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)

An urea-ENFET (Enzyme field effect transistor) probe was made by covering one of the grids of the dual ISFET (Ion sensitive field effect transistor) with a membrane of cross-linked bovine serum albumin (BSA)-urease and the other with cross-linked BSA, and the response characteristics of the probe was then tested through differential measurements. In different concentrations of phosphate buffer, the sensor responded to various concentrations of urea solution within 10-60s. From the calibration curve plotted on logarithmic scales a linear concentration range of 1.0-8.0 mg/dl was acquired, and the correlation coefficient and response sensitivity were 0.997 and 50mV/dec. (mg/dl), respectively. However, in dilute urea solution, the sensor responded linearly to the contents of urea over the range of concentration of 0.1-1.0 mg/dl with a correlation coefficient of 0.998 and a response sensitivity of 12-15mV/mg/dl. The standard deviation and the variation coefficient for 20 performances responding to 100mg/dl urea in 0.01M pH7.0 phosphate buffer were found to be 1.39mV and 1.44%, respectively. The urea-ENFET was used for the determination of BUN (Blood urea nitrogen) and the BUN values were compared with those determined by enzymatic method, the repression equation and correlation coefficient for 50 assays were y = -0.1272 + 0.9695x and r = 0.9912, respectively. When the urea-ENFET was used for determining urea either in buffer solution or in serum for 250 runs over a period of 1.5 months (the enzyme FET was stored at 4 degrees C between measurements during this period), the observed decrease of response sensitivity was only about 10%.(ABSTRACT TRUNCATED AT 250 WORDS)
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PMID:Miniature urea sensor based on H(+)-ion sensitive field effect transistor and its application in clinical analysis. 133 30

Cysteine 319 in the large subunit of Klebsiella aerogenes urease was identified as an essential catalytic residue based on chemical modification studies (Todd, M.J., and Hausinger, R.P. (1991) J. Biol. Chem. 266, 24327-24331). Through site-directed mutagenesis, this cysteine has been changed independently to alanine, serine, aspartate, and tyrosine. None of these mutations (C319A, C319S, C319D, and C319Y, respectively) affected the size or level of synthesis of the urease subunits as monitored by polyacrylamide gel electrophoresis. The wild type enzyme and each of the mutant proteins was purified and their properties were compared. The C319Y protein possessed no detectable activity, while activity was reduced in C319A, C319S, and C319D to 48, 4.5, and 0.03% of wild type levels under normal assay conditions. All of the active mutants had a small increase in Km when compared to the wild type value. The active mutants displayed a greatly reduced sensitivity to inactivation by iodoacetamide in comparison to the wild type enzyme, confirming our previous assignment of the essential cysteine to this residue based on active site peptide mapping. In contrast to the wild type enzyme, inactivation of the mutant proteins was not affected by the presence of the competitive inhibitor phosphate, suggesting that the remaining slow rate of iodoacetamide inactivation is due to modification away from the active site. The pH dependence of urease activity was substantially altered in the active mutants with C319S and C319D showing a pH optimum near 5.2, and C319A near 6.7, compared to the pH 7.75 optimum of wild type urease. These data are consistent with Cys-319 facilitating catalysis at neutral and basic pH values by participating as a general acid.
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PMID:Site-directed mutagenesis of the active site cysteine in Klebsiella aerogenes urease. 140 Mar 17

The advent of genetic engineering has resulted in a proliferation of protein pharmaceuticals available for a variety of therapeutic needs. However, the formulation and delivery of these proteins remain an intriguing challenge. Polymer-based protein drug delivery systems continue to be investigated, although many of the fabrication techniques used to incorporate proteins into the polymer matrix or device result in irreversible inactivation (denaturation) of the proteins. A well-characterized model enzyme, urease, was formulated in 33% (w/w) poloxamer 407 (Pluronic F-127) vehicle and injected intraperitoneally (ip) into rats in an attempt to achieve both preservation of biological activity and sustained release of the protein. The resulting ammonia concentration in plasma-time profiles were compared with those for rats injected with an identical dose (27.6 units of activity per 200 g of body weight) of urease dissolved in pH 7 phosphate buffer. Neither a pH 7 phosphate buffer solution nor poloxamer 407 (33%, w/w) dissolved in pH 7 phosphate buffer, when injected ip into rats, resulted in elevated ammonia levels in plasma. The time to reach a maximum ammonia level in plasma was increased approximately threefold following the injection of the urease-poloxamer 407 formulation, compared with that in control rats administered an identical dose of urease in solution. In addition, hyperammonemia was extended almost threefold in treated rats compared with control rats, without untoward effects. However, prolonged hyperammonemia in animals receiving an ip injection of the urease-poloxamer 407 formulation may have potentially resulted from the reduced clearance of ammonia and ammonium ion in the proximal tubules of the rats.(ABSTRACT TRUNCATED AT 250 WORDS)
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PMID:Biological activity of urease formulated in poloxamer 407 after intraperitoneal injection in the rat. 140 93

