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Peter C. Maloney - One of the best experts on this subject based on the ideXlab platform.
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Closure of the Cytoplasmic Gate Formed by TM5 and TM11 during Transport in the Oxalate/Formate Exchanger from Oxalobacter Formigenes
2015Co-Authors: Osigbemhe Iyalomhe, Dawn Z. Herrick, David S. Cafiso, Peter C. MaloneyAbstract:OxlT, the oxalate/formate exchanger of Oxalobacter Formigenes, is a member of the major facilitator superfamily of transporters. In the present work, substrate (oxalate) was found to enhance the reactivity of the cysteine mutant S336C on the cytoplasmic end of helix 11 to methanethiosulfonate ethyl carboxylate. In addition, S336C is found to spontaneously cross-link to S143C in TM5 in either native or reconstituted membranes under conditions that support transport. Continuous wave EPR measurements are consistent with this result and indicate that positions 143 and 336 are in close proximity in the presence of substrate. These two residues are localized within helix interacting GxxxG-like motifs (G140LASG144 and S336DIFG340) at the cytoplasmic poles of TM5 and TM11. Pulse EPR measurements were used to determine distances and distance distributions across the cytoplasmic or periplasmic ends of OxlT and were compared with the predictions of an inside-open homology model. The data indicate that a significant population of transporter is in an outside-open configuration in the presence of substrate; however, each end of the transporter exhibits significant conformational heterogeneity, where both inside-open and outside-open configurations are present. These data indicate that TM5 and TM11, which form part of the transport pathway, transiently close during transport and that there is a conformational equilibrium between inside-open and outside-open states of OxlT in the presence of substrate
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closure of the cytoplasmic gate formed by tm5 and tm11 during transport in the oxalate formate exchanger from Oxalobacter Formigenes
Biochemistry, 2014Co-Authors: Osigbemhe Iyalomhe, Dawn Z. Herrick, David S. Cafiso, Peter C. MaloneyAbstract:OxlT, the oxalate/formate exchanger of Oxalobacter Formigenes, is a member of the major facilitator superfamily of transporters. In the present work, substrate (oxalate) was found to enhance the reactivity of the cysteine mutant S336C on the cytoplasmic end of helix 11 to methanethiosulfonate ethyl carboxylate. In addition, S336C is found to spontaneously cross-link to S143C in TM5 in either native or reconstituted membranes under conditions that support transport. Continuous wave EPR measurements are consistent with this result and indicate that positions 143 and 336 are in close proximity in the presence of substrate. These two residues are localized within helix interacting GxxxG-like motifs (G140LASG144 and S336DIFG340) at the cytoplasmic poles of TM5 and TM11. Pulse EPR measurements were used to determine distances and distance distributions across the cytoplasmic or periplasmic ends of OxlT and were compared with the predictions of an inside-open homology model. The data indicate that a significant ...
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Closure of the cytoplasmic gate formed by TM5 and TM11 during transport in the oxalate/formate exchanger from Oxalobacter Formigenes.
Biochemistry, 2014Co-Authors: Osigbemhe Iyalomhe, Dawn Z. Herrick, David S. Cafiso, Peter C. MaloneyAbstract:OxlT, the oxalate/formate exchanger of Oxalobacter Formigenes, is a member of the major facilitator superfamily of transporters. In the present work, substrate (oxalate) was found to enhance the reactivity of the cysteine mutant S336C on the cytoplasmic end of helix 11 to methanethiosulfonate ethyl carboxylate. In addition, S336C is found to spontaneously cross-link to S143C in TM5 in either native or reconstituted membranes under conditions that support transport. Continuous wave EPR measurements are consistent with this result and indicate that positions 143 and 336 are in close proximity in the presence of substrate. These two residues are localized within helix interacting GxxxG-like motifs (G140LASG144 and S336DIFG340) at the cytoplasmic poles of TM5 and TM11. Pulse EPR measurements were used to determine distances and distance distributions across the cytoplasmic or periplasmic ends of OxlT and were compared with the predictions of an inside-open homology model. The data indicate that a significant ...
