Journal articles on the topic 'Bacterial urea transport'
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1
Godara, Geeta, Craig Smith, Janine Bosse, Mark Zeidel, and John Mathai. "Functional characterization ofActinobacillus pleuropneumoniaeurea transport protein, ApUT." American Journal of Physiology-Regulatory, Integrative and Comparative Physiology 296, no. 4 (April 2009): R1268—R1273. http://dx.doi.org/10.1152/ajpregu.90726.2008.
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Urea transporters (UTs) effect rapid flux of urea across biological membranes. In the mammalian kidney, UT activity is essential for effective urine concentration. In bacteria, UT-mediated urea uptake permits intracellular urease to degrade urea to ammonia and CO2, a process that either buffers acid loads or provides nutrient nitrogen. We have characterized the urea transport channel protein ApUT from Actinobacillus pleuropneumoniae. Kinetic analysis of bacterial inside-out membranes enriched in ApUT showed ∼28-fold increase in urea permeability (3.3 ± 0.4 × 10−4cm/s) compared with control vesicles (0.11 ± 0.02 × 10−4cm/s). In addition to urea, ApUT also conducts water. Urea and water transport across the channel was phloretin and mercury inhibitable, and the site of inhibition may be located on the cytoplasmic side of the protein. Glycerol and urea analogs, such as methylamine, dimethylurea, formamide, acetamide, methylurea, propanamide, and ethylamine did not permeate across ApUT.
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Scott, David R., Elizabeth A. Marcus, Yi Wen, Siddarth Singh, Jing Feng, and George Sachs. "Cytoplasmic Histidine Kinase (HP0244)-Regulated Assembly of Urease with UreI, a Channel for Urea and Its Metabolites, CO2, NH3, and NH4+, Is Necessary for Acid Survival of Helicobacter pylori." Journal of Bacteriology 192, no. 1 (October 23, 2009): 94–103. http://dx.doi.org/10.1128/jb.00848-09.
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ABSTRACT Helicobacter pylori colonizes the normal human stomach by maintaining both periplasmic and cytoplasmic pH close to neutral in the presence of gastric acidity. Urease activity, urea flux through the pH-gated urea channel, UreI, and periplasmic α-carbonic anhydrase are essential for colonization. Exposure to pH 4.5 for up to 180 min activates total bacterial urease threefold. Within 30 min at pH 4.5, the urease structural subunits, UreA and UreB, and the Ni2+ insertion protein, UreE, are recruited to UreI at the inner membrane. Formation of this complex and urease activation depend on expression of the cytoplasmic sensor histidine kinase, HP0244. Its deletion abolishes urease activation and assembly, impairs cytoplasmic and periplasmic pH homeostasis, and depolarizes the cells, with an ∼7-log loss of survival at pH 2.5, even in 10 mM urea. Associated with this assembly, UreI is able to transport NH3, NH4 +, and CO2, as shown by changes in cytoplasmic pH following exposure to NH4Cl or CO2. To be able to colonize cells in the presence of the highly variable pH of the stomach, the organism expresses two pH-sensor histidine kinases, one, HP0165, responding to a moderate fall in periplasmic pH and the other, HP0244, responding to cytoplasmic acidification at a more acidic medium pH. Assembly of a pH-regulatory complex of active urease with UreI provides an advantage for periplasmic buffering.
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Hong, Wu, Kouichi Sano, Shinichi Morimatsu, David R. Scott, David L. Weeks, George Sachs, Toshiyuki Goto, et al. "Medium pH-dependent redistribution of the urease of Helicobacter pylori." Journal of Medical Microbiology 52, no. 3 (March 1, 2003): 211–16. http://dx.doi.org/10.1099/jmm.0.05072-0.
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Helicobacter pylori is an aetiological agent of gastric disease. Although the role of urease in gastric colonization of H. pylori has been shown, it remains unclear as to where urease is located in this bacterial cell. The purpose of this study was to define the urease-associated apparatus in the H. pylori cytoplasm. H. pylori was incubated at both a neutral and an acidic pH in the presence or absence of urea and examined by double indirect immunoelectron microscopy. The density of gold particles for UreA was greatest in the inner portion of the wild-type H. pylori cytoplasm at neutral pH but was greatest in the outer portion at acidic pH. This difference was independent of the presence of urea and was not observed in the ureI-deletion mutant. Also, the eccentric shift of urease in acidic pH was not observed in UreI. After a 2 day incubation period at acidic pH, it was observed that the urease gold particles in H. pylori assembled and were associated with UreI gold particles. Urease immunoreactivity shifted from the inner to the outer portion of H. pylori as a result of an extracellular decrease in pH. This shift was urea-independent and UreI-dependent, suggesting an additional role of UreI in urease-dependent acid resistance. This is the first report of the intracellular transport of molecules in bacteria in response to changes in the extracellular environment.
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Rodela, T. M., J. S. Ballantyne, and P. A. Wright. "Carrier-mediated urea transport across the mitochondrial membrane of an elasmobranch (Raja erinacea) and a teleost (Oncorhynchus mykiss) fish." American Journal of Physiology-Regulatory, Integrative and Comparative Physiology 294, no. 6 (June 2008): R1947—R1957. http://dx.doi.org/10.1152/ajpregu.00251.2007.
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In osmoregulating teleost fish, urea is a minor nitrogen excretory product, whereas in osmoconforming marine elasmobranchs it serves as the major tissue organic solute and is retained at relatively high concentrations (∼400 mmol/l). We tested the hypothesis that urea transport across liver mitochondria is carrier mediated in both teleost and elasmobranch fishes. Intact liver mitochondria in rainbow trout ( Oncorhynchus mykiss) demonstrated two components of urea uptake, a linear component at high concentrations and a phloretin-sensitive saturable component [Michaelis constant ( Km) = 0.58 mmol/l; maximal velocity ( Vmax) = 0.12 μmol·h−1·mg protein−1] at lower urea concentrations (<5 mmol/l). Similarly, analysis of urea uptake in mitochondria from the little skate ( Raja erinacea) revealed a phloretin-sensitive saturable transport ( Km= 0.34 mmol/l; Vmax= 0.054 μmol·h−1·mg protein−1) at low urea concentrations (<5 mmol/l). Surprisingly, urea transport in skate, but not trout, was sensitive to a variety of classic ionophores and respiration inhibitors, suggesting cation sensitivity. Hence, urea transport was measured in the reverse direction using submitochondrial particles in skate. Transport kinetics, inhibitor response, and pH sensitivity were very similar in skate submitochondrial particle submitochondrial particles ( Km= 0.65 mmol/l, Vmax= 0.058 μmol·h−1·mg protein−1) relative to intact mitochondria. We conclude that urea influx and efflux in skate mitochondria is dependent, in part, on a bidirectional proton-sensitive mechanism similar to bacterial urea transporters and reminiscent of their ancestral origins. Rapid equilibration of urea across the mitochondrial membrane may be vital for cell osmoregulation (elasmobranch) or nitrogen waste excretion (teleost).
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Zhong, Chongliang, Laura L. Griffin, Orla Heussaff, Ruairi O’Dea, Conor Whelan, and Gavin Stewart. "Sex-Related Differences in UT-B Urea Transporter Abundance in Fallow Deer Rumen." Veterinary Sciences 9, no. 2 (February 8, 2022): 73. http://dx.doi.org/10.3390/vetsci9020073.
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Rumen studies have focused almost exclusively on livestock species under strictly regimented diets. This means that the ruminal condition of free-living and free-feeding wildlife remains practically unstudied. Urea nitrogen salvaging, a process by which urea is passed into the rumen, to both provide a valuable source of nitrogen for bacterial growth and to buffer the potentially harmful acidic effects of bacterial short chain fatty acids, has remained unexplored in wild ruminants, such as deer. UT-B2 transporters are the key proteins reported to facilitate the transepithelial ruminal urea transport. In this study, we investigate the expression, abundance and localisation of urea transporters in the rumen of a semi-wild fallow deer (Dama dama) population. Physical measurements confirmed that males had larger rumen than females, while adults had longer papillae than juveniles. Initial RT-PCR experiments confirmed the expression of UT-B2, while immunolocalisation studies revealed that strong UT-B staining was present in the stratum basale of deer rumen. Western blotting analysis demonstrated that a 50 kDa UT-B2 protein was significantly more abundant in adult females compared to adult males. This study confirms the presence of UT-B2 urea transporters in deer rumen and suggests that sex-related differences occur, bringing new insight into our understanding of rumen physiology.
