Gene/Protein Disease Symptom Drug Enzyme Compound
Pivot Concepts: Gene/Protein Disease Symptom Drug Enzyme Compound   Target Concepts: Gene/Protein Disease Symptom Drug Enzyme Compound

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Query: EC:3.5.1.5 (urease)
7,257 document(s) hit in 31,850,051 MEDLINE articles (0.00 seconds)

Gyrocotyle fimbriata isolated from the spiral valve of Hydrolagus colliei were washed, then held in a filtered seawater-penicillin-Tris buffer medium. Ammonia and urea release to the medium declined together and ammonia production was minimal when the urea concentration was below detectable limits. Alanine and smaller amounts of glycine were released to the medium at a more constant rate. After 12 hr the alanine-glycine excretion was more than 20 times the ammonia excretion. L-arginine, L-serine, L-histidine, and urea were most effective in stimulating ammonia production by whole worms; other L-amino acids were essentially ineffective. L-glutamate dehydrogenase, L-amino acid oxidase, uricase, and ornithine transcarbamylase were below detectable levels. L-serine dehydrase, L-arginase, L-histidase, and urease were detected in tissue homogenates and probably account for most of the endogenous ammonia production. L-arginase has a molecular weight of 28,000 by Sehpadex gel filtration. The high levels of glutamate-pyruvate transaminase and lower levels of glutamate-oxalacetate transaminase correlate with the high level of alanine excretion. It is concluded that (1) ammonia production is not strongly linked to the overall energy metabolism of Gyrocotyle and is probably a result of a series of unrelated enzymatic reactions such as the action of urease of urea from the tissue of the rat fish, and (2) alanine and glycine are the major nitrogen excretory products and their production is linked to the energy metabolism of Gyrocotyle.
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PMID:Ammonia formation and amino acid excretion by Gyrocotyle fimbriata (Cestoidea). 111 78

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

We describe a coupled-enzyme equilibrium method for measuring urea in serum, which is performed on supernates prepared by treating each specimen with Ba(OH)2 and ZnSO4 (Somogyi reagent). Analytical recovery of [14C]urea added to a variety of matrices was essentially complete (mean, 100.6%) for the supernates after precipitation. Nine variables were univariately examined in arriving at the reaction conditions for the method: glutamate dehydrogenase, urease, 2-oxoglutarate, ADP, Tris . HCI, NADH, EDTA, pH, and temperature. The reagent is stable for at least 48 days at--20 degrees C and for 23 days at 4 degrees C. Mean analytical recovery of urea (14 mmol/L) added to seven different specimens (three different matrices) was 100.8%. The analytical linear range of the method extends to 30 mmol of urea per liter. Of 22 potential interferents, only bilirubin at 1 mmol/L (580 mg/L), hemoglobin at 10 g/L, and hydroxyurea at 6 mmol/L showed more than 2% interference. We discuss precision and effects of specimen dilution, and compare results for 100 human serum specimens with those measured for the same specimens with four other urea methods. We examined the effects of measuring a blank, consisting of sample and reagent without urease, with each specimen.
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PMID:A coupled-enzyme equilibrium method for measuring urea in serum: optimization and evaluation of the AACC study group on urea candidate reference method. 737 2

Portions of the 16S RNA from a urease-positive Bilophila wadsworthia strain were sequenced, and a probe was constructed. The probe was end labeled with [32P]ATP and polynucleotide kinase and hybridized on a nylon filter (by dot blot hybridization) to the immobilized rRNA of 12 B. wadsworthia strains and eight other anaerobic isolates. The probe efficiently hybridized only to the Bilophila strains. Cross-reactivity at high RNA levels (2,000 ng) was observed with one strain of Bacteroides thetaiotamicron and one strain of Bacteroides fragilis (with 10x SET buffer [20x SET buffer is 0.5 M NaCl, 0.03 M Tris, and 2 mM EDTA]) but was not seen at lower RNA levels or with 5x SET buffer. When tested against mixed cultures of aerobic and anaerobic isolates representative of appendiceal abscess flora, the probe did not react with mixed cultures containing no Bilophila cells and could detect > or = 10(5) Bilophila CFU/ml when the mixture was seeded with Bilophila cells. This probe is of potential use in the rapid identification of pure isolates and in the direct identification of B. wadsworthia in clinical specimens.
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PMID:Preliminary study using species-specific oligonucleotide probe for rRNA of Bilophila wadsworthia. 752 41