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

The composition of 3,084 urinary calculi was determined using an infrared spectrophotometer. Mixed calcium oxalate-calcium phosphate stones were most frequently implicated. Of the urinary calculi analyzed 199 were associated with urinary tract infection. Escherichia coli was most frequently isolated (43 strains) and urease-producing organisms, such as Proteus mirabilis, were cultured from 40 patients. The core culture of 20 staghorn calculi yielded 15 isolates from 14 stones. There were 13 identical species isolated from the urine and stone specimens of 13 patients (65%), including 7 strains of P. mirabilis. These results suggest that cultures of urine specimens of urolithiasis patients, especially those with staghorn calculi, may help to elucidate the bacteriology of the stones.
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PMID:Composition of urinary calculi related to urinary tract infection. 150 58

Three cases of encrusted cystitis caused by Corynebacterium group D2 are described. The vesical damage previous to the establishment of this bacteria is noteworthy and the very rapid increase in urease activity explains the pathogenesis of the situation. Thus allowing for its identification and is relevant to treatment. Cloudy urine with a strong smell of ammonium, alkaline pH and crystals of ammonium magnesium phosphate in the sediment will bring this microorganism and its characteristic growth pattern to mind thus avoiding a falsely negative report. Treatment combining an antimicrobial agent and cystoscopic resection of the encrusted stones, where Corynebacterium group D2 has lodged, has proved efficacious. Vancomycin and teicoplanin have always been active and are eliminated through the kidneys.
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PMID:[Incrusted cystitis with isolation of Corynebacterium group D2]. 153 60

Although successful in reducing urea levels, the use of oral microcapsules containing a urease-silica adduct and a zirconium phosphate ion exchanger result in a number of problems, including a negative calcium balance. In this study, it is demonstrated that the use of microcapsules containing a urease-zeolite preparation may be a potential route to urea removal. The use of zeolite ion exchangers, and zeolite W in particular, can alleviate the problems encountered with zirconium phosphate. Unlike zirconium phosphate, zeolite W is nonselective toward calcium ions and is stable at the high pH found in the intestinal tract. Zeolite W, when present in the sodium form, has a high ammonium capacity of 3.6 mEq NH4+/g zeolite under simulated intestinal conditions; its reactivity to ammonium is also higher. The application of enzyme envelopes to zeolite particles is a novel immobilization procedure that does not involve the use of colloidal silica and can reduce the amount of ingested material by as much as 25%. The current in vitro study shows that cellulose acetate butyrate microcapsules, containing a urease-zeolite preparation, remove up to 80% of urea in less than 1 hour. These microcapsules can be dried and retain activity when sealed in a jar at 4 degrees C.
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PMID:The potential of a microencapsulated urease-zeolite oral sorbent for the removal of urea in uremia. 164 15

It is reasonable to assume that the rate of pH increase in urine induced by urease-producing microorganisms is one of the factors which determine whether crystallisation with subsequent stone formation will occur or not. To evaluate how the time needed to increase urine pH varies between different urine samples and how it depends on urine composition, a standardised amount of urease was added to different human urine samples. The incubations were performed in a pH-stat. This allowed simultaneous study of how urease enzymatic activity depends on urine pH and how it varies between different urines. The enzymatic activity was found to be negatively correlated to urine pH and to vary between different urines. The rate of the pH increase varied markedly between different urines. Small pH increases depended on the native urine pH and urease enzymatic activity. Higher pH increases up to the levels of phosphate crystallisation depended more on urine phosphate, the major urine buffer. The results presented show that urine composition influences the urease-induced pH increase. This might have clinical implications.
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PMID:The influence of pH and urine composition on urease enzymatic activity in human urine. 173 85

During reaction with [14C]iodoacetamide at pH 6.3, radioactivity was incorporated primarily into a single Klebsiella aerogenes urease peptide concomitant with activity loss. This peptide was protected from modification at pH 6.3 by inclusion of phosphate, a competitive inhibitor of urease, which also protected the enzyme from inactivation. At pH 8.5, several peptides were alkylated; however, modification of one peptide, identical to that modified at pH 6.3, paralleled activity loss. The N-terminal amino acid sequence and composition of the peptide containing the essential thiol was determined. Previous enzyme inactivation studies of K. aerogenes urease could not distinguish whether one or two essential thiols were present per active site (Todd, M. J., and Hausinger, R. P. (1991) J. Biol. Chem. 266, 10260-10267); we conclude that there is a single essential thiol present and identify this residue as Cys319 in the large subunit of the heteropolymeric enzyme.
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PMID:Identification of the essential cysteine residue in Klebsiella aerogenes urease. 176 35

The urease-induced precipitation of phosphate salts on indwelling catheters was studied in an experimental in vitro model. The precipitation was strongly pH-related and was much higher in synthetic urine than in human urine. In the latter, it was significantly lower on silicone catheters than on latex catheters, including those with a hydrophilic coating. The precipitation on silicone catheters that had been in situ was not increased as compared with that on unused catheters, in contrast to latex catheters with a hydrophilic coating, among which the precipitation on used catheters was higher.
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PMID:Urease-induced precipitation of phosphate salts in vitro on indwelling catheters made of different materials. 194 29


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