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Skouloubris, Stéphane, Jean-Michel Thiberge, Agnès Labigne, and Hilde De Reuse. "The Helicobacter pylori UreI Protein Is Not Involved in Urease Activity but Is Essential for Bacterial Survival In Vivo." Infection and Immunity 66, no. 9 (September 1, 1998): 4517–21. http://dx.doi.org/10.1128/iai.66.9.4517-4521.1998.
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ABSTRACT We produced defined isogenic Helicobacter pylori ureImutants to investigate the function of UreI, the product of one of the genes of the urease cluster. The insertion of a catcassette had a strong polar effect on the expression of the downstream urease genes, resulting in very weak urease activity. Urease activity, measured in vitro, was normal in a strain in which ureI was almost completely deleted and replaced with a nonpolar cassette. In contrast to previous reports, we thus found that the product ofureI was not necessary for the synthesis of active urease. Experiments with the mouse-adapted H. pylori SS1 strain carrying the nonpolar ureI deletion showed that UreI is essential for H. pylori survival in vivo and/or colonization of the mouse stomach. The replacement of ureIwith the nonpolar cassette strongly reduced H. pylorisurvival in acidic conditions (1-h incubation in phosphate-buffered saline solution at pH 2.2) in the presence of 10 mM urea. UreI is predicted to be an integral membrane protein and may therefore be involved in a transport process essential for H. pylori survival in vivo.
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Zeuthen, Thomas, Edurne Gorraitz, Ka Her, Ernest M. Wright, and Donald D. F. Loo. "Structural and functional significance of water permeation through cotransporters." Proceedings of the National Academy of Sciences 113, no. 44 (October 18, 2016): E6887—E6894. http://dx.doi.org/10.1073/pnas.1613744113.
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Membrane transporters, in addition to their major role as specific carriers for ions and small molecules, can also behave as water channels. However, neither the location of the water pathway in the protein nor their functional importance is known. Here, we map the pathway for water and urea through the intestinal sodium/glucose cotransporter SGLT1. Molecular dynamics simulations using the atomic structure of the bacterial transporter vSGLT suggest that water permeates the same path as Na+ and sugar. On a structural model of SGLT1, based on the homology structure of vSGLT, we identified and mutated residues lining the sugar transport pathway to cysteine. The mutants were expressed in Xenopus oocytes, and the unitary water and urea permeabilities were determined before and after modifying the cysteine side chain with reversible methanethiosulfonate reagents. The results demonstrate that water and urea follow the sugar transport pathway through SGLT1. The changes in permeability, increases or decreases, with side-chain modifications depend on the location of the mutation in the region of external or internal gates, or the sugar binding site. These changes in permeability are hypothesized to be due to alterations in steric hindrance to water and urea, and/or changes in protein folding caused by mismatching of side chains in the water pathway. Water permeation through SGLT1 and other transporters bears directly on the structural mechanism for the transport of polar solutes through these proteins. Finally, in vitro experiments on mouse small intestine show that SGLT1 accounts for two-thirds of the passive water flow across the gut.
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Wang, Jianxiu, Yanxia Long, Yu Zhao, Weiqiang Pan, Jianxun Qu, Tianliang Yang, Xinlei Huang, Xiaotian Liu, and Na Xu. "Laboratory Experiment on Formation of MICP Horizontal Seepage-Reducing Body in Confined Aquifer for Deep Excavation." Applied Sciences 13, no. 1 (December 22, 2022): 104. http://dx.doi.org/10.3390/app13010104.
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Using a soil improvement biotechnology based on microbial-induced carbonate precipitation (MICP) to form a horizontal water barrier in foundation pit dewatering can not only achieve a good dewatering control effect, but also control the cost and obtain good economic and environmental benefits. In this study, taking the Shanghai layer ⑨ sand as the research object, the solution injected into the layer ⑨ sand by MICP grouting technology includes bacterial solution (Sporosarcina pasteurii) and cementation solution (urea and CaCl2 solution). The migration mechanism of the bacterial solution and cementation solution in groundwater was analyzed through batch experiment and column test, and the mathematical model of migration was established. The dynamic adsorption characteristics of the S. pasteurii, Ca2+, and urea were obtained by dynamic adsorption test. Through isothermal adsorption test, the Freundlich isotherm was used to describe the isothermal adsorption of S. pasteurii onto fine sand, and the partition coefficient of the S. pasteurii was found to be 1.5 × 10−7 L/mg. The Langmuir isotherm can be used truly reflect the isothermal adsorption of Ca2+ onto fine sand. The distribution coefficient of the Ca2+ was 7 × 10−8 L/mg. The maximum adsorption capacity of the solute was 2404.8 mg/kg. The adsorption capacity of the urea was very small. According to the sand column test, the vertical dispersion was 11.5 cm. Due to the obvious size effect of dispersion, the sensitivity of dispersion in actual working conditions should be analyzed in a subsequent numerical simulation. The determined solute transport parameters provide support for the further study of the numerical simulation of the solute transport process of the HSRB formed by MICP grouting technology and provide a basis for further field application.
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Sanders, M. R., H. E. Findlay, and P. J. Booth. "Lipid bilayer composition modulates the unfolding free energy of a knotted α-helical membrane protein." Proceedings of the National Academy of Sciences 115, no. 8 (February 5, 2018): E1799—E1808. http://dx.doi.org/10.1073/pnas.1714668115.
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α-Helical membrane proteins have eluded investigation of their thermodynamic stability in lipid bilayers. Reversible denaturation curves have enabled some headway in determining unfolding free energies. However, these parameters have been limited to detergent micelles or lipid bicelles, which do not possess the same mechanical properties as lipid bilayers that comprise the basis of natural membranes. We establish reversible unfolding of the membrane transporter LeuT in lipid bilayers, enabling the comparison of apparent unfolding free energies in different lipid compositions. LeuT is a bacterial ortholog of neurotransmitter transporters and contains a knot within its 12-transmembrane helical structure. Urea is used as a denaturant for LeuT in proteoliposomes, resulting in the loss of up to 30% helical structure depending upon the lipid bilayer composition. Urea unfolding of LeuT in liposomes is reversible, with refolding in the bilayer recovering the original helical structure and transport activity. A linear dependence of the unfolding free energy on urea concentration enables the free energy to be extrapolated to zero denaturant. Increasing lipid headgroup charge or chain lateral pressure increases the thermodynamic stability of LeuT. The mechanical and charge properties of the bilayer also affect the ability of urea to denature the protein. Thus, we not only gain insight to the long–sought-after thermodynamic stability of an α-helical protein in a lipid bilayer but also provide a basis for studies of the folding of knotted proteins in a membrane environment.
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Moran, M. A., R. Belas, M. A. Schell, J. M. González, F. Sun, S. Sun, B. J. Binder, et al. "Ecological Genomics of Marine Roseobacters." Applied and Environmental Microbiology 73, no. 14 (May 25, 2007): 4559–69. http://dx.doi.org/10.1128/aem.02580-06.
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ABSTRACT Bacterioplankton of the marine Roseobacter clade have genomes that reflect a dynamic environment and diverse interactions with marine plankton. Comparative genome sequence analysis of three cultured representatives suggests that cellular requirements for nitrogen are largely provided by regenerated ammonium and organic compounds (polyamines, allophanate, and urea), while typical sources of carbon include amino acids, glyoxylate, and aromatic metabolites. An unexpectedly large number of genes are predicted to encode proteins involved in the production, degradation, and efflux of toxins and metabolites. A mechanism likely involved in cell-to-cell DNA or protein transfer was also discovered: vir-related genes encoding a type IV secretion system typical of bacterial pathogens. These suggest a potential for interacting with neighboring cells and impacting the routing of organic matter into the microbial loop. Genes shared among the three roseobacters and also common in nine draft Roseobacter genomes include those for carbon monoxide oxidation, dimethylsulfoniopropionate demethylation, and aromatic compound degradation. Genes shared with other cultured marine bacteria include those for utilizing sodium gradients, transport and metabolism of sulfate, and osmoregulation.
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Park, Kwon-Sam, Tetsuya Iida, Yoshiharu Yamaichi, Tomohito Oyagi, Koichiro Yamamoto, and Takeshi Honda. "Genetic Characterization of DNA Region Containing the trh and ure Genes of Vibrio parahaemolyticus." Infection and Immunity 68, no. 10 (October 1, 2000): 5742–48. http://dx.doi.org/10.1128/iai.68.10.5742-5748.2000.