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Helix proximity in OxlT, the oalate:formate antiporter of Oxalobacter Formigenes. Cross-linking between TM2 and TM11.
The Journal of biological chemistry, 2001Co-Authors: Young Mog Kim, Peter C. MaloneyAbstract:Experiments were designed to evaluate the proximity of transmembrane helices two (TM2) and eleven (TM11) in the tertiary structure of OxlT, the oxalate:formate exchange transporter of Oxalobacter Formigenes. A tandem duplication of the Factor Xa protease cleavage site (IEGRIEGR) was inserted into the central cytoplasmic loop of an OxlT cysteine-less derivative in which an endogenous cleavage site had been eliminated by mutagenesis (R248Q). Using this host, double cysteine derivatives were constructed so as to pair one of seventeen positions in TM2 with one of four positions in TM11. Following treatment of membrane vesicles with Cu(II)(1,10-phenanthroline)(3), molecular iodine, or N,N'-o-phenylenedimaleimide, samples were exposed to Factor Xa, and disulfide bond formation was assessed after SDS-polyacrylamide gel electrophoresis by staining with antibody directed against the OxlT C terminus. In the absence of disulfide bond formation, exposure to Factor Xa revealed the expected C-terminal 22-kDa fragment, a result unaffected by the presence of reductant. By contrast, after disulfide formation, OxlT mobility remained at 35 kDa, and appearance of the 22-kDa fragment required addition of 200 mm dithiothreitol prior to electrophoresis. The four TM11 positions chosen for cysteine substitution lie on a helical face known to interact with substrate. Similarly, TM2 positions supporting disulfide trapping were also confined to a single helical face. We conclude that TM2 and TM11 are in close juxtaposition to one another in the tertiary structure of OxlT.
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Topology of OxlT, the Oxalate Transporter of Oxalobacter Formigenes, Determined by Site-Directed Fluorescence Labeling
Journal of bacteriology, 2001Co-Authors: Zhenzhen Jia, Thomas S. Jung, Peter C. MaloneyAbstract:The topology of OxlT, the oxalate:formate exchange protein of Oxalobacter Formigenes, was established by site-directed fluorescence labeling, a simple strategy that generates topological information in the context of the intact protein. Accessibility of cysteine to the fluorescent thiol-directed probe Oregon green maleimide (OGM) was examined for a panel of 34 single-cysteine variants, each generated in a His9-tagged cysteine-less host. The reaction with OGM was readily scored by examining the fluorescence profile after sodium dodecyl sulfate-polyacrylamide gel electrophoresis of material purified by Ni 21 -linked affinity chromatography. A position was assigned an external location if its single-cysteine derivative reacted with OGM added to intact cells; a position was designated internal if OGM labeling required cell lysis. We also showed that labeling of external, but not internal, positions was blocked by prior exposure of cells to the impermeable and nonfluorescent thiol-specific agent ethyltrimethylammonium methanethiosulfonate. Of the 34 positions examined in this way, 29 were assigned unambiguously to either an internal or external location; 5 positions could not be assigned, since the target cysteine failed to react with OGM. There was no evidence of false-positive assignment. Our findings document a simple and rapid method for establishing the topology of a membrane protein and show that OxlT has 12 transmembrane segments, confirming inferences from hydropathy analysis. The gram-negative bacterium Oxalobacter Formigenes sustains a proton motive force by utilizing a “virtual” proton pump based on the transport and metabolism of oxalate. An electric
Ammon B Peck - One of the best experts on this subject based on the ideXlab platform.
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Colonization of the neonatal rat intestinal tract from environmental exposure to the anaerobic bacterium Oxalobacter Formigenes.
