Search PubMed⌕ Search

SEARCH · Search PubMed

Results for “UREASE”

Search indexed PubMed citations on genomics, clinical trials, systematic reviews and public health. Explore titles, authors and supplied subject terms, then open the PubMed record.

Quote a phrase for an exact phrase match. Source license links do not imply unrestricted reuse.

At least 487 records · Page 27Linked to original sources
PubMed1990

Sequence of the Klebsiella aerogenes urease genes and evidence for accessory proteins facilitating nickel incorporation.

A 4.8-kilobase-pair region of cloned DNA encoding the genes of the Klebsiella aerogenes urease operon has been sequenced. Six closely spaced open reading frames were found: ureA (encoding a peptide of 11.1 kilodaltons [kDa]), ureB (11.7-kDa peptide), ureC (60.3-kDa peptide), ureE (17.6-kDa peptide), ureF (25.2-kDa peptide), and ureG (21.9-kDa peptide). Immediately after the ureG gene is a putative rho-dependent transcription terminator. The three subunits of the nickel-containing enzyme are encoded by ureA, ureB, and ureC based on protein structural studies and sequence homology to jack bean urease. Potential roles for ureE, ureF, and ureG were explored by deleting these accessory genes from the operon. The deletion mutant produced inactive urease, which was partially purified and found to have the same subunit stoichiometry and native size as the active enzyme but which contained no significant levels of nickel. The three accessory genes were able to activate apo-urease in vivo when they were cloned into a compatible expression vector and cotransformed into cells carrying the plasmid containing ureA, ureB, and ureC. Thus, one or more of the ureE, ureF, or ureG gene products are involved in nickel incorporation into urease.

Amino Acid Sequence↗
PubMed1992

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.

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.

Amino Acid Sequence↗
PubMed1994

Cloning, sequencing, and expression of thermophilic Bacillus sp. strain TB-90 urease gene complex in Escherichia coli.

The urease of thermophilic Bacillus sp. strain TB-90 is composed of three subunits with molecular masses of 61, 12, and 11 kDa. By using synthetic oligonucleotide probes based on N-terminal amino acid sequences of each subunit, we cloned a 3.2-kb EcoRI fragment of TB-90 genomic DNA. Moreover, we cloned two other DNA fragments by gene walking starting from this fragment. Finally, we reconstructed in vitro a 6.2-kb DNA fragment which expressed catalytically active urease in Escherichia coli by combining these three DNA fragments. Nucleotide sequencing analysis revealed that the urease gene complex consists of nine genes, which were designed ureA, ureB, ureC, ureE, ureF, ureG, ureD, ureH, and ureI in order of arrangement. The structural genes ureA, ureB, and ureC encode the 11-, 12-, and 61-kDa subunits, respectively. The deduced amino acid sequences of UreD, UreE, UreF, and UreG, the gene products of four accessory genes, are homologous to those of the corresponding Ure proteins of Klebsiella aerogenes. UreD, UreF, and UreG were essential for expression of urease activity in E. coli and are suggested to play important roles in the maturation step of the urease in a co- and/or posttranslational manner. On the other hand, UreH and UreI exhibited no significant similarity to the known accessory proteins of other bacteria. However, UreH showed 23% amino acid identity to the Alcaligenes eutrophus HoxN protein, a high-affinity nickel transporter.

Alcaligenes↗
PubMed2002

Energetics of Helicobacter pylori and its implications for the mechanism of urease-dependent acid tolerance at pH 1.