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ABSTRACT We have demonstrated that possession of the gene for thermostable direct hemolysin-related hemolysin (trh) coincides with the presence of the urease gene among clinical Vibrio parahaemolyticus strains and that the location of the two genes are in close proximity on the chromosome. Here, we cloned and sequenced the 15,754-bp DNA region containing the trh gene and the gene cluster for urease production from the chromosome of clinicalV. parahaemolyticus (TH3996). We found 16 open reading frames (ORFs) and a lower G+C content (41%) compared with the total genome of this bacterium (46 to 47%). The ure cluster consisted of eight genes, namely, ureDABCEFG andureR. ureR was located 5.2 kb upstream of the other seven genes in the opposite direction. The genetic organization and sequences of the ure genes resembled those found in Proteus mirabilis. Between ureR and the other uregenes, there were five ORFs, which are homologous with the nickel transport operon (nik) of Escherichia coli. We disrupted each of the ureR, ureC, andnikD genes in TH3996 by homologous recombination and analyzed the phenotype of the mutants. In the presence of urea these mutant strains had dramatically less urease activity than the strain they were derived from. Disruption of ureR,nikD, or ureC, however, had no effect on TRH production. The DNA region containing the trh,nik, and ure genes was found in onlytrh-positive strains and not in Kanagawa phenomenon-positive and environmental V. parahaemolyticusstrains. At the end of the region, an insertion sequence-like element existed. These results suggest that the DNA region was introduced intoV. parahaemolyticus in the past through a mechanism mediated by insertion sequences. This is the first reported case that the genes for an ATP-binding cassette-type nickel transport system, which may play a role in nickel transport through bacterial cytoplasmic membrane, are located adjacent to the ure cluster on the genome of an organism.
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Bennett, W. M., L. M. Mela-Riker, D. C. Houghton, D. N. Gilbert, and W. C. Buss. "Microsomal protein synthesis inhibition: an early manifestation of gentamicin nephrotoxicity." American Journal of Physiology-Renal Physiology 255, no. 2 (August 1, 1988): F265—F269. http://dx.doi.org/10.1152/ajprenal.1988.255.2.f265.
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Aminoglycoside antibiotics achieve bacterial killing by binding to bacterial ribosomes and inhibiting protein synthesis. To examine whether similar mechanisms could be present in renal tubular cells prior to the onset of overt proximal tubular necrosis due to these drugs, we isolated microsomes from Fischer rats given 20 mg/kg gentamicin every 12 h subcutaneously for 2 days and from vehicle-injected controls. Concomitant studies of renal structure, function, and mitochondrial respiration were carried out. [3H]leucine incorporation into renal microsomes of treated animals was reduced by 21.9% (P less than 0.01), whereas brain and liver microsomes from the same animals were unaffected. Gentamicin concentration in the renal microsomal preparation was 56 micrograms/ml, a value 7- to 10-fold above concentrations necessary to inhibit bacterial growth. Conventional renal function studies were normal (blood urea, serum creatinine, creatinine clearance). Treated animals showed only a mild reduction of inulin clearance, 0.71 compared with 0.93 ml.min-1.100 g-1 in controls (P less than 0.05), and an increase in urinary excretion of N-acetylglucosaminidase of 20 compared with 14.8 units/l (P less than 0.05). Renal slice transport of p-aminohippuric acid, tetraethylammonium, and the fractional excretion of sodium were well preserved. There was no evidence, as seen by light microscopy, of proximal tubular necrosis. Mitochondrial cytochrome concentrations were normal and respiratory activities only slightly reduced. Processes similar to those responsible for bacterial killing could be involved in experimental gentamicin nephrotoxicity before overt cellular necrosis.
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Calvert, Paul. "3D printing of gels with living photosynthetic algae." MRS Advances 1, no. 36 (2016): 2569–72. http://dx.doi.org/10.1557/adv.2016.455.
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ABSTRACTChlorella is a green, photosynthetic single-celled genus of algae. It can be 3D printed as a suspension in sodium alginate and gelled with calcium solutions. We have made “log pile” structures with channels between the gel lines to allow easy transport of nutrients and products. Under white light and immersed in solutions of bicarbonate and phosphate and urea “plant food” the algae multiply with the gel and produce oxygen at a rate comparable to that reported for suspensions of Chlorella. The system is stable for one or two weeks at least. In principle this can be extended to other plant tissues but there are concerns relating to bacterial and fungal infection and toxicity of the gel components. In addition a tougher gel is needed if this was to be converted to a practical bioreactor system.
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Burall, Laurel S., Janette M. Harro, Xin Li, C. Virginia Lockatell, Stephanie D. Himpsl, J. Richard Hebel, David E. Johnson, and Harry L. T. Mobley. "Proteus mirabilis Genes That Contribute to Pathogenesis of Urinary Tract Infection: Identification of 25 Signature-Tagged Mutants Attenuated at Least 100-Fold." Infection and Immunity 72, no. 5 (May 2004): 2922–38. http://dx.doi.org/10.1128/iai.72.5.2922-2938.2004.
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ABSTRACT Proteus mirabilis, a common cause of urinary tract infections (UTI) in individuals with functional or structural abnormalities or with long-term catheterization, forms bladder and kidney stones as a consequence of urease-mediated urea hydrolysis. Known virulence factors, besides urease, are hemolysin, fimbriae, metalloproteases, and flagella. In this study we utilized the CBA mouse model of ascending UTI to evaluate the colonization of mutants of P. mirabilis HI4320 that were generated by signature-tagged mutagenesis. By performing primary screening of 2,088 P. mirabilis transposon mutants, we identified 502 mutants that ranged from slightly attenuated to unrecoverable. Secondary screening of these mutants revealed that 114 transposon mutants were reproducibly attenuated. Cochallenge of 84 of these single mutants with the parent strain in the mouse model resulted in identification of 37 consistently out-competed P. mirabilis transposon mutants, 25 of which were out-competed >100-fold for colonization of the bladder and/or kidneys by the parent strain. We determined the sequence flanking the site of transposon insertion in 29 attenuated mutants and identified genes affecting motility, iron acquisition, transcriptional regulation, phosphate transport, urease activity, cell surface structure, and key metabolic pathways as requirements for P. mirabilis infection of the urinary tract. Two mutations localized to a ∼42-kb plasmid present in the parent strain, suggesting that the plasmid is important for colonization. Isolation of disrupted genes encoding proteins with homologies to known bacterial virulence factors, especially the urease accessory protein UreF and the disulfide formation protein DsbA, showed that the CBA mouse model and mutant pools are a reliable source of attenuated mutants with mutations in virulence genes.
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Martins, Nathana Beatriz, Lucas Arthur Ricardo Ferreira, Andre Luiz Quagliatto Santos, Rafael Rocha De Souza, Wilson Junior Oliveira, Thaís De Almeida Moreira, Caroline Lopes Queiroz, and Anna Monteiro Correira Lima. "Dermatopathy Caused by Enterobacter aerogenes and Pseudomonas aeruginosa in Boa constrictor amarali." Acta Scientiae Veterinariae 45 (June 27, 2017): 4. http://dx.doi.org/10.22456/1679-9216.86173.
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Background: Bacterial diseases are the main cause of the high mortality rates of snakes, especially those caused by gramnegative agents. However, studies on dermatopathy caused by these bacterial agents in snakes are scarce; and no reports have been found on Enterobacter aerogenes as causative agent of dermatopathy in snake species. Thus, the objective of this study was to describe the clinical signs, and lesion evolution of a dermatopathy in a male snake (Boa constrictor amarali) specimen of approximately seven years old; and to describe the isolation and identification of the Enterobacter aerogenes and Pseudomonas aeruginosa agents involved in the cause of this disease.Case: The Boa constrictor amarali evaluated presented blackened cutaneous lesions in the dorsal, snout-vent and tail regions; and well-defined subcutaneous nodules of 2.0-3.0 cm diameter, with soft consistency, reddish color, cutaneous flaccidity, and areas of scale ulceration in the dorsolateral region. The clinical evaluation of the animal showed dehydration signs and pale mucous membranes. The blackened lesions were subjected to mycological analysis - after procedure of deep scale scraping - which showed presence of septate hyphae. The nodule was punctured for microbiological and biochemical analysis. The sample was collected with a sterilized alginate cotton tip swab, and was stored in a plastic tube containing a semi-solid Stuart transport medium, for microbiological analysis. Then, this sample was incubated in a bacteriological oven at 37°C for 24 h. Typical colonies of Pseudomonas and Enterobacter grew on MacConkey agar medium; these bacteria were identified by the colony morphology and their typical odor. The colonies grown in MacConkey agar were also identified through biochemical tests in the mediums: Phenol red, Lysine, Phenylalanine, Citrate, Urea and SIM (Sulfide, Indole, Motility). The results of these tests were able to confirm and identify the P. aeruginosa and E. aerogenes species. The animal died within 36 h, before the identification of the causative agents of the disease, thus, no pharmacological interference was possible.Discussion: Immunodepression, malnutrition, and temperatures and humidity outside the animal thermal comfort zone, are predisposing factors for the development of bacterial diseases in reptiles. Little information about pathogen agents affecting Boa constrictor specimens in their native area is available; however, captive snakes are subject to a wide variety of diseases - most of which caused or intensified by the captivity conditions. Among the bacteria involved in reptile diseases, few are primary causative agents. In general, clinical bacterial infections tend to be secondary to viral infections. The bacterial agents found in this study are commonly described in scientific literature with location in the oral cavity, differently from the results found in this study. Moreover, the bacterium E. aerogenes has not yet been described in other studies as a causative agent of dermatopathy. Reptiles are considered reservoirs of important zoonotic microorganisms, such as P. aeruginosa, which can be transmitted by fecal contact, bites and wounds. However, the little information on P. aeruginosa in captive reptiles indicates the need for further studies to establish its zoonotic potential. A most adequate management conditions for the snake species could have decreased the severity of the lesions. The occurrence of P. aeruginosa and E. aerogenes found in this work may alert professionals for future clinical suspicions and adequate therapeutic management.