Journal of Medical Microbiology, 2004Co-Authors: Janet G. Cornelius, Ammon B PeckAbstract:Oxalobacter Formigenes, an anaerobic bacterium that inhabits the mammalian gastrointestinal tract, has an important symbiotic relationship with its vertebrate hosts by regulating oxalic acid homeostasis. Epidemiological studies of O. Formigenes colonization in man have shown that colonization occurs in young children, that every child can become colonized naturally, that >20 % lose colonization during adolescence or as adults and that stable colonization can be disrupted by antibiotic use or changes in diet, greatly affecting subsequent health. As O. Formigenes is a fastidious anaerobe that seldom re-colonizes adults, the question arises as to how initial colonization occurs. To investigate this question, non-colonized female laboratory rats were placed on diets high in oxalate and were colonized by oesophageal gavage with O. Formigenes either before or after being impregnated. Faecal specimens from their offspring were tested for the presence of O. Formigenes. Although the bacterium was first detected in a few neonates as early as 7 days post-partum, colonization of all the offspring did not occur until after weaning. In each case, the offspring were colonized with the bacterial strain carried by their mothers. To determine whether O. Formigenes colonization occurs vertically or horizontally, newborn rats were placed with foster mothers that were either non-colonized or colonized with an O. Formigenes strain different from that of their natural mothers. Colonization occurred temporally in a manner similar to natural colonization but all offspring became colonized only with the O. Formigenes strain of the foster mothers. These data indicate that intestinal colonization occurs horizontally, but does not answer the question of how O. Formigenes survives the aerobic environment in order to be transmitted.
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RAPID REVERSAL OF HYPEROXALURIA IN A RAT MODEL AFTER PROBIOTIC ADMINISTRATION OF Oxalobacter Formigenes
The Journal of urology, 2001Co-Authors: Harmeet Sidhu, Milton J. Allison, J.o. May Chow, Amy Clark, Ammon B PeckAbstract:Purpose: The gut inhabiting bacterium Oxalobacter Formigenes may be a negative risk factor in recurrent calcium oxalate kidney stone disease that apparently maintains oxalic acid homeostasis in its host via the degradation of dietary oxalate. The possibility of using this bacterium as probiotic treatment to reduce urinary oxalate was investigated in a rat model.Materials and Methods: Male Sprague-Dawley rats were placed on a diet supplemented with ammonium oxalate to induce a state of severe hyperoxaluria. Subgroups of these rats received an esophageal gavage of 1 × 103, 105, 107 or 109 O. Formigenes per feeding for a 2-week period. Each rat was followed for general health and changes in urinary oxalate.Results: Rats with chronic hyperoxaluria resulting from high dietary oxalate that were treated with O. Formigenes showed decreased urinary oxalate within 2 days of initiating probiotic supplementation. The amount of the decrease in a 2-week period proved directly proportional to the dose of bacteria. Urina...
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Direct Quantification of the Enteric Bacterium Oxalobacter Formigenes in Human Fecal Samples by Quantitative Competitive-Template PCR
Journal of clinical microbiology, 1999Co-Authors: H Sidhu, Milton J. Allison, Ross P. Holmes, Ammon B PeckAbstract:Homeostasis of oxalic acid appears to be regulated, in part, by the gut-associated bacterium Oxalobacter Formigenes. The loss of this bacterium from the gut flora is associated with an increased susceptibility to hyperoxaluria, a condition which can lead to the formation of calcium oxalate crystalluria and kidney stones. In order to identify and quantify the presence of O. Formigenes in clinical specimens, a quantitative-PCR-based assay system utilizing a competitive DNA template as an internal standard was developed. This quantitative competitive-template PCR test allows for the rapid, highly specific, and reproducible quantification of O. Formigenes in fecal samples and provides a prototype for development of DNA-based quantitative assays for enteric bacteria.