Urease from pigeonpea was entrapped in polyacrylamide gel with 50% immobilization at 10% total monomer (containing 5% cross-linker) with high mechanical stability of the gel. Approximately 0.61 mg of protein could be loaded per 5 ml of gel. The immobilized enzyme had a t1/2 of approx. 200 days when stored in 0.1 M Tris/acetate buffer, pH 6.5, at 4 degrees C. The gel strips were used 4-5 times for urea assay over a period of 6 h with less than 2% loss of activity. Approximately 50% immobilization of urease in calcium alginate was observed at 3% alginate with 0.12 mg protein/ml alginate. The resultant enzyme beads showed a t1/2 of approx. 75 days when stored in 0.1 M Tris/acetate buffer, pH 6.5, at 4 degrees C. The beads were used 4-5 times for urea assay over a period of 6 h with about 40% loss of activity. In both cases, the enzyme activity was directly proportional to the amount of immobilized enzyme. There was practically no leaching of the entrapped enzyme over a period of 48 h from either of the polymers. Both the immobilized enzyme preparations were used to analyse the blood urea of some clinical samples from the University hospital. The results obtained compared favourably with those obtained by the usual method employed in the clinical pathology laboratory.
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PMID:Immobilization of urease from pigeonpea (Cajanus cajan L.) in polyacrylamide gels and calcium alginate beads. 947 53

A simple novel method was introduced for determination of an inhibitor binding constant (Ki) and enthalpy of binding by isothermal titration microcalorimetry technique. This method was applied to the binding of fluoride ion, as an inhibitor, with the active sites of jack bean urease at pH = 7.0 (Tris 30 mM) and T = 300 degrees K. The dissociation equilibrium constant measured by this method was markedly consistent with the inhibition constant obtained from assay of enzyme activity in the presence of fluoride ion.
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PMID:A simple novel method for determination of an inhibition constant by isothermal titration microcalorimetry. The effect of fluoride ion on urease. 950 48

We describe the development of an immunoligand assay (ILA) in conjunction with a light-addressable potentiometric sensor (LAPS) for the rapid detection of Escherichia coli O157:H7 cells in buffered saline. The ILA protocol consists of "sandwiching" bacterial analyte between biotinylated and fluoresceinated antibodies, indirect enzyme labeling of the bacteria with urease-labeled anti-fluorescein antibody, and active capture of the immune complex at a biotinylated bovine serum albumin-blocked nitrocellulose filter membrane with streptavidin. Using live E. coli O157:H7, the efficiency of the ILA was compared using various ratios of the biotinylated and fluoresceinated antibodies. Simultaneous addition of equimolar biotinylated and fluoresceinated antibodies effected optimal urease labeling and subsequent active capture of the bacteria in the ILA. Equimolar concentrations of the antibodies were varied to achieve optimal LAPS detection response for the live bacteria. Using ILA with LAPS, a minimum detectable level of ca. 7.1 x 10(2) cells/ml of heat-killed or ca. 2.5 x 10(4) cells/ml of live E. coli O157:H7 bacteria was achieved in Tris-buffered saline in an assay time of ca. 45 or ca. 30 min, respectively.
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PMID:Use of a light-addressable potentiometric sensor for the detection of Escherichia coli O157:H7. 957 Aug 43

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

The biosensor with urease entrapped in PVC layer at the surface of pH-sensitive iridium oxide electrode was applied for testing of mercury and other metal ions inhibition on enzymatic reaction. The calculation of inhibition effect was based on the measurement of initial rate of decrease of biosensor potential (proportional to the initial rate of enzymatic reaction) after addition of substrate after inhibition step. Some differences of inhibition extent were observed for various mercury forms (Hg(NO3)2, HgCl2, PhHgCl and Hg2(NO3)2) as well as for other heavy metal ions investigated as potential interferents. Because the method was not specific, it was applied for the determination of total inhibition effect caused by heavy metal ions in water samples. In the case of most cations tested the total recovery of enzyme activity was possible using Tris buffer solution with EDTA and thioacetamide after less than 10 min regeneration time.
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PMID:Inhibitive determination of mercury and other metal ions by potentiometric urea biosensor. 1121 29


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