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Gramignoli, Roberto, Aarati R. Ranade, Raman Venkataramanan, and Stephen C. Strom. "Effects of Pro-Inflammatory Cytokines on Hepatic Metabolism in Primary Human Hepatocytes." International Journal of Molecular Sciences 23, no. 23 (November 28, 2022): 14880. http://dx.doi.org/10.3390/ijms232314880.
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Three decades of hepatocyte transplantation have confirmed such a cell-based approach as an adjunct or alternative treatment to solid organ transplantation. Donor cell survival and engraftment were indirectly measured by hepatospecific secretive or released metabolites, such as ammonia metabolism in urea cycle defects. In cases of sepsis or viral infection, ammonia levels can significantly and abruptly increase in these recipients, erroneously implying rejection. Pro-inflammatory cytokines associated with viral or bacterial infections are known to affect many liver functions, including drug-metabolizing enzymes and hepatic transport activities. We examined the influence of pro-inflammatory cytokines in primary human hepatocytes, isolated from both normal donors or patients with metabolic liver diseases. Different measures of hepatocyte functions, including ammonia metabolism and phase 1–3 metabolism, were performed. All the hepatic functions were profoundly and significantly suppressed after exposure to concentrations of from 0.1 to 10 ng/mL of different inflammatory cytokines, alone and in combination. Our data indicate that, like phase I metabolism, suppression of phase II/III and ammonia metabolism occurs in hepatocytes exposed to pro-inflammatory cytokines in the absence of cell death. Such inflammatory events do not necessarily indicate a rejection response or loss of the cell graft, and these systemic inflammatory signals should be carefully considered when the immunosuppressant regiment is reduced or relieved in a hepatocyte transplantation recipient in response to such alleged rejection.
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Pyatchenkov, M. O., A. A. Vlasov, E. V. Sherbakov, A. N. Belskykh, E. V. Kryukov, and A. G. Markov. "Features of assessing the intestinal barrier permeability in chronic kidney disease." Experimental and Clinical Gastroenterology, no. 11 (January 23, 2023): 46–59. http://dx.doi.org/10.31146/1682-8658-ecg-207-11-46-59.
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Numerous studies have shown that changes in the intestinal microbial-tissue complex are a risk factor for the progression of chronic kidney disease (CKD) to end-stage renal disease and, therefore, a potential target for new therapeutic interventions. Thus, reliable and sensitive diagnostic tools for measuring intestinal permeability in the clinical setting are necessary. Modern genome sequencing and multi-omics technologies have established that patients with CKD are characterized by a specific imbalance between the saccharolytic and proteolytic microbiota, contributing to the accumulation of numerous toxic microbial products, such as indoxyl sulphate, p-cresyl sulphate, trimethylamine-N-oxide. Progressive kidney function decline leads to compensatory urea accumulation in the gastrointestinal tract. In the intestinal lumen, urea is hydrolyzed by microbial urease, forming a large amount of ammonium hydroxide, which may be accompanied by disruption of the epithelial barrier integrity with an increase in intestinal permeability for microbial molecules that initiate systemic inflammation. Experimental approaches to studying the intestinal barrier in CKD include the assessment of electrophysiological parameters of the intestinal epithelium and the transport of fluorescently labelled tracers in the Ussing chamber. Actively improving various cell-based in vitro methods, which may be useful for studying the effect of microbiota on the barrier functions of the intestinal epithelium. Gene expression and protein content of tight junctions are estimated using polymerase chain reaction, immunohistochemical methods and Western blotting. Using various biomolecular methods, it was found that renal failure is characterized by the presence of inflammatory and atrophic changes throughout the gastrointestinal tract, destruction of the mucin layer, damage to tight junctions with a decrease in the amount of claudine-1, occludin and ZO-1 as well as a decrease in transepithelial electrical resistance. Clinical examination of intestinal permeability by methods based on the urine excretion of orally administered sugars, polyethylene glycol polymers and labelled tracers indicate a distortion of the results in patients with CKD due to altered renal clearance. Alternatively, quantitative determination of bacterial DNA and D-lactate levels in the blood is considered. Identification of serum non-coding microRNAs, confocal laser endomicroscopy and impedance spectroscopy have the potential to be used as methods for assessing intestinal barrier function.
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Collins, D., D. C. Winter, A. M. Hogan, L. Schirmer, A. W. Baird, and G. S. Stewart. "Differential protein abundance and function of UT-B urea transporters in human colon." American Journal of Physiology-Gastrointestinal and Liver Physiology 298, no. 3 (March 2010): G345—G351. http://dx.doi.org/10.1152/ajpgi.00405.2009.
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Facilitative UT-B urea transporters enable the passage of urea across cell membranes. Gastrointestinal urea transporters are thought to play a significant role in the urea nitrogen salvaging process that occurs between mammalian hosts and their gut bacteria. This study investigated the expression of UT-B urea transporters in different segments of human colon. Immunoblot analysis showed that human colon expressed a 35-kDa glycosylated UT-B protein in the colonic mucosa. The 35-kDa UT-B transporter was predominantly located in plasma membrane-enriched samples ( P < 0.001; n = 6), and its expression was greater in the ascending colon compared with the descending colon ( P < 0.01; n = 3). At the cellular level, UT-B transporters were located throughout colonocytes situated in the upper portion of the colonic crypts. Bidirectional trans-epithelial urea transport was significantly greater in the ascending colon than the descending colon ( P < 0.05; n = 6). In addition, the facilitative urea transporter inhibitor 1,3,dimethylurea significantly reduced urea transport in the ascending colon ( P < 0.05; n = 6) but had no effect in the descending colon (NS; n = 6). These results illustrate differential protein abundance of functional UT-B protein in different sections of the human colon, strongly correlating to regions that contain the largest populations of intestinal bacteria. This study suggests an important role for UT-B urea transporters in maintaining the symbiotic relationship between humans and their gut bacteria.
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Sebbane, Florent, Marie-Andrée Mandrand-Berthelot, and Michel Simonet. "Genes Encoding Specific Nickel Transport Systems Flank the Chromosomal Urease Locus of Pathogenic Yersiniae." Journal of Bacteriology 184, no. 20 (October 15, 2002): 5706–13. http://dx.doi.org/10.1128/jb.184.20.5706-5713.2002.
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ABSTRACT 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 Ni2+ 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.
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Sachs, G., J. A. Kraut, Y. Wen, J. Feng, and D. R. Scott. "Urea Transport in Bacteria: Acid Acclimation by Gastric Helicobacter spp." Journal of Membrane Biology 212, no. 2 (September 2006): 71–82. http://dx.doi.org/10.1007/s00232-006-0867-7.
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Hanuš, O., J. Vegricht, J. Frelich, A. Macek, M. Bjelka, F. Louda, and L. Janů. "Analysis of raw cow milk quality according to free fatty acid contents in the Czech Republic." Czech Journal of Animal Science 53, No. 1 (January 14, 2008): 17–30. http://dx.doi.org/10.17221/2717-cjas.