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absence of Oxalobacter Formigenes in cystic fibrosis patients a risk factor for hyperoxaluria
The Lancet, 1998Co-Authors: Harmeet Sidhu, Bernd Hoppe, Klaus Tenbrock, Sabine Bromme, Ernst Rietschel, Ammon B PeckAbstract:Summary Background Patients with cystic fibrosis have an increased risk of hyperoxaluria, and of subsequent nephrocalcinosis and calcium-oxalate urolithiasis. Oxalate homoeostasis is controlled, in part, by the intestinal bacterium, Oxalobacter Formigenes. The loss of this bacterium from the gut flora is associated with an increased risk of hyperoxaluria and calcium-oxalate urolithiasis. We investigated whether the absence of O Formigenes and the presence of hyperoxaluria are correlated in cystic fibrosis (CF) patients. Methods Stool specimens from 43 patients with CF aged 3–9 years and from 21 similarly aged healthy volunteers were examined for O Formigenes by culture and DNA analysis. At the same time, 24 h urine samples were collected and analysed for oxalate and other factors that promote or inhibit stone formation. Findings 15 (71%) of 21 healthy volunteers but only seven (16%) of 43 CF patients were colonised with O Formigenes. Detection of O Formigenes in six of these seven patients required DNA-based identification, suggesting low numbers of colony-forming units, and the CF patient with normal numbers of O Formigenes was the only one of the 43 patients who had not been treated with antibiotics. All seven CF patients colonised with O Formigenes had normal urinary oxalate levels, but 19 (53%) of 36 patients not colonised with O Formigenes were hyperoxaluric, with the most severe hyperoxaluria occurring in young patients. Interpretation Absence of O Formigenes from the intestinal tract of CF patients appears to lead to increased absorption of oxalate, thereby increasing the risk of hyperoxaluria and its complications (eg, nephrocalcinosis, urolithiasis). Prolonged widespread use of antibiotics, and alterations of the gastrointestinal tract that occur in CF, may induce a permanent decolonisation in CF patients.
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DNA sequencing and expression of the formyl coenzyme A transferase gene, frc, from Oxalobacter Formigenes.
Journal of bacteriology, 1997Co-Authors: Harmeet Sidhu, A L Baetz, Hui-yu Lung, S D Ogden, Benjamin G. Luttge, Ammon B PeckAbstract:Oxalic acid, a highly toxic by-product of metabolism, is catabolized by a limited number of bacterial species utilizing an activation-decarboxylation reaction which yields formate and CO2. frc, the gene encoding formyl coenzyme A transferase, an enzyme which transfers a coenzyme A moiety to activate oxalic acid, was cloned from the bacterium Oxalobacter Formigenes. DNA sequencing revealed a single open reading frame of 1,284 bp capable of encoding a 428-amino-acid protein. A presumed promoter region and a rho-independent termination sequence suggest that this gene is part of a monocistronic operon. A PCR fragment containing the open reading frame, when overexpressed in Escherichia coli, produced a product exhibiting enzymatic activity similar to the purified native enzyme. With this, the two genes necessary for bacterial catabolism of oxalate, frc and oxc, have now been cloned, sequenced, and expressed.
Harmeet Sidhu - One of the best experts on this subject based on the ideXlab platform.
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The role of Oxalobacter Formigenes colonization in calcium oxalate stone disease
Kidney international, 2013Co-Authors: Roswitha Siener, G.e. Von Unruh, Harmeet Sidhu, Ursula Bangen, Ruth Hönow, Albrecht HesseAbstract:About 75% of urinary stones contain oxalate. As Oxalobacter Formigenes is a Gram-negative anaerobic bacterium that degrades oxalate in the intestinal tract, we assessed the role of O. Formigenes in oxalate metabolism by evaluating its intestinal absorption, plasma concentration, and urinary excretion. Of 37 calcium oxalate stone formers, 26 tested negative for O. Formigenes and were compared with the 11 patients who tested positive. Patients provided 24-h urine samples on both a self-selected and a standardized diet. Urinary oxalate excretion did not differ significantly on the self-selected diet, but was significantly lower in O. Formigenes -positive than in O. Formigenes -negative patients under controlled, standardized conditions. Intestinal oxalate absorption, measured using [ 13 C 2 ]oxalate, was similar in the patients with or without O. Formigenes . Plasma oxalate concentrations were significantly higher in noncolonized (5.79μmol/l) than in colonized stone formers (1.70μmol/l). Colonization with O. Formigenes was significantly inversely associated with the number of stone episodes. Our findings suggest that O. Formigenes lowers the intestinal concentration of oxalate available for absorption at constant rates, resulting in decreased urinary oxalate excretion. Thus, dietary factors have an important role in urinary oxalate excretion. The data indicate that O. Formigenes colonization may reduce the risk of stone recurrence.
