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)

Whole-cell, outer-membrane protein, flagellum-associated antigens and partially purified urease of Campylobacter pylori were analyzed by two-dimensional gel electrophoresis. C. pylori strains were readily distinguished from strains of Campylobacter jejuni, C. coli, and C. fetus by absence of major outer membrane proteins with Mrs of 41,000 to 45,000. C. pylori strains also lacked the acidic surface-array proteins at Mr 100,000 to 149,000 identified previously in serum-resistant strains of C. fetus. Surface labeling of intact C. pylori cells with 125I revealed two common major proteins, which we have designated protein 2 (pI 5.6 to 5.8, Mr 66,000) and protein 3 (pI 5.2 to 5.5, Mr 63,000). Proteins 2 and 3 were also the major components (subunits) observed in partially purified urease. Partially purified preparations of flagella consistently contained proteins 2 and 3. Thus, urease appears to be associated with both outer membranes and flagella of C. pylori. C. pylori strains also possessed an antigen at Mr 59,000 which was cross-reactive with antiserum against flagella of C. jejuni. However, the antigen did not appear to be associated with flagella per se in C. pylori. Protein 2 was unique to C. pylori among the Campylobacter species studied. It was not recognized by antibody against whole cells of C. jejuni or C. fetus or flagella of C. jejuni. Protein 3 was cross-reactive with antiserum against whole cells of C. jejuni and C. fetus, as were several other major protein antigens. Because protein 2 is a major outer membrane protein that is apparently unique to C. pylori, development of monospecific antibodies against this antigen may be useful for the identification of C. pylori in tissues, and purified antigen may be useful for serologic tests for specific diagnosis of C. pylori infections.
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PMID:Two-dimensional gel electrophoresis and immunoblotting of Campylobacter pylori proteins. 272 41

Yersinia enterocolitica is a facultative intracellular parasite, displaying the ability to grow saprophytically or invade and persist intracellularly in the mammalian reticuloendothelial system. The transition between such diverse environments requires the co-ordinated regulation of specific sets of genes on both the chromosome and virulence plasmid. Temperature has a profound pleiotropic effect on gene expression and phenotypically promotes alterations in cell morphology, outer-membrane protein synthesis, urease production, lipopolysaccharide synthesis, motility, and synthesis of genes involved in invasion of eukaryotic host cells. By examining thermoregulated flagella biosynthesis, we have determined that motility is repressed at 25 degrees C (permissive temperature) with subinhibitory concentrations of novobiocin. These conditions also induce virulence gene expression suggesting novobiocin addition simulates, at least partially, a high-temperature environment. Furthermore, temperature-shift experiments, using Y. enterocolitica containing pACYC184 as a reporter plasmid, indicate that thermo-induced alterations of DNA supercoiling coincide with temperature-induced phenotypic changes. A class of putative DNA gyrase mutant (novobiocin resistant) likewise demonstrates the 37 degrees C phenotype when cultured at 25 degrees C; it is non-motile, urease negative, calcium growth dependent, and positive for Yop expression. These results support a model implicating DNA topology as a contributing factor of Y. enterocolitica thermoregulation.
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PMID:Thermoregulation in Yersinia enterocolitica is coincident with changes in DNA supercoiling. 805 44

The present study includes 178 Haemophilus influenzae strains isolated in different pediatric hospitals from Havana, Cuba, during 1991-1994, associated to divers infections (meningitis, respiratory sepsis, primary bacteremia). A combination of various typing and subtyping methods was used as epidemiological markers: serotyping (slide agglutination with diagnostical serum a-f and latex agglutination), biotyping according to Killian's procedures (by determination of indole production, urease and ornithine decarboxylase activity), subtyping by fermentative profiles according to Roberts' methods (glucose, maltose, xylose and fructose) and outer membrane protein profile subtyping (vesicles extraction by a modified Barenkamp's method, analysis by lineal and gradient SDS-PAGE and assessment according to our own classification system). Serotype b was identified in 89.3%, biotype I was the most frequent (79.1%), other biotypes (II, III, IV and V) were also identified. Fermentative profile D (glucose, maltose, xylose and fructose positive) was the most frequent (52.8%) while profile G (glucose, maltose, xylose positive and fructose negative) represented 20.2%. Other known profiles were present. PA2 (33.7%) was the most frequent OMP subtype. Even though 11 different protein subtypes were found, the 77.5% of the strains were located in only three OMP electrophoretic subtypes (PA2, PC1, LA2).
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PMID:[Utilization of different microbiological markers in the study of Haemophilus influenzae]. 902 20