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The concentration (<I>c</I>) of free fatty acids (FFAs) in milk is an indicator of dairy cow nutrition, milk straining, its bacterial contamination and storage quality. High FFA concentrations (<I>cs</I>) caused by lipolysis can damage the quality properties of milk products. Therefore the FFA content is introduced thanks to an increase in the efficiency of modern analytical methods as a milk quality indicator and as an indicator for its price as well. The goal of this paper was to analyse the FFA relations to the other milk quality indicators. The data set (<I>n</I> = 11 586) was evaluated by regression methods. In November and December the respective FFA means were 0.614 ± 0.458 and 0.835 ± 0.491 mmol/100 g with a relatively high variability of 74.6 and 58.8%. The frequency of unsatisfactory FFA values (> 1.3) was 7.51 and 13.93%. Casein content (<I>r</I> = –0.17; <I>P</I> < 0.01) and crude protein content (<I>r</I> = –0.12; <I>P</I> < 0.01) were related more closely with FFA <I>c</I>. The FFAs can increase by 0.066 mmol/100 g with casein decrease by 0.10%. The FFAs in milk fat can slightly increase by the supply of energy to dairy cows (protein and casein decrease) and rise with the deteriorating health state of mammary gland (lactose, <I>r</I> = –0.14; <I>P</I> < 0.01) as well. The somatic cell count correlated with FFAs more weakly (<I>r</I> = 0.07; <I>P</I> < 0.05), similarly like the total mesophilic bacteria count (<I>r</I> = 0.11; <I>P</I> < 0.01), relatively more closely the psychrotrophic bacteria count (<I>r</I> = 0.27; <I>P</I> < 0.05). The deterioration of almost all hygienic indicators signified an FFA c increase. The urea content correlated with FFAs weakly (<I>r</I> = –0.08; <I>P</I> < 0.05) and the fat content imperceptibly as a component of similar substance like FFAs. The mechanical milk stress led to FFA liberation from fat esters proportionally to the intervention intensity (<I>P</I> < 0.001). Even a relatively small mechanical stress caused by mixing comparable to the current milking technology, milk transport and storage increased the FFA c of milk fat from 1.11 ± 0.19 to 1.80 ± 0.40 mmol/100 g (<I>P</I> < 0.05). The highest experimental stress up to 6.88 ± 0.55 mmol/100 g (<I>P</I> < 0.001).
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Rodionov, Dmitry A., Peter Hebbeln, Mikhail S. Gelfand, and Thomas Eitinger. "Comparative and Functional Genomic Analysis of Prokaryotic Nickel and Cobalt Uptake Transporters: Evidence for a Novel Group of ATP-Binding Cassette Transporters." Journal of Bacteriology 188, no. 1 (January 1, 2006): 317–27. http://dx.doi.org/10.1128/jb.188.1.317-327.2006.
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ABSTRACT The transition metals nickel and cobalt, essential components of many enzymes, are taken up by specific transport systems of several different types. We integrated in silico and in vivo methods for the analysis of various protein families containing both nickel and cobalt transport systems in prokaryotes. For functional annotation of genes, we used two comparative genomic approaches: identification of regulatory signals and analysis of the genomic positions of genes encoding candidate nickel/cobalt transporters. The nickel-responsive repressor NikR regulates many nickel uptake systems, though the NikR-binding signal is divergent in various taxonomic groups of bacteria and archaea. B12 riboswitches regulate most of the candidate cobalt transporters in bacteria. The nickel/cobalt transporter genes are often colocalized with genes for nickel-dependent or coenzyme B12 biosynthesis enzymes. Nickel/cobalt transporters of different families, including the previously known NiCoT, UreH, and HupE/UreJ families of secondary systems and the NikABCDE ABC-type transporters, showed a mosaic distribution in prokaryotic genomes. In silico analyses identified CbiMNQO and NikMNQO as the most widespread groups of microbial transporters for cobalt and nickel ions. These unusual uptake systems contain an ABC protein (CbiO or NikO) but lack an extracytoplasmic solute-binding protein. Experimental analysis confirmed metal transport activity for three members of this family and demonstrated significant activity for a basic module (CbiMN) of the Salmonella enterica serovar Typhimurium transporter.
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Krishnamurthy, Partha, Mary Parlow, Jason B. Zitzer, Nimish B. Vakil, Harry L. T. Mobley, Marilyn Levy, Suhas H. Phadnis, and Bruce E. Dunn. "Helicobacter pylori Containing Only Cytoplasmic Urease Is Susceptible to Acid." Infection and Immunity 66, no. 11 (November 1, 1998): 5060–66. http://dx.doi.org/10.1128/iai.66.11.5060-5066.1998.
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ABSTRACT Helicobacter pylori, an important etiologic agent in a variety of gastroduodenal diseases, produces large amounts of urease as an essential colonization factor. We have demonstrated previously that urease is located within the cytoplasm and on the surface of H. pylori both in vivo and in stationary-phase culture. The purpose of the present study was to assess the relative contributions of cytoplasmic and surface-localized urease to the ability of H. pylori to survive exposure to acid in the presence of urea. Toward this end, we compared the acid resistance in vitro of H. pylori cells which possessed only cytoplasmic urease to that of bacteria which possessed both cytoplasmic and surface-localized or extracellular urease. Bacteria with only cytoplasmic urease activity were generated by using freshly subcultured bacteria or by treating repeatedly subcultured H. pylori with flurofamide (1 μM), a potent, but poorly diffusible urease inhibitor. H. pyloriwith cytoplasmic and surface-located urease activity survived in an acid environment when 5 mM urea was present. In contrast, H. pylori with only cytoplasmic urease shows significantly reduced survival when exposed to acid in the presence of 5 mM urea. Similarly,Escherichia coli SE5000 expressing H. pyloriurease and the Ni2+ transport protein NixA, which expresses cytoplasmic urease activity at levels similar to those in wild-typeH. pylori, survived minimally when exposed to acid in the presence of 5 to 50 mM urea. We conclude that cytoplasmic urease activity alone is not sufficient (although cytoplasmic urease activity is likely to be necessary) to allow survival of H. pyloriin acid; the activity of surface-localized urease is essential for resistance of H. pylori to acid under the assay conditions used. Therefore, the mechanism whereby urease becomes associated with the surface of H. pylori, which involves release of the enzyme from bacteria due to autolysis followed by adsorption of the enzyme to the surface of intact bacteria (“altruistic autolysis”), is essential for survival of H. pylori in an acid environment. The ability of H. pylori to survive exposure to low pH is likely to depend on a combination of both cytoplasmic and surface-associated urease activities.
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Wiklund, E., C. Rehbinder, G. Malmfors, I. Hansson, and M.-L. Danielsson-Tham. "Ultimate pH values and bacteriological condition of meat and stress metabolites in blood of transported reindeer bulls." Rangifer 21, no. 1 (March 1, 2001): 3. http://dx.doi.org/10.7557/2.21.1.1522.
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Twenty-three reindeer bulls, aged 2-3 years, fed during two winter months at the Vuolda reindeer research station in Arjeplog, Sweden, were used in the study. The first group of eight reindeer was moved from their feeding corral to a selection corral, captured by lasso and stunned with a captive bolt outside the selection corral. The second group of seven reindeer was moved to the selection corral, captured by lasso and restrained, after which they were loaded onto a lorry- and transported for 1 hour and then slaughtered. The third group of eight reindeer was moved to the selection corral and herded directly onto the lorry, without any manual handling. They were transported for 5 h and then slaughtered. In both transport groups, four reindeer were fitted with pre-programmed automatic blood sampling equipment (ABSE). ABSE sampled blood at predetermined times via a jugular vein catheter. Ultimate pH-values in three muscles (Mm. longissimus, triceps brachii and biceps femoris) were significantly lower in the group carefully handled and transported for 5 h compared with the other two groups. The physiological mechanisms behind these results are discussed. Samples from M. semimembranosus were collected at slaughter and after 2, 6 and 10 days of refrigerated storage (+4 °C). The samples were analysed for total counts of aerobic bacteria (pour-plated in Tryptone Glucose Extract Agar, Difco, incubated at 20 °C and 30 °C, respectively for 72 h), coliform bacteria 37 °C (pour-plated in Violet Red Bile Agar, Oxoid, incubated at 37 °C for 24 h), Enterococci (surface-plated onto Slantez and Bartley Agar, Oxoid, incubated at 44 °C for 48 h) and Bacillus cereus (surface-plated onto Blood Agar Plates (Blood Agar Base, Difco, supplemented with 5% defibrinated horse blood) 30 °C for 24 h). All samples fell in the range 'fit for consumption'. At slaughter, there was no difference in ASAT activity, urea and Cortisol concentrations between the two transported groups. However, the plasma ASAT activity and urea concentrations at slaughter were significantly lower in the non-transported group. In both transport groups, the plasma Cortisol concentrations increased during loading onto and unloading from the lorry. Abomasal lesions were observed in all treatment groups. It was concluded that reindeer showed an acute stress response to manual handling and transport.