In the presence of urea the neutrophilic human pathogen Helicobacter pylori survives for several hours at pH 1 with concomitant cytoplasmic pH homeostasis. To study this effect in detail, the transmembrane proton motive force and cytoplasmic urease activity of H. pylori were determined at various pH values. In the absence of urea, the organism maintained a close-to-neutral cytoplasm and an internally negative membrane potential at external pH values greater than 4 to 5. In the presence of urea, H. pylori accomplished cytoplasmic pH homeostasis down to an external pH of 1.2. At this external pH, the cytoplasmic pH was 4.9 and the membrane potential was slightly negative inside. The latter finding is in contrast to the situation in acidophiles, which develop inside-positive membrane potentials under similar conditions. Measurements of the time course of the membrane potential confirmed that addition of urea to the cells led to hyperpolarization. Most likely, this effect was due to electrogenic export of ammonium cations from the cytoplasm. The urease activity of intact cells increased nearly exponentially with decreasing external pH. This activation was not due to enhanced gene expression at low external pH values. In cell extracts the pH optimum of urease activity was dependent on the buffer system and was about pH 5 in sodium citrate buffer. Since this is the cytoplasmic pH of the cells at pH 1 to 2, we propose that cytoplasmic pH is a factor in the in vivo activation of the urease at low external pH values. The mechanism by which urease activity leads to cytoplasmic pH homeostasis in H. pylori is discussed.

Citrates↗
PubMed2002

Genes encoding specific nickel transport systems flank the chromosomal urease locus of pathogenic yersiniae.

The transition metal nickel is an essential cofactor for a number of bacterial enzymes, one of which is urease. Prior to its incorporation into metalloenzyme active sites, nickel must be imported into the cell. Here, we report identification of two loci corresponding to nickel-specific transport systems in the gram-negative, ureolytic bacterium Yersinia pseudotuberculosis. The loci are located on each side of the chromosomal urease gene cluster ureABCEFGD and have the same orientation as the latter. The yntABCDE locus upstream of the ure genes encodes five predicted products with sequence homology to ATP-binding cassette nickel permeases present in several gram-negative bacteria. The ureH gene, located downstream of ure, encodes a single-component carrier which displays homology to polypeptides of the nickel-cobalt transporter family. Transporters with homology to these two classes are also present (again in proximity to the urease locus) in the other two pathogenic yersiniae, Y. pestis and Y. enterocolitica. An Escherichia coli nikA insertion mutant recovered nickel uptake ability following heterologous complementation with either the ynt or the ureH plasmid-borne gene of Y. pseudotuberculosis, demonstrating that each carrier is necessary and sufficient for nickel transport. Deletion of ynt in Y. pseudotuberculosis almost completely abolished bacterial urease activity, whereas deletion of ureH had no effect. Nevertheless, rates of nickel transport were significantly altered in both ynt and ureH mutants. Furthermore, the ynt ureH double mutant was totally devoid of nickel uptake ability, thus indicating that Ynt and UreH constitute the only routes for nickel entry. Both Ynt and UreH show selectivity for Ni(2+) ions. This is the first reported identification of genes coding for both kinds of nickel-specific permeases situated adjacent to the urease gene cluster in the genome of a microorganism.

ATP-Binding Cassette Transporters↗
PubMed1966

Urease activity and antibiotic sensitivity of bacteria.

Gale, Glen R. (Veterans Administration Hospital, Durham, N.C.). Urease activity and antibiotic sensitivity of bacteria. J. Bacteriol. 91:499-506. 1966.-An investigation was made of the responses of certain urease-positive bacteria to various antibacterial drugs in the presence of highly specific urease inhibitors, in a test of the hypothesis proposed by other workers that inhibition of bacterial urease enhances the sensitivity of the cells to antimicrobial agents. Urease inhibitors employed were seven hydroxamic acids (HA). Six of the seven HA reduced the sensitivity of nine Proteus strains to ampicillin and methenamine mandelate. Two HA increased the sensitivity to colistin, and six HA increased the sensitivity to kanamycin. Investigation of the mechanism of action of the synergistic effect between kanamycin and HA led to the tentative conclusion that potentiation was mediated through an initial alteration of cell permeability by the aminoglycoside antibiotic which permitted accumulation of each of the six HA into the cell, at which point each interacted with pyridoxal phosphate. The single HA which failed to yield synergism with kanamycin failed to interact with pyridoxal phosphate in a nonenzymatic system; the other six HA produced alterations of the normal ultraviolet absorption spectrum of the coenzyme.