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Detection and characterization of merohedral twinning in crystals of oxalyl-coenzyme A decarboxylase from Oxalobacter Formigenes.
Biochimica et biophysica acta, 2005Co-Authors: Catrine L. Berthold, Stefano Ricagno, Nigel G. J. Richards, Harmeet Sidhu, Ylva LindqvistAbstract:Abstract Oxalyl-coenzyme A decarboxylase is a thiamin diphosphate dependent enzyme active in the catabolism of the highly toxic compound oxalate. The enzyme from Oxalobacter Formigenes has been expressed as a recombinant protein in Escherichia coli , purified to homogeneity and crystallized. Two crystal forms were obtained, one showing poor diffraction and the other merohedral twinning. Crystals in the former category belong to the tetragonal space group P4 2 2 1 2. Data to 4.1 A resolution were collected from these crystals and an incomplete low resolution structure was initially determined by molecular replacement. Crystals in the latter category were obtained by co-crystallizing the protein with coenzyme A, thiamin diphosphate and Mg 2+ -ions. Data to 1.73 A were collected from one of these crystals with apparent point group 622. The crystal was found to be heavily twinned, and a twin ratio of 0.43 was estimated consistently by different established methods. The true space group P3 1 21 was deduced, and a molecular replacement solution was obtained using the low resolution structure as template when searching in detwinned data.
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oxalate degrading enzymes from Oxalobacter Formigenes a novel device coating to reduce urinary tract biomaterial related encrustation
Journal of Endourology, 2003Co-Authors: James D Watterson, Harmeet Sidhu, Peter A Cadieux, Darren T Beiko, Anthony Cook, Jeremy P Burton, Robert R Harbottle, Clarence C Lee, Elaine Rowe, Gregor ReidAbstract:Background and Purpose: The long-term placement of biomaterials within the urinary tract is limited by the development of encrustation. In a noninfected urinary environment, encrustation often results from the deposition of calcium oxalate on the biomaterial surface. There is an association between the absence of Oxalobacter Formigenes, a commensal colonic bacterium capable of degrading oxalate, and calcium oxalate stone formation. This pilot study was designed to evaluate several oxalate-degrading enzymes produced by O. Formigenes as a potential biomaterial coating to reduce urinary tract encrustation. Materials and Methods: Circular silicone disks of 6-mm diameter were incubated for 48 hours in oxalylcoenzyme A decarboxylase (OXC), formyl-coenzyme A transferase (FRC), and coenzyme A, while control disks were incubated in distilled water. The adsorption of OXC and FRC was assessed using enhanced chemiluminescence (ECL) and atomic force microscopy (AFM). Coated and uncoated disks (20 of each) were implant...
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Intestinal Oxalobacter Formigenes colonization in calcium oxalate stone formers and its relation to urinary oxalate.
Journal of endourology, 2003Co-Authors: Scott A. Troxel, Harmeet Sidhu, Poonam Kaul, Roger K. LowAbstract:Background and Purpose: Oxalobacter Formigenes is an anaerobic commensal colonic bacterium capable of degrading oxalate through the enzyme oxalyl-CoA decarboxylase. It has been theorized that individuals who lack this bacterium have higher intestinal oxalate absorption, leading to a higher urinary oxalate concentration and an increased risk of calcium oxalate urolithiasis. We performed a prospective, controlled study to evaluate O. Formigenes colonization in calcium oxalate stone formers and to correlate colonization with urinary oxalate and other standard urinary stone risk factors. Patients and Methods: Thirty-five first-time calcium oxalate stone formers were compared with 10 control subjects having no history of urolithiasis and a normal renal ultrasound scan. All subjects underwent standard metabolic testing by submitting serum and 24-hour urine specimens. In addition, all subjects submitted stool samples for culture and detection of O. Formigenes by Xentrix O. Formigenes Monitor. Results: Intestinal...