ureI encodes an integral cytoplasmic membrane protein. It is present in the urease gene cluster of Helicobacter pylori and is essential for infection and acid survival, but its role is unknown. To determine the function of UreI protein, we produced H. pylori ureI deletion mutants and measured the pH dependence of urease activity of intact and lysed bacteria and the effect of urea on the membrane potential. We also determined ureI expression, urease activity, and the effect of urea on membrane potential of several gastric and nongastric Helicobacter species. ureI was found to be present in the genome of the gastric Helicobacter species and absent in the nongastric Helicobacter species studied, as determined by PCR. Likewise, Western blot analysis confirmed that UreI was expressed only in the gastric Helicobacter species. When UreI is present, acidic medium pH activation of cytoplasmic urease is found, and urea addition increases membrane potential at acidic pH. The addition of a low concentration of detergent raised urease activity of intact bacteria at neutral pH to that of their homogenates, showing that urease activity was membrane limited. No acidic pH activation or urea induced membrane potential changes were found in the nongastric Helicobacter species. The ureI gene product is probably a pH activated urea transporter or perhaps regulates such a transporter as a function of periplasmic pH.
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PMID:Expression of the Helicobacter pylori ureI gene is required for acidic pH activation of cytoplasmic urease. 1063 6

Acidic media trigger cytoplasmic urease activity of the unique human gastric pathogen Helicobacter pylori. Deletion of ureI prevents this activation of cytoplasmic urease that is essential for bacterial acid resistance. UreI is an inner membrane protein with six transmembrane segments as shown by in vitro transcription/translation and membrane separation. Expression of UreI in Xenopus oocytes results in acid-stimulated urea uptake, with a pH profile similar to activation of cytoplasmic urease. Mutation of periplasmic histidine 123 abolishes stimulation. UreI-mediated transport is urea specific, passive, nonsaturable, nonelectrogenic, and temperature independent. UreI functions as a H+-gated urea channel regulating cytoplasmic urease that is essential for gastric survival and colonization.
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PMID:A H+-gated urea channel: the link between Helicobacter pylori urease and gastric colonization. 1064 49

ureI encodes an inner membrane protein of Helicobacter pylori. The role of the bacterial inner membrane and UreI in acid protection and regulation of cytoplasmic urease activity in the gastric microorganism was studied. The irreversible inhibition of urease when the organism was exposed to a protonophore (3,3',4', 5-tetrachlorsalicylanide; TCS) at acidic pH showed that the inner membrane protected urease from acid. Isogenic ureI knockout mutants of several H. pylori strains were constructed by replacing the ureI gene of the urease gene cluster with a promoterless kanamycin resistance marker gene (kanR). Mutants carrying the modified ureAB-kanR-EFGH operon all showed wild-type levels of urease activity at neutral pH in vitro. The mutants resisted media of pH > 4.0 but not of pH < 4.0. Whereas wild-type bacteria showed high levels of urease activity below pH 4.0, this ability was not retained in the ureI mutants, resulting in inhibition of metabolism and cell death. Gene complementation experiments with plasmid-derived H. pylori ureI restored wild-type properties. The activation of urease activity found in structurally intact but permeabilized bacteria treated with 0.01% detergent (polyoxy-ethylene-8-laurylether; C12E8), suggested a membrane-limited access of urea to internal urease at neutral pH. Measurement of 14C-urea uptake into Xenopus oocytes injected with ureI cRNA showed acid activation of uptake only in injected oocytes. Acceleration of urea uptake by UreI therefore mediates the increase of intracellular urease activity seen under acidic conditions. This increase of urea permeability is essential for H. pylori survival in environments below pH 4.0. ureI-independent urease activity may be sufficient for maintenance of bacterial viability above pH 4.0.
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PMID:Acid resistance of Helicobacter pylori depends on the UreI membrane protein and an inner membrane proton barrier. 1076 Jan 71

Experiments were performed using the standardized murine model of Helicobacter pylori infection to determine the immunogenicity of H. pylori outer membrane vesicles in immune protection. These vesicles, which are naturally shed from the surface of the bacterium, induce a protective response when administered intragastrically to mice in the presence of cholera holotoxin, despite the absence of the urease enzyme and associated Hsp54 chaperonin. Immunoblotting identified a specific serum immunoglobulin G (IgG) response to an 18-kDa outer membrane protein in a significant number of immunized animals. This commonly expressed, immunodominant protein was subsequently identified as lipoprotein 20 (Lpp20). Hybridoma backpacks secreting an IgG1 subclass monoclonal antibody to Lpp20 were generated in H. pylori-infected mice and were found to significantly reduce bacterial numbers, providing evidence that this surface-exposed antigen is a true vaccine candidate and not merely an antigenic marker for successful, protective immunization.
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PMID:Immune response to an 18-kilodalton outer membrane antigen identifies lipoprotein 20 as a Helicobacter pylori vaccine candidate. 1081 82