Ampicillin↗
PubMed1981

Transferable urease activity in Providencia stuartii.

Six urea-positive Providencia stuartii strains were tested for transmissible urease determinants. Two strains, when implanted with "helper" conjugative plasmids, were found to be capable of transferring urease genes to Escherichia coli or urea-negative P. stuartii. Recombination of the urease genes with the helper plasmid in P. stuartii was noted in one case. One of the urea-positive P. staurtii strains was found to harbor a conjugative plasmid which mediated both urease activity and ampicillin resistance. This large plasmid (molecular weight, approximately 140 x 10(6)) was transmissible to and stably maintained in E. coli strains. The demonstration of transmissible genes for urease activity in P. stuartii is significant in that it accounts for previous problems associated with classifying urea-positive strains of this species.

Anti-Bacterial Agents↗
PubMed1989

Urease-positive, Kanagawa-negative Vibrio parahaemolyticus from patients and the environment in the Pacific Northwest.

We previously reported the occurrence of Vibrio parahaemolyticus in patients and the environment in the Pacific Northwest. The present studies compare the biochemical characteristics, Kanagawa hemolysin reactions, and plasmid profiles of 13 patient and 221 environmental isolates of the organism. Classical biochemical testing of the isolates revealed similar reactions for the clinical and environmental strains, and analysis in agarose gels revealed that 13 to 15% of the isolates had plasmids. The strains were tested for production of Kanagawa hemolysin on Wagatsuma agar, and 1.4% of environmental isolates and 23% of clinical isolates were positive. Clinical isolates from locally acquired extraintestinal infections were urease negative and Kanagawa hemolysin negative, isolates from locally acquired gastroenteritis cases were urease positive and Kanagawa negative, and isolates from traveler's diarrhea were urease negative and Kanagawa positive. Eight percent of the local environmental isolates were also urease positive and Kanagawa hemolysin negative. These findings suggest that expression of the Kanagawa hemolysin is not essential for the pathogenesis of V. parahaemolyticus infections. In addition, our findings suggest that V. parahaemolyticus gastroenteritis in the Pacific Northwest is associated with a urease-positive, Kanagawa-negative biotype of the organism.

Animals↗
PubMed1992

Assay of urease-inhibiting activity in serum from children infected with Helicobacter pylori.

In order to provide a basis for obtaining further information concerning the host response to Helicobacter pylori urease, four assay methods for detecting urease-inhibiting activity in serum were examined. A quantitative assay, established in a COBAS BIO centrifugal fast analyzer and based on detection of the consumption of NADH by glutamate dehydrogenase stimulated by ammonia production, was considered most suitable for large-scale serological work. Serum samples from 63 children (aged 5 to 16 years), 28 of whom had seropositive H. pylori gastritis, were assayed. One of the serum samples in this latter group showed significant inhibitory activity. This serum sample was one of 13 in the seropositive group known to bind to urease antigen. It showed no inhibitory activity against Bacillus pasteurii or jack bean urease. Protein A binding and heat treatment indicated that the inhibitory activity was immunoglobulin G mediated. The patient from whom this sample was collected showed no distinctive features in his illness. The COBAS BIO analyzer-based urease inhibition assay provides a new tool for studying one aspect of the host response to H. pylori infection.

Child↗
PubMed1999

Evaluation of the one-minute ultra-rapid urease test for diagnosing Helicobacter pylori.

To determine the diagnostic accuracy of the one-minute ultra-rapid urease test for diagnosing Helicobacter pylori infection, two biopsies were taken from both the gastric corpus and antrum from 1000 patients undergoing upper gastrointestinal endoscopy. All the biopsies were subjected to the one-minute ultra-rapid urease test before imprint smears were prepared from them. Thereafter, the biopsies were fixed in 10% formalin and histological sections were examined for the presence of H pylori by a pathologist who was not aware of the clinical details or the results of the urease test. The prevalence of H pylori in the gastric antrum and corpus was 86.7% and 53.3%, respectively. The sensitivity, specificity, positive and negative predictive value and the overall diagnostic accuracy of the ultra-rapid urease test to diagnose H pylori infection in the gastric antrum were 92%, 100%, 100%, 66%, and 93%, respectively. The corresponding figures for the gastric corpus were 83%, 100%, 100%, 85%, and 91%, respectively. It is concluded that the one-minute ultra-rapid urease test has a high sensitivity and specificity and may be used as a rapid and cheap method to diagnose H pylori infection.