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Flores Castellanos, Ana Sofía, Lucio Rodríguez Sifuentes, Yeni Natalia Carolina Pérez Gelvez, and Gerardo Gutiérrez Sánchez. "Influencia de fuentes de fosfato inorgánico en la expresión de proteínas de una cepa bacteriana solubilizadora de fosfato nativa de La Comarca Lagunera." Nova Scientia 10, no. 21 (September 7, 2018): 173–90. http://dx.doi.org/10.21640/ns.v10i21.1464.
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Introducción: El presente trabajo tuvo como objetivo identificar las proteínas implicadas en la solubilizacion de fosfatos inorgánicos (Pi) en Klebsiella aerogenes y evaluar la capacidad de solubilizacion de Pi bajo la influencia de fuentes de carbono (C) y nitrógeno (N) en bacterias aisladas de la Comarca Lagunera.Método: Se trabajó con ocho cepas bacterianas aisladas de cultivos forrajeros de La Comarca Lagunera y conservadas en glicerol a -20 °C. Se determinó el Índice de Solubilización (IS) en medio sólido Pikovskaya (PVK) y se cuantificó la solubilización de Pi en medio líquido con diferentes fuentes de C, N y fósforo (P). Se realizó una identificación molecular de la cepa que presentó la mayor capacidad para solubilizar el Pi. Para los estudios de expresión de proteínas, se realizaron cultivos bacterianos por duplicado con dos diferentes fuentes de Pi {(fosfato de calcio (FC) y fosfato de aluminio (FA)} en medio líquido PVK. Se llevó a cabo una precipitación de proteínas con ácido tricloroacético (TCA) y una cuantificación con BCATM Protein assay kit. Las proteínas se corrieron en gel de poliacrilamida al 12%, se realizó una digestión con la enzima tripsina y se analizaron los péptidos obtenidos por HPLC-espectrometría de masas. La identificación de la función de las proteínas se llevó a cabo mediante el uso de herramientas bioinformáticas.Resultados: K. aerogenes fue la bacteria que presentó una mayor capacidad para solubilizar FC (2.31 y 2.59) en medio sólido PVK Glucosa (G)-Urea (U) y G- Nitrato de Amonio (NA) respectivamente, FC (1.97 ppm) y FA (1.36 ppm) en medio líquido PVK U-G. En los cultivos bacterianos se identificaron doscientas dieciséis proteínas entre las cuales se encontraron: ABC transportadoras, histidina quinasa Pho, proteína del sistema de transporte específico de fosfato Pho U, involucradas con el aprovechamiento del P.Discusión o Conclusión: Las fuentes de C y N influyen en la capacidad de solubilización de Pi en bacterias nativas de La Comarca Lagunera. Los mejores resultados de solubilización se presentaron cuando la G se encontraba en el medio como fuente de C y la U como fuente de N. Bajo las condiciones de experimentación, no se observaron proteínas únicas de K. aerogenes para solubilizar el FC y FA. Se observó únicamente diferencias en el nivel de expresión de algunas proteínas identificadas.
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Konopleva, Marina Y., Philip L. Lorenzi, Sanaz Ghotbaldini, Yoko Tabe, Tianyu Cai, and Stefano Tiziani. "Contribution of Amino Acid Metabolism to Hematologic Malignancies." Blood 132, Supplement 1 (November 29, 2018): SCI—10—SCI—10. http://dx.doi.org/10.1182/blood-2018-99-109469.
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Abstract Tumor cells rewire metabolic pathways to meet the high metabolic demands of proliferation, frequently developing auxotrophy to specific amino acid(s) (AAs) required to satisfy protein biosynthesis. Thus specific metabolic inhibitors or AA-depleting enzymes have been developed and tested as cancer therapeutics. For example, depletion of asparagine by bacterial L-asparaginase (ASNase) has proven efficacious against hematologic malignancies, especially leukemia and lymphoma, by starving tumors lacking asparagine synthetase (ASNS). We and others have reported that the glutaminase (GLS) activity of ASNase is required for anticancer activity against ASNS-positive leukemia cell types in vitro.1 In vivo, we have found that durable response to ASNase in pre-clinical models of leukemia also requires glutaminase activity, even against ASNS-negative leukemia models; a glutaminase-deficient mutant of ASNase yielded subsequent leukemia recurrence. We speculate that the underlying anti-leukemia mechanism mediated by ASNase glutaminase activity involves a deeper depletion of asparagine within the tumor microenvironment, since ASNS in nearby cells (adipocytes, mesenchymal stromal cells, etc.) can use glutamine as a precursor for asparagine synthesis. Nevertheless, since L-glutamine depletion is thought to cause the significant side effects of ASNase, enzyme variants with reduced glutaminase coactivity are being developed and tested. Another viable therapeutic strategy involving glutamine starvation via GLS inhibitor has shown significant pre-clinical activity in acute myeloid leukemia (AML) and multiple myeloma (MM) models; this approach is synergistic with hypomethylating agents and BCL2 inhibitors in AML, and with proteasome inhibitors in MM. Recent findings highlight the switch to glutamine metabolism as a metabolic dependency of tyrosine kinase-driven AML, and targeting GLS in conjunction with tyrosine kinase inhibition has been proposed.2 Targeting arginine metabolism has been shown to be another viable therapeutic strategy. Arginine (ARG) depletion using pegylated arginine deaminase (ADI-PEG 20) or pegylated arginase (PEG-ARGase), the 2 critical enzymes of the ARG metabolism/urea cycle, reduced leukemia tumor burden in AML models characterized by low arginosuccinate synthetase (ASS) and high uptake of ARG. However, recently reported Phase I/II clinical trials of recombinant PEG-arginase and of ADI-PEG 20 showed minimal efficacy in relapsed/refractory AML and in solid tumors despite efficient depletion of arginine and low ASS1 expression in tumors, indicating that depletion of arginine alone is insufficient for clinical activity. As a final example of AA metabolic pathways targeted in the treatment of hematologic malignancies, exogenous L-cysteine is required for the synthesis of glutathione for antioxidant cellular defense. In pre-clinical studies, multiple malignancy subtypes were sensitive to cysteine and cystine degradation by an engineered human cyst(e)inase enzyme, including AML, acute lymphocytic leukemia, poor-risk chronic lymphocytic leukemia (CLL), and MM.3 In all therapeutic strategies targeting AA metabolism, the tumor microenvironment may contribute to resistance. For example, bone marrow stromal cells efficiently import cystine, convert it to cysteine, and transport it to CLL cells, facilitating leukemia chemoresistance. Mesenchymal stromal cells and bone marrow adipocytes secrete asparagine and glutamine, respectively, and protect leukemia cells from ASNase cytotoxicity. Recent insights into the immune tumor microenvironment highlight interplay between tumor, AAs, and immune cell functions. Some AAs, such as arginine and glutamine, are essential nutrients for immune cell proliferation and metabolism; excessive tumor consumption of glutamine, or secretion of arginase by myeloid-derived suppressor cells or AML blasts, may deprive immune cells, impair T cell proliferation, and induce immunosuppressive phenotypes. GLS inhibitors that block glutamine consumption and arginase inhibitors that increase plasma arginine, increase availability of their respective target nutrients for immune cells and are, therefore, being explored in ongoing clinical trials as monotherapies and in combination with anti-PD1 blockade. Chan WK, Lorenzi PL, Anishkin A, et al. The glutaminase activity of L-asparaginase is not required for anticancer activity against ASNS-negative cells. Blood. 2014;123:3596-3606. Gallipoli P, Giotopoulos G, Tzelepis K, et al. Glutaminolysis is a metabolic dependency in FLT3(ITD) acute myeloid leukemia unmasked by FLT3 tyrosine kinase inhibition. Blood. 2018;131:1639-1653. Zhang W, Trachootham D, Liu J, et al. Stromal control of cystine metabolism promotes cancer cell survival in chronic lymphocytic leukaemia. Nat Cell Biol. 2012;14:276-286. Disclosures Konopleva: Stemline Therapeutics: Research Funding. Lorenzi:Erytech Pharma: Consultancy; NIH: Patents & Royalties.
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Wilson, MR, JM Hush, and NA Walker. "The involvement of Microbial Epiphytes in Uptake Measurements with the Giant-celled Alga Chara australis." Functional Plant Biology 15, no. 4 (1988): 483. http://dx.doi.org/10.1071/pp9880483.
