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:6.3.4.6 (urease)
7,490 document(s) hit in 31,850,051 MEDLINE articles (0.00 seconds)

The urease of Helicobacter pylori is suspected to play a role in the pathogenesis of gastritis. Although all clinical isolates of H. pylori are urease positive (U+), we have selected and characterized several spontaneously arising urease-negative (U-) variants from wild-type strain 60190. Urease-negative variants were identified by growth in medium containing 60 mM urea and arose at a frequency of 10(-5) to 10(-6). The urease activity of the wild-type strain inhibited growth of this strain in the presence of 60 mM urea. U- variants retained the U- phenotype for more than 100 passages on medium with or without urea. The urease activities of the original U+ and derived U- cells were 9.55 to 16.7 and 0.01 to 0.17 U/mg of protein, respectively. Colonial growth and other biochemical characteristics were identical for the strains. U- variants showed three classes of whole-cell sodium dodecyl sulfate-polyacrylamide gel electrophoresis profiles: (i) identical to U+; (ii) change in the migration of the 61-kDa urease subunit; and (iii) lack of 61- and 30-kDa subunits. These differences were confirmed by immunoblotting and by protein separation using fast protein liquid chromatography. The U+ strain but not U- variants tolerated exposure to pH 4.0 for 60 min in the presence of urea. Supernatants of the U+ strain and U- variants contained vacuolating cytotoxin activity for HeLa cells in similar titers. By enzyme-linked immunosorbent assay, human serum samples recognized water extract from the U+ strain significantly better than extract from a U- variant lacking urease subunits. In conclusion, this study demonstrates that U- H. pylori variants may arise spontaneously, that urease activity enhances survival at acid pH, and that urease and cytotoxin activities are disparate phenotypes.
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PMID:Characteristics of Helicobacter pylori variants selected for urease deficiency. 150 Jan 74

Helicobacter pylori, a gram-negative, microaerophilic, spiral-shaped bacterium, is an etiologic agent of human gastritis and peptic ulceration and is highly restricted to the gastric mucosa of humans. Urease, synthesized at up to 6% of the soluble cell protein, hydrolyzes urea, thereby releasing ammonia, which may neutralize acid, allowing survival of the bacterium and initial colonization of the gastric mucosa. The urease protein is encoded by two subunit genes, ureA and ureB; however, accessory genes are necessary for enzyme activity. H. pylori urease genes were isolated from a cosmid gene bank and subcloned on a 5.8-kb Sau3A partial fragment carrying ureCDAB, corresponding to four open reading frames described by A. Labigne, V. Cussac, and P. Courcoux (J. Bacteriol. 173:1920-1931, 1991). Clones were confirmed as ureas gene sequences by polymerase chain reaction amplification. The recombinant enzyme was purified from the soluble protein of French press lysates of Escherichia coli DH5 alpha(pHP402) by chromatography on DEAE-Sepharose, Phenyl-Sepharose, Mono-Q, and Superose 6 resins. Fractions containing a catalytically inactive apoenzyme were identified by an enzyme-linked immunosorbent assay (ELISA) by using antisera to native UreA (29.5 kDa) and UreB (66 kDa). Purified recombinant urease was indistinguishable from native enzyme on a Superose 6 column and on Coomassie blue-stained sodium dodecyl sulfate-polyacrylamide gels. The protein reacted specifically on Western blots (immunoblots) with anti-UreA and anti-UreB antibodies and was recognized with an intensity equal to that of the native enzyme in an ELISA using human sera. Clones containing only ureA and ureB also produced an assembled but inactive enzyme. Enzyme activity was not restored by in trans complementation with cloned urease accessory gene sequences from Proteus mirabilis or Morganella morganii. H. pylori urease genes (ureCDAB) subcloned into pACYC184 were also not complemented with any of 1,000 cosmid clones containing H. pylori chromosomal sequences. However, larger clones containing 4.5 kb of DNA downstream of ureB synthesized catalytically active urease when grown in minimal medium. These data indicate that the ureA and ureB genes encoding H. pylori urease are transcribed and translated in E. coli and that these genes alone are sufficient for the synthesis and assembly of the native size enzyme. Genes downstream of ureB, however, are necessary for production of a catalytically active urease.
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PMID:Purification of recombinant Helicobacter pylori urease apoenzyme encoded by ureA and ureB. 161 35