Bacteriological Techniques↗
PubMed1987

Purification of urease from Ureaplasma urealyticum.

We have purified urease from the Mollicutes, Ureaplasma urealyticum, using high performance liquid chromatography methods and DEAE-Sephadex chromatography. While only small amounts of material could be utilized in these methods, urease was purified at least 180-fold, yield a major band on SDS-PAGE of 66,000 daltons, a minor band of 64,000 daltons, and several faint bands of lower molecular mass. These results suggest that the 380,000 dalton intact urease is a pentamer or hexamer of these two larger subunits. The highly purified urease from DEAE-Sephadex retained full activity for at least 20 days at 4 degrees C in sodium phosphate buffer (pH 7.2) with 1% bovine serum albumin. The estimated specific activity of the DEAE peak fractions, 180 IU/micrograms, is at least 90-fold greater than that of jack bean urease.

Animals↗
PubMed1995

Helicobacter pylori urease inhibition by rabeprazole, a proton pump inhibitor.

We investigated the inhibitory effects of four gastric proton pump inhibitors (PPIs): rabeprazole, a novel benzimidazole PPI, omeprazole, lansoprazole and AG-2000, on the urease activity of Helicobacter pylori (H. pylori). Their 50% inhibitory concentrations (I50s) were found to be 0.29, 5.4, 9.3 and 0.3 microM respectively. Rabeprazole and omeprazole were also potent inhibitors of Jack bean and Proteus mirabilis cellular ureases. The thioether derivative of rabeprazole, one of its metabolites, had no inhibitory effect on H. pylori urease, despite being reported as a more potent inhibitor of H. pylori growth than rabeprazole. The inhibitory effect of rabeprazole was prevented completely and reversed considerably by the addition of sulfhydryl compounds, such as beta-mercaptoethanol, glutathione and dithiothreitol. Moreover, the addition of beta-mercaptoethanol recovered the urease activity inhibited by rabeprazole. From these results, we expected that rabeprazole inhibited H. pylori urease activity by forming disulfide bonds between it and the active site of the enzyme.

2-Pyridinylmethylsulfinylbenzimidazoles↗
PubMed1996

Kinetic studies of Helicobacter pylori urease inhibition by a novel proton pump inhibitor, rabeprazole.

Urease is an important virulence factor of pathogenicity of gastric Helicobacter pylori. The inhibition of H. pylori urease by the novel proton pump inhibitor, rabeprazole, was investigated kinetically. It was found to act as an irreversible noncompetitive inhibitor of the enzyme. The inhibitory potency of rabeprazole was dependent on the pH of reaction mixture and its Ki values were 0.14 microM (pH 5.0), 0.34 microM (pH 7.0) and 6.10 microM (pH 8.5). Progressive inactivation of urease by rabeprazole initially proceeded according to pseudo-first-order kinetics with respect to the remaining enzymatic activity at pH 7.0 and 37 degrees C, with a second-order rate constant of 0.0017 microM-1 s-1. When the inactivation half-life was plotted versus the reciprocal of the rabeprazole concentration, a straight line was obtained with a slope of -3.12. From an Arrhenius-plot of the temperature-dependence of the inactivation (over the range of 5-37 degrees C), an activation energy of 13.2 kcal/mol was calculated. Recovery of activity was incomplete for H. pylori urease inhibited by rabeprazole, suggesting that the rabeprazole-urease complex is very stable.