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RAPID REVERSAL OF HYPEROXALURIA IN A RAT MODEL AFTER PROBIOTIC ADMINISTRATION OF Oxalobacter Formigenes
The Journal of urology, 2001Co-Authors: Harmeet Sidhu, Milton J. Allison, J.o. May Chow, Amy Clark, Ammon B PeckAbstract:Purpose: The gut inhabiting bacterium Oxalobacter Formigenes may be a negative risk factor in recurrent calcium oxalate kidney stone disease that apparently maintains oxalic acid homeostasis in its host via the degradation of dietary oxalate. The possibility of using this bacterium as probiotic treatment to reduce urinary oxalate was investigated in a rat model.Materials and Methods: Male Sprague-Dawley rats were placed on a diet supplemented with ammonium oxalate to induce a state of severe hyperoxaluria. Subgroups of these rats received an esophageal gavage of 1 × 103, 105, 107 or 109 O. Formigenes per feeding for a 2-week period. Each rat was followed for general health and changes in urinary oxalate.Results: Rats with chronic hyperoxaluria resulting from high dietary oxalate that were treated with O. Formigenes showed decreased urinary oxalate within 2 days of initiating probiotic supplementation. The amount of the decrease in a 2-week period proved directly proportional to the dose of bacteria. Urina...
John Knight - One of the best experts on this subject based on the ideXlab platform.
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response of germfree mice to colonization by Oxalobacter Formigenes and altered schaedler flora
Applied and Environmental Microbiology, 2016Co-Authors: Xingsheng Li, Melissa L Ellis, Alexander E Dowell, Trenton R Schoeb, Casey D Morrow, Ranjit Kumar, John KnightAbstract:ABSTRACT Colonization with Oxalobacter Formigenes may reduce the risk of calcium oxalate kidney stone disease. To improve our limited understanding of host-O. Formigenes and microbe-O. Formigenes interactions, germfree mice and mice with altered Schaedler flora (ASF) were colonized with O. Formigenes. Germfree mice were stably colonized with O. Formigenes, which suggests that O. Formigenes does not require other organisms to sustain its survival. Examination of intestinal material indicated no viable O. Formigenes in the small intestine and ∼4 × 106 CFU O. Formigenes per 100 mg contents in the cecum and proximal colon, with ∼0.02% of total cecal O. Formigenes cells being tightly associated with the mucosa. O. Formigenes did not alter the overall microbial composition of ASF, and ASF did not affect the capacity of O. Formigenes to degrade dietary oxalate in the cecum. Twenty-four-hour collections of urine and feces in metabolic cages in semirigid isolators demonstrated that the introduction of ASF into germfree mice significantly reduced urinary oxalate excretion. These experiments also showed that O. Formigenes-monocolonized mice excreted significantly more urinary calcium than did germfree mice, which may be due to degradation of calcium oxalate crystals by O. Formigenes and subsequent intestinal absorption of free calcium. In conclusion, the successful establishment of mouse models with defined flora and O. Formigenes should improve our understanding of O. Formigenes-host and O. Formigenes-microbe interactions. These data support the use of O. Formigenes as a probiotic that has limited impact on the composition of the resident microbiota but provides an efficient oxalate-degrading function. IMPORTANCE Despite evidence suggesting that a lack of Oxalobacter Formigenes colonization is a risk factor for calcium oxalate stone formation, little is known about O. Formigenes biology. This study is the first to utilize germfree mice to examine the response to monocolonization with O. Formigenes, as well as the impact of a defined bacterial cocktail (i.e., ASF) on O. Formigenes colonization. This study demonstrated that germfree mice receiving their regular diet remained monocolonized with O. Formigenes, and it suggests that further studies with O. Formigenes gnotobiotic mouse models could improve our understanding of O. Formigenes biology and host-O. Formigenes and microbe-O. Formigenes interactions.