The region located immediately upstream from the Klebsiella aerogenes urease structural genes was sequenced and shown to possess an open reading frame capable of encoding a 29.8-kDa peptide. Deletions were generated in this gene, denoted ureD, and in each of the genes (ureE, ureF, and ureG) located immediately downstream of the three structural genes. Transformation of the mutated plasmids into Escherichia coli resulted in high levels of urease expression, but the enzyme was inactive (deletions in ureD, ureF, or ureG) or only partially active (deletions in ureE). Ureases were purified from the recombinant cells and shown to be identical to control enzyme when analyzed by gel filtration chromatography and sodium dodecyl sulfate-polyacrylamide gel electrophoresis; however, in every case the activity levels correlated to nickel contents as analyzed by atomic absorption analysis. UreD, UreE, UreF, and UreG peptides were tentatively identified by gel electrophoretic comparison of mutant and control cell extracts, by in vivo expression of separately cloned genes, or by in vitro transcription-translation analyses; the assignments were confirmed for UreE and UreG by amino-terminal sequencing. The latter peptides (apparent M(r)s, 23,900 and 28,500) were present at high levels comparable to those of the urease subunits, whereas the amounts of UreF (apparent M(r), 27,000) and UreD (apparent M(r), 29,300) were greatly reduced, perhaps because of the lack of good ribosome binding sites in the regions upstream of these open reading frames. These results demonstrate that all four accessory genes are necessary for the functional incorporation of the urease metallocenter.
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PMID:Klebsiella aerogenes urease gene cluster: sequence of ureD and demonstration that four accessory genes (ureD, ureE, ureF, and ureG) are involved in nickel metallocenter biosynthesis. 162 27

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

Urinary tract infection with Proteus mirabilis may lead to serious complications, including cystitis, acute pyelonephritis, fever, bacteremia, and death. In addition to the production of hemolysin and the enzyme urease, fimbriae and flagellum-mediated motility have been postulated as virulence factors for this species. We purified mannose-resistant/proteuslike (MR/P) fimbriae and flagella from strains CFT322 and HU2450, respectively. Electron microscopy revealed highly concentrated preparations of fimbriae and flagella. Fimbrial and flagellar structural subunits were estimated by sodium dodecyl sulfate-polyacrylamide gel electrophoresis to be 18.5 and 41 kDa, respectively. N-terminal sequencing revealed that 10 of the first 20 amino acids of the major MR/P subunit matched the sequence of the P. mirabilis uroepithelial cell adhesin N terminus and 11 of 20 amino acids matched the predicted amino acid sequence of the Escherichia coli P fimbriae structural subunit, PapA. In addition, 90 and 80% homologies were found between the first 20 amino acids of P. mirabilis flagellin and those of Salmonella typhimurium phase-1 flagellin and the E. coli hag gene product, respectively. An enzyme-linked immunosorbent assay using purified antigens showed a strong reaction between the MR/P fimbriae or flagella and sera of CBA mice challenged transurethrally with P. mirabilis. A possible role for MR/P fimbriae in the pathogenesis of urinary tract infection is supported by (i) a strong immune response to the antigen in experimentally infected animals, (ii) amino acid sequence similarity to other enteric surface structure, and (iii) our previously reported observation that MR/P fimbriae are expressed preferentially as the sole fimbrial type in human pyelonephritis isolates.
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PMID:Proteus mirabilis flagella and MR/P fimbriae: isolation, purification, N-terminal analysis, and serum antibody response following experimental urinary tract infection. 168 Jan 6

Two 21-d trials were conducted to evaluate the effect of heating time and sodium metabisulfite (SMBS) on the nutritional value of full-fat soybeans for chicks. In Trial 1, four pen-replicates of eight chicks each were fed corn-based diets (19% CP; 3,167 kcal of ME/kg) containing either 44% CP soybean meal or full-fat soybeans. The soybeans either were unheated or were autoclaved at 121 degrees C for 10, 20, 30, 40, 60, or 90 min. Soybean oil was added to the soybean meal diet to make it isoenergetic with the soybean diets. Trypsin inhibitor, urease activity, and the solubility of protein in the soybeans decreased as heating time increased. Weight gain increased and feed:gain and pancreas weights decreased quadratically (P less than .01) with heating time. Rate and efficiency of gain were maximized when the soybeans were heated for 40 min; further heating for 60 or 90 min reduced performance. In Trial 2, SMBS was added at levels of 0, 1, or 2% to full-fat, unheated soybeans or to soybeans before autoclaving at 121 degrees C for 10, 20, or 40 min. Four pen-replicates of seven chicks each were fed corn-soybean diets (19% CP; 3,144 kcal of ME/kg) with 12 treatments in a factorial arrangement of heating times and SMBS levels. The rate and efficiency of chick weight gain improved linearly (P less than .01) and pancreas weights decreased linearly (P less than .01) as heating time increased. Less heating time was required to maximize performance and minimize pancreas weights when SMBS was added, resulting in a heating time x SMBS interaction (P less than .05). Under the conditions of this research, chicks fed full-fat soybeans achieved maximum performance when the soybeans were heated at 121 degrees C for 40 min, and SMBS decreased by one-half the heating time required to inactivate the trypsin inhibitors. Trypsin inhibitor activity in soybeans was more closely related to their nutritional value than was urease activity.
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PMID:Effects of heating time and sodium metabisulfite on the nutritional value of full-fat soybeans for chicks. 175 23