Considering a suspected link between Helicobacter pylori infection and human stomach cancer, a new H. pylori gene for membrane protein 1 (HP-MP1) was recently cloned. Because HP-MP1 induces release of inflammatory cytokines and tumor necrosis factor-alpha acts as both initiator and tumor promoter, we studied the possible involvement of HP-MP1 in carcinogenesis of H. pylori. Two cell lines, BALB/3T3 cells as control and v-Ha-ras-transfected BALB/3T3 cells (Bhas 42 cells) as putative initiated cells, were each transfected with HP-MP1, urease B genes, or vector alone. All of the Bhas/mpl clones showed strong expression of tumor necrosis factor-alpha gene and produced tumors in 100% of nude mice. Two Bhas/ure clones showed weak tumorigenicity; the other Bhas and BALB clones showed none. Results indicate strong carcinogenic activity of HP-MP1 in cooperation with viral Ras protein and weak activity of urease B.
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PMID:Helicobacter pylori membrane protein 1: a new carcinogenic factor of Helicobacter pylori. 1152 25

Helicobacter pylori is a human gastric pathogen that survives the strong acidity of the stomach by virtue of its urease activity. This activity produces ammonia, which neutralizes the bacterial microenvironment. UreI, an inner membrane protein, is essential for resistance to low pH and for the gastric colonization of mice by H. pylori. In the heterologous Xenopus oocytes expression system, UreI behaves like an H+-gated urea channel, and His-123 was found to be important for low pH activation. We investigated the role of UreI directly in H. pylori and showed that, in the presence of urea, strains expressing wild-type UreI displayed very rapid stimulation of extracellular ammonia production upon exposure to pH </= 5. This response was not observed when acetamide was used as a source of ammonia; therefore, it is specific for urea hydrolysis. To identify residues critical for UreI activity or activation, we constructed H. pylori strains carrying individual chromosomal mutations of UreI (i) in the four conserved histidine residues (H71, H123, H131, H193) and (ii) in a conserved region of the third intracellular loop (L165, G166, K167, F168). The distal H193 (and not H123) was found to be crucial for stimulating the production of ammonia at low pH; a single mutation in this residue uncoupled the UreI activity from its acid activation. The third intracellular loop of UreI was shown to be important for UreI activity. Thus, in H. pylori, UreI is necessary for the adaptation of urease activity to the extracellular pH. UreI behaves like a novel type of urea transporter, and the identification of residues essential for its function in H. pylori provides new insight into the unusual molecular mechanism of low pH activation.
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PMID:The Helicobacter pylori UreI protein: role in adaptation to acidity and identification of residues essential for its activity and for acid activation. 1173 44

Previous studies have demonstrated that Helicobacter pylori (Hp) delays its entry into macrophages and persists inside megasomes, which are poorly acidified and accumulate early endosome autoantigen 1. Herein, we explored the role of Hp urease in bacterial survival in murine peritoneal macrophages and J774 cells. Plasmid-free mutagenesis was used to replace ureA and ureB with chloramphenicol acetyltransferase in Hp Strains 11637 and 11916. ureAB null Hp lacked detectable urease activity and did not express UreA or UreB as judged by immunoblotting. Deletion of ureAB had no effect on Hp binding to macrophages or the rate or extent of phagocytosis. However, intracellular survival of mutant organisms was impaired significantly. Immunofluorescence microscopy demonstrated that (in contrast to parental organisms) mutant Hp resided in single phagosomes, which were acidic and accumulated the lysosome marker lysosome-associated membrane protein-1 but not early endosome autoantigen 1. A similar phenotype was observed for spontaneous urease mutants derived from Hp Strain 60190. Treatment of macrophages with bafilomycin A1, NH4Cl, or chloroquine prevented acidification of phagosomes containing mutant Hp. However, only ammonium chloride enhanced bacterial viability significantly. Rescue of ureAB null organisms was also achieved by surface adsorption of active urease. Altogether, our data indicate a role for urease and urease-derived ammonia in megasome formation and Hp survival.
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PMID:Role of urease in megasome formation and Helicobacter pylori survival in macrophages. 1654 3


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