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The giant-celled algae Chara and Nitella have been used extensively in studies of the transport of ions and other solutes. Before this report, uptake measurements were made without a detailed consideration of the effects of microbial epiphytes living on the external surface of the algal cell wall. Examination of the cell walls of Chara and Nitella with the scanning electron microscope revealed the presence of an often dense microflora consisting of diatoms, many types of bacteria, and even fungi. The contribution of this microflora to uptake measurements made with radiotracers was estimated by using isolated Chara cell wall cylinders internally filled with a silicone compound. For the Chara tested by us, the microflora was shown to make a substantial contribution to the uptake of urea, uric acid and glycine measured for untreated internodes. It was shown that, by briefly soaking Chara internodes in an artificial pond water, pH 8.5, containing 3 mM EGTA and then wiping their surfaces with a Kimwipe (a lint-free paper tissue), the microbial epiphytes can be quickly and simply removed. This treatment could be very useful in radiotracer studies of giant-celled algae in which the activity of the surface microflora complicates measurements of solute uptake.
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Stepanova, S. I., V. V. Boynik, T. M. Gontova, and S. A. Kozyra. "Determination of ureid content in shoots of russian pea shrub and siberian pea tree." Farmatsevtychnyi zhurnal, no. 2 (May 7, 2020): 59–66. http://dx.doi.org/10.32352/0367-3057.2.20.06.
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Among natural ureides, allantoin (5-ureidohydantoin or 2,5-dioxoimidazolidin-4-yl urea) is the most famous compound which have been found in various organisms: bacteria, plants, and animals. Allantoin is a product of purine catabolism in mammals (except for primates and humans) and insect larvae. It is found in many plants: comfrey (Symphytum officinale L.), soybean (Glycine hispida (Moench) Maxim), alfalfa (Medicago sativa L.), purslane (Portulaca oleracea L.).
Allantoin has a reparative, antimicrobial, and anti-inflammatory effect, that substantiate it use in the composition of numerous cosmetic and therapeutic dermatological agents. It can be used also used for the treatment of gastroduodenal ulcers and chronic gastritis. In most species of the legume family allantoin is a transport form of nitrogen, besides, its role in the adaptation of plants to stress factors is known.
The plants of the genus Caragana (Caragana) of the legume family (Fabaceae), due to nitrogen-fixing bacteria, receive nitrogen from the soil and convert it into transport forms, namely allantoin and allantoic acid. These substances were previously isolated from the shoots of russian pea shrub (Caragana frutex (L.) C. Koch) and siberian pea tree (Caragana arborescens Lam.) and identified, but the quantitative content of ureids in these raw materials has not been studied.
The aim of the work was to determine the content of ureides in the shoots of russian pea shrub tree and siberian pea tree.
The objects of study were shoots of russian pea shrub tree and siberian pea tree harvested in Kharkov in May 2019 (during flowering period). The total ureide content was determined by spectrophotometric method based on the reaction of chemical decomposition of allantoin to glyoxylic acid with subsequent formation of coloured phenylhydrazone, which had absorption maximum at 530 nm.
The study results have shown that the content of ureides (expressed as allantoin) is slightly lower in the shoots of russian pea shrub than in the shoots of siberian pea tree and equals 0.34 ± 0.03% and 0.41 ± 0.02% respectively.
The content of total ureids in the shoots of russian pea shrub and siberian pea tree was determined. The flowering shoots of these plants can be used as herbal raw material for the development of therapeutic and prophylactic dermatological and antiulcer drugs possessing reparative, anti-inflammatory, and astringent effects.
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Benz, Roland, Claudio Piselli, and Andrew Potter. "Channel Formation by LktA of Mannheimia (Pasteurella) haemolytica in Lipid Bilayer Membranes and Comparison of Channel Properties with Other RTX-Cytolysins." Toxins 11, no. 10 (October 17, 2019): 604. http://dx.doi.org/10.3390/toxins11100604.
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Cytolysin LktA is one of the major pathogenicity factors of Mannheimia haemolytica (formerly Pasteurella haemolytica) that is the cause of pasteurellosis, also known as shipping fever pneumonia, causing substantial loss of sheep and cattle during transport. LktA belongs to the family of RTX-toxins (Repeats in ToXins) that are produced as pathogenicity factors by a variety of Gram-negative bacteria. Sublytic concentrations of LktA cause inflammatory responses of ovine leukocytes. Higher concentrations result in formation of transmembrane channels in target cells that may cause cell lysis and apoptosis. In this study we investigated channel formation by LktA in artificial lipid bilayer membranes made of different lipids. LktA purified from culture supernatants by polyethylene glycol 4000 precipitation and lyophilization had to be activated to frequently form channels by solution in 6 M urea. The LktA channels had a single-channel conductance of about 60 pS in 0.1 M KCl, which is about one tenth of the conductance of most RTX-toxins with the exception of adenylate cyclase toxin of Bordetella pertussis. The LktA channels are highly cation-selective caused by negative net charges. The theoretical treatment of the conductance of LktA as a function of the bulk aqueous concentration allowed a rough estimate of the channel diameter, which is around 1.5 nm. The size of the LktA channel is discussed with respect to channels formed by other RTX-toxins. We present here the first investigation of LktA in a reconstituted system.
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Li, Wei, Ashutosh Upadhyay, Fabio L. Fontes, E. Jeffrey North, Yuehong Wang, Debbie C. Crans, Anna E. Grzegorzewicz, et al. "Novel Insights into the Mechanism of Inhibition of MmpL3, a Target of Multiple Pharmacophores in Mycobacterium tuberculosis." Antimicrobial Agents and Chemotherapy 58, no. 11 (August 18, 2014): 6413–23. http://dx.doi.org/10.1128/aac.03229-14.
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ABSTRACTMmpL3, a resistance-nodulation-division (RND) superfamily transporter, has been implicated in the formation of the outer membrane ofMycobacterium tuberculosis; specifically, MmpL3 is required for the export of mycolic acids in the form of trehalose monomycolates (TMM) to the periplasmic space or outer membrane ofM. tuberculosis. Recently, seven series of inhibitors identified by whole-cell screening againstM. tuberculosis, including the antituberculosis drug candidate SQ109, were shown to abolish MmpL3-mediated TMM export. However, this mode of action was brought into question by the broad-spectrum activities of some of these inhibitors against a variety of bacterial and fungal pathogens that do not synthesize mycolic acids. This observation, coupled with the ability of three of these classes of inhibitors to kill nonreplicatingM. tuberculosisbacilli, led us to investigate alternative mechanisms of action. Our results indicate that the inhibitory effects of adamantyl ureas, indolecarboxamides, tetrahydropyrazolopyrimidines, and the 1,5-diarylpyrrole BM212 on the transport activity of MmpL3 in actively replicatingM. tuberculosisbacilli are, like that of SQ109, most likely due to their ability to dissipate the transmembrane electrochemical proton gradient. In addition to providing novel insights into the modes of action of compounds reported to inhibit MmpL3, our results provide the first explanation for the large number of pharmacophores that apparently target this essential inner membrane transporter.
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Morrow, Carl A., and James A. Fraser. "Is the Nickel-Dependent Urease Complex of Cryptococcus the Pathogen’s Achilles’ Heel?" mBio 4, no. 4 (June 25, 2013). http://dx.doi.org/10.1128/mbio.00408-13.
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ABSTRACT The nitrogen-scavenging enzyme urease has been coopted in a variety of pathogenic organisms as a virulence factor, most notoriously to neutralize stomach acid and establish infection by the gastric pathogen Helicobacter pylori. The opportunistic fungal pathogen Cryptococcus neoformans also utilizes urease as a virulence factor, only in this case to invade the central nervous system (CNS) via the blood-brain barrier and cause life-threatening meningoencephalitis. A recent study [A. Singh, R. Panting, A. Varma, T. Saijo, K. Waldron, A. Jong, P. Ngamskulrungroj, Y. Chan, J. Rutherford, K. Kwon-Chung, mBio 4(3):e00220-13] genetically and biochemically characterizes the accessory proteins required for successful activation of the urease protein complex, including the essential nickel cofactor. The accessory proteins Ure4, Ure6, and Ure7 are all essential for urease function. Ure7 appears to combine the roles of two bacterial accessory proteins: it incorporates both the GTPase activity and nickel chaperone properties of UreE, a bacterial protein whose homolog is missing in the fungi. An accompanying nickel transporter, Nic1, is responsible for most, but not all, nickel uptake into the fungal cell. Mutants of the core urease protein Ure1, accessory protein Ure7, and transporter Nic1 are all attenuated for invasion of the CNS of mice, and urease activity may directly affect integrity of the tight junction of the endothelial cells of the blood-brain barrier, the network of proteins that limits paracellular permeability. This work highlights the potential of urease, its accessory proteins, and nickel transport as potential chemotherapeutic targets.
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Breusing, Corinna, N. Hagen Klobusnik, Michelle A. Hauer, and Roxanne A. Beinart. "Genome assembly of the chemosynthetic endosymbiont of the hydrothermal vent snail Alviniconcha adamantis from the Mariana Arc." G3 Genes|Genomes|Genetics, August 23, 2022. http://dx.doi.org/10.1093/g3journal/jkac220.