Lactic acid production, urease activity and genetic stability were investigated in five selected rumen strains of Enterococcus faecium. The average value of urease activity in the tested strains was 16.5 +/- 0.953 nkat per ml, two strains were urease-negative. The values of E. faecium strains produced lactic acid ranged from 1.087 +/- 0.134 to 1.787 +/- 0.213 mmol per 1 l. Cultivation of the strains in ethidium bromide (EB) eliminated urease activity of these strains already in the first subculture, but the elimination effects of sodium dodecyl sulphate (SDS) and acridine orange (AO) were manifested later on (1-8 subcultures). Lactic acid production was eliminated in all strains from 1st to 8th subculture after cultivation in SDS, EB and also AO.
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PMID:[Lactic acid production and urease activity in strains of Enterococcus faecium found in the rumen and their genetic stability]. 180 29

Previous investigators have suggested that urinary tract infections with urea-splitting organisms may be a primary etiologic factor in the acidosis which is seen after urinary diversion. This study employs a model in which small intestinal segments are perfused with an artificial urine solution over a three hour period. Urease is then added in order to determine its effect on acid-base balance and net intestinal electrolyte transport. Urease created no significant increase in acid load (delta HCO3- = -7.5 +/- 2.2 for controls vs. -8.7 +/- 2.9 for urease group), but did increase the osmolality of the intestinal contents and resulted in a 24% increase in free water loss (p = .037). Analysis of sodium and chloride movement following the addition of urease to the perfusate suggests that both ammonium and bicarbonate are absorbed by the intestinal segment. Thus any acidosis resulting from increased ammonium absorption following the addition of urease appears to be offset by concomitant bicarbonate absorption. The azotemia of urinary diversion appears to be primarily the result of urea absorption, partially the result of ammonium absorption, and is not significantly increased by urease.
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PMID:Urease and the acidosis of urinary intestinal diversion. 185 52

Nephrolithiasis is a heterogeneous disorder, with varying chemical composition and pathophysiologic background. Although kidney stones are generally composed of calcium oxalate or calcium phosphate, they may also consist of uric acid, magnesium-ammonium phosphate, or cystine. Stones develop from a wide variety of metabolic or environmental disturbances, including varying forms of hypercalciuria, hypocitraturia, undue urinary acidity, hyperuricosuria, hyperoxaluria, infection with urease-producing organisms, and cystinuria. The cause of stone formation may be ascertained in most patients using the reliable diagnostic protocols that are available for the identification of these disturbances. Effective medical treatments, capable of correcting underlying derangements, have been formulated. They include sodium cellulose phosphate, thiazide, and orthophosphate for hypercalciuric nephrolithiasis; potassium citrate for hypocitraturic calcium nephrolithiasis; acetohydroxamic acid for infection stones; and D-penicillamine and alpha-mercaptopropionylglycine for cystinuria. Using these treatments, new stone formation can now be prevented in most patients.
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PMID:Etiology and treatment of urolithiasis. 196 46

A mutant strain of Helicobacter pylori with weak urease activity was created by using N-methyl-N'-nitro-N-nitrosoguanidine. The urease activity of the mutant (0.036 +/- 0.009 nmol of urea per micrograms of bacterial protein per min) was 0.4% of that of the parental strain (8.20 +/- 2.30 nmol of urea per micrograms of bacterial protein per min). The mutant was otherwise indistinguishable from the parental strain. Both demonstrated prominent catalase and oxidase activities, and both produced vacuolating cytotoxin. Restriction endonuclease and sodium dodecyl sulfate-polyacrylamide gel electrophoresis (SDS-PAGE) patterns and ultrastructure were identical for the two strains. The mutant was fully motile, as evaluated by spreading in soft agar and by direct microscopic examination. Growth rate and colony size and morphology were identical for the mutant and parental strains. Seventeen gnotobiotic piglets were challenged with either the mutant or the parental strain and sacrificed 3 or 21 days after challenge. Gastric tissue was examined histologically and cultured for H. pylori. Of seven piglets challenged with the parental strain, all became infected. H. pylori was not recovered from any of 10 piglets challenged with the urease-negative strain. Lymphofollicular gastritis was present in all seven piglets challenged with the parental strain but in none of the piglets challenged with the urease-negative strain. These results suggest that prominent urease activity is essential for colonization by H. pylori.
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PMID:Essential role of urease in pathogenesis of gastritis induced by Helicobacter pylori in gnotobiotic piglets. 205 Apr 11


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