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Probiotic properties of Oxalobacter Formigenes: an in vitro examination.
Archives of microbiology, 2016Co-Authors: Melissa L Ellis, Alexander E Dowell, John KnightAbstract:Oxalobacter Formigenes (O. Formigenes) is a nonpathogenic, Gram-negative, obligate anaerobic bacterium that commonly inhabits the human gut and degrades oxalate as its major energy and carbon source. Results from a case-controlled study suggested that lack of O. Formigenes colonization is a risk factor for recurrent calcium oxalate stone formation. Hence, O. Formigenes colonization may prove to be an efficacious method for limiting calcium oxalate stone risk. However, challenges exist in the preparation of O. Formigenes as a successful probiotic due to it being an anaerobe with fastidious growth requirements. Here we examine in vitro properties expected of a successful probiotic strain. The data show that the Group 1 O. Formigenes strain OxCC13 is sensitive to pH < 5.0, persists in the absence of oxalate, is aerotolerant, and survives for long periods when freeze-dried or mixed with yogurt. These findings highlight the resilience of this O. Formigenes strain to some processes and conditions associated with the manufacture, storage and distribution of probiotic strains.
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Proteome Dynamics of the Specialist Oxalate Degrader Oxalobacter Formigenes.
Journal of proteomics & bioinformatics, 2016Co-Authors: Melissa L Ellis, Ross P. Holmes, James A. Mobley, John KnightAbstract:Oxalobacter Formigenes is a unique intestinal organism that relies on oxalate degradation to meet most of its energy and carbon needs. A lack of colonization is a risk factor for calcium oxalate kidney stone disease. The release of the genome sequence of O. Formigenes has provided an opportunity to increase our understanding of the biology of O. Formigenes. This study used mass spectrometry based shotgun proteomics to examine changes in protein levels associated with the transition of growth from log to stationary phase. Of the 1867 unique protein coding genes in the genome of O. Formigenes strain OxCC13, 1822 proteins were detected, which is at the lower end of the range of 1500-7500 proteins found in free-living bacteria. From the protein datasets presented here it is clear that O. Formigenes contains a repertoire of metabolic pathways expected of an intestinal microbe that permit it to survive and adapt to new environments. Although further experimental testing is needed to confirm the physiological and regulatory processes that mediate adaptation with nutrient shifts, the O. Formigenes protein datasets presented here can be used as a reference for studying proteome dynamics under different conditions and have significant potential for hypothesis development.
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Role of Oxalobacter Formigenes Colonization in Calcium Oxalate Kidney Stone Disease
The Role of Bacteria in Urology, 2016Co-Authors: John Knight, Ross P. HolmesAbstract:Oxalobacter Formigenes is part of the bacterial flora in the large intestine of humans and many other mammalian species. It is unique in that it requires oxalate both as an energy and carbon source. A lack of colonization with O. Formigenes is a risk factor for idiopathic recurrent calcium oxalate stone disease. Protection against calcium oxalate stone disease appears to be due to the oxalate degradation that occurs in the gut as measurements of 24 h urinary oxalate indicate that O. Formigenes colonized calcium oxalate stone formers excrete less oxalate compared to non-colonized individuals when ingesting standardized diets. There is also some evidence that suggests a possible mechanism involving intestinal oxalate secretion triggered by the bacterium itself, as O. Formigenes colonization appears to lower plasma oxalate. Whether high oral doses of this organism can promote sufficient intestinal oxalate secretion to diminish the oxalate burden on the kidney in individuals with Primary Hyperoxaluria is currently being tested by OxThera, Inc. in a phase