Full textAbstract:
Abstract Chemosynthetic animal-microbe symbioses sustain hydrothermal vent communities in the global deep sea. In the Indo-Pacific Ocean, hydrothermal ecosystems are often dominated by gastropod species of the genus Alviniconcha, which live in association with chemosynthetic Gammaproteobacteria or Campylobacteria. While the symbiont genomes of most extant Alviniconcha species have been sequenced, no genome information is currently available for the gammaproteobacterial endosymbiont of A. adamantis – a comparatively shallow living species that is thought to be the ancestor to all other present Alviniconcha lineages. Here, we report the first genome sequence for the symbiont of A. adamantis from the Chamorro Seamount at the Mariana Arc. Our phylogenomic analyses show that the A. adamantis symbiont is most closely related to Chromatiaceae endosymbionts of the hydrothermal vent snails A. strummeri and Chrysomallon squamiferum, but represents a distinct bacterial species or possibly genus. Overall, functional capacity of the A. adamantis symbiont appeared to be similar to other chemosynthetic Gammaproteobacteria, though several flagella and chemotaxis genes were detected, which are absent in other gammaproteobacterial Alviniconcha symbionts. These differences might suggest potential contrasts in symbiont transmission dynamics, host recognition or nutrient transfer. Furthermore, an abundance of genes for ammonia transport and urea usage could indicate adaptations to the oligotrophic waters of the Mariana region, possibly via recycling of host- and environment-derived nitrogenous waste products. This genome assembly adds to the growing genomic resources for chemosynthetic bacteria from hydrothermal vents and will be valuable for future comparative genomic analyses assessing gene content evolution in relation to environment and symbiotic lifestyle.
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Tang, Jin-Ming, Min Nina Xu, Yuxuan Lin, Huangxin Chen, Haoquan Jin, Li-Li Han, Wenbin Zou, and Shuh-Ji Kao. "The biological transformation of ammonium and urea in a eutrophic estuarine system in Southern China." Frontiers in Marine Science 9 (December 12, 2022). http://dx.doi.org/10.3389/fmars.2022.1040554.
Full textAbstract:
Estuaries channel large amounts of anthropogenic nitrogen (N) from continents to the offshore where productivity is widely limited by N and phosphorus. Ammonium and urea, two reduced forms of anthropogenic N commonly observed, are the preferred substrates for various microorganisms (e.g., uptake by phytoplankton or bacteria and oxidation by nitrifier). Yet, it remains underexplored how they transform and their concentrations vary during transport in the estuary which may influence the offshore microbial community. We applied 15N isotope tracer incubation techniques to determine the two main bio-consumption processes, i.e., uptake and oxidation, of ammonium and urea, in the dark for the Jiulong River Estuary, a eutrophic estuary in southeastern China. Results showed that light penetration depth ranged from 0.8-3.3 m leaving 76% of estuary water bodies to stay in dark situations throughout a day. Ammonia oxidation, which favors dark conditions, dominates the estuarine regenerated-N cycle, showing the rank: ammonia oxidation > ammonium uptake >> urea uptake ≈ urea oxidation. By compiling the reported case studies, we found the relatively low ratio of urea oxidation to ammonia oxidation was accompanied by a relatively high ammonium concentration. Microorganisms’ high preference for ammonium over urea may lead to an inhibitory-like phenomenon. An analogous effect was evident by the increased urea uptake at downstream when ammonium was down to a few µM. The obstructed urea utilization resulted in 10 times longer lifetime for urea relative to ammonium (surface: 19 ± 9 days; bottom 12 ± 7 days). Such an inhibitory-like effect allows urea to be preserved in the estuary and allows urea to be transported farther offshore to stimulate microorganisms capable of utilizing urea, which may have significant impacts on offshore ecology.
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Gajewski, Adrian, Eliza Mnich, Karol Szymański, Krzysztof Hinc, Michał Obuchowski, Anthony P. Moran, and Magdalena Chmiela. "Helicobacter pylori antigens, acetylsalicylic acid, LDL and 7-ketocholesterol - their potential role in destabilizing the gastric epithelial cell barrier. An in vitro model of Kato III cells." Acta Biochimica Polonica 63, no. 1 (November 30, 2015). http://dx.doi.org/10.18388/abp.2015_1122.
Full textAbstract:
Colonization of gastric tissue in humans by H. pylori Gram-negative bacteria initiates gastric and duodenal ulcers and even gastric cancers. Infections promote inflammation and damage to gastric epithelium which might be followed by the impairment of its barrier function. The role of H. pylori components in these processes has not been specified. H. pylori cytotoxicity may potentially increase in the milieu of anti-inflammatory drugs including acetylsalicylic acid (ASA). The lipid transport-associated molecule such as low density lipoprotein (LDL), which is a classic risk factor of coronary heart disease (CHD) and 7-ketocholesterol (7-kCh) a product of cholesterol oxidation, which may occur during the oxidative stress in LDL could also be considered as pro-inflammatory. The aim of this study was to evaluate the cytotoxicity of H. pylori antigens, ASA, LDL and 7-kCh towards Kato III gastric epithelial cells, on the basis of the cell ability to reduce tetrazolium salt (MTT) and morphology of cell nuclei assessed by 4',6-diamidino-2-phenylindole (DAPI) staining. Kato III cells were stimulated for 24 h, at 37°C and 5% CO2, with H. pylori antigens: cytotoxin associated gene A (CagA) protein, the urease A subunit (UreA), lipopolysaccharide (LPS) and ASA, LDL or 7-kCh. H. pylori LPS, ASA, LDL and 7-kCh, but not H. pylori glycine acid extract (GE), demonstrated cytotoxicity against Kato III cells, which was related to a diminished percentage of MTT reducing cells and to an increased cell population with the signs of DNA damage. The results suggest that damage to gastric epithelial cells can be induced independently by H. pylori antigens, ASA and endogenous lipid transport-associated molecules. During H. pylori infection in vivo, especially in CHD patients, synergistic or antagonistic interactions between these factors might possibly influence the disease course. Further study is necessary to explain these potential effects.
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Kozmin, Stanislav G., Elena I. Stepchenkova, Stephen C. Chow, and Roel M. Schaaper. "A Critical Role for the Putative NCS2 Nucleobase Permease YjcD in the Sensitivity of Escherichia coli to Cytotoxic and Mutagenic Purine Analogs." mBio 4, no. 6 (October 29, 2013). http://dx.doi.org/10.1128/mbio.00661-13.
Full textAbstract:
ABSTRACTThe base analogs 6-N-hydroxylaminopurine (HAP) and 2-amino-HAP (AHAP) are potent mutagens in bacteria and eukaryotic organisms. Previously, we demonstrated that a defect in theEscherichia coli ycbXgene, encoding a molybdenum cofactor-dependent oxidoreductase, dramatically enhances sensitivity to the toxic and mutagenic action of these agents. In the present study, we describe the discovery and properties of a novel suppressor locus,yjcD, that strongly reduces the HAP sensitivity of theycbXstrain. Suppressor effects are also observed for other purine analogs, like AHAP, 6-mercaptopurine, 6-thioguanine, and 2-aminopurine. In contrast, theyjcDdefect did not affect the sensitivity to the pyrimidine analog 5-fluorouracil. Homology searches have predicted thatyjcDencodes a putative permease of the NCS2 family of nucleobase transporters. We further investigated the effects of inactivation of all other members of the NCS2 family, XanQ, XanP, PurP, UacT, UraA, RutG, YgfQ, YicO, and YbbY, and of the NCS1 family nucleobase permeases CodB and YbbW. None of these other defects significantly affected sensitivity to either HAP or AHAP. The combined data strongly suggest that YjcD is the primary importer for modified purine bases. We also present data showing that this protein may, in fact, also be a principal permease involved in transport of the normal purines guanine, hypoxanthine, and/or xanthine.IMPORTANCENucleotide metabolism is a critical aspect of the overall metabolism of the cell, as it is central to the core processes of RNA and DNA synthesis. At the same time, nucleotide metabolism can be subverted by analogs of the normal DNA or RNA bases, leading to highly toxic and mutagenic effects. Thus, understanding how cells process both normal and modified bases is of fundamental importance. This work describes a novel suppressor of the toxicity of certain modified purine bases in the bacteriumEscherichia coli. This suppressor encodes a putative high-affinity nucleobase transporter that mediates the import of the modified purine bases. It is also a likely candidate for the long-sought high-affinity importer for the normal purines, like guanine and hypoxanthine.