->NH3 transition, and a mechanism established to release the NH3 into the oral cavity. Preliminary in vivo data confirm the chemiresistive sensor is sufficiently sensitive to NH3 to distinguish H. pylori-negative subjects who have taken 10 mg NH4Cl from those who have not, and clearly distinguish H. pylori-negative subjects from H. pylori-positive subjects. Ingestion of urea or other labelled tracers is not required, nor is belching; and the sensor takes less than two minutes to reach a maximum response. The data provide good evidence that the chemiresistive detection of NH3 has considerable potential as a rapid, point-of-care diagnostic test for H. pylori infection.
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PMID:Ammonia vapour in the mouth as a diagnostic marker for Helicobacter pylori infection: preliminary 'proof of principle' pharmacological investigations. 1144 Feb 9


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