2-Pyridinylmethylsulfinylbenzimidazoles↗
PubMed1995

Avirulent, urease-deficient Helicobacter pylori colonizes gastric epithelial explants ex vivo.

BACKGROUND: Urease-negative Helicobacter pylori generated by insertional mutagenesis fails to colonize gnotobiotic piglets, and this effect is largely independent of gastric pH. The purpose of this study was to determine whether urease-negative H. pylori colonized gastric explants ex vivo. METHODS: Gastric mucosal explants derived from neonatal germ-free piglets were inoculated with either wild-type H. pylori or one of two mutants derived by insertional mutagenesis. RESULTS: All three bacterial strains colonized explants. The level of colonization increased over the duration of the experiment, reaching 10(8)-10(9) cfu/g gastric mucosa by 72 h after inoculation. Morphologic evidence of colonization was similar to that observed in gnotobiotic piglets. CONCLUSIONS: Colonization of explants was not affected by lack of urease. These results contrast with previous findings showing that urease activity is essential for colonization of piglets by H. pylori. Thus, urease-dependent colonization is dependent on an intact gastric microenvironment.

Animals↗
PubMed1990

E. coli and urease-induced crystallisation in urine.

The effects of urine preinoculation with E. coli for 20 h on urease activities in urine have been studied in synthetic as well as human urine. The E. coli preinoculation increased pH in both synthetic and human urine. Urease enzymatic activity was enhanced in E. coli-preinoculated synthetic urine. The intraluminal urease-induced precipitation was increased in E. coli-preinoculated synthetic urine. The precipitation on glass rods, which more closely reflects crystal growth and aggregation, was reduced. The process of urease-induced crystallisation thus appears to be influenced by E. coli. In human urine, the effects of E. coli preinoculation were less uniform but a significantly increased urease-induced precipitation after E. coli preinoculation could be reproduced in human urine also.

Adult↗
PubMed1996

Immobilization of urease on activated methoxypolyethyleneglycol-5000.

Urease (E.C 3.5.1.5) was covalently immobilized on activated methoxypolyethyleneglycol-5000 which is linear, uncharged, soluble in water and nonimmunogenic. mPEG is bound to the epsilon-NH2 groups of Lysin in urease. Previously different molar ratios of urease -Lys/activated-mPEG were searched for immobilization. Storage stabilities, molecular weights and the values of blocked amino groups were determined for each immobilized urease and the best conditions was found 1:3 urease-Lys/activated mPEG. Furthermore physical characterization, kinetic constants (Km, Vmax), heat and temperature stabilites were also determined.

Drug Carriers↗
PubMed1977

Rumen bacterial urease requirement for nickel.

Lambs were fed a basal purified diet low in nickel (60 ppb) or the basal diet supplemented with 5 ppm of nickel to determine if rumen bacterial urease was a nickel-requiring enzyme. Two collection periods with lambs fed a diet in which all the nitrogen was supplied as preformed protein (casein) indicated that ruminal urease activity was much lower in lambs fed the low nickel diet. When 1% urea was added to the basal diet, urease activity increased slightly with both treatments; however, bacterial urease activity was still much higher in the lambs receiving 5 ppm of nickel. Ruminal volatile fatty acids were not influenced by dietary nickel. Ruminal urease requires nickel for maximal activity.

Ammonia↗
PubMed1988

Improved growth in the progeny of hens immunized with jackbean urease.

Three experiments were conducted with Single Comb White Leghorn and broiler breeder hens. Half of the experimental hens were injected with jackbean urease enzyme and the other half served as uninjected or adjuvant-injected controls. Chicks hatched from urease-injected or control hens were placed factorially (2 x 2) on diets with or without antibiotics. Urease injection of hens resulted in significant improvements in percent fertility and hatch compared with values for control hens. Depending on age, both urease injection and antibiotic feeding stimulated chick growth, independently of one another. Additive effects of these treatments resulted in significant improvements in body weight of chicks injected with urease and consuming antibiotics at all time periods compared with body weights of untreated controls.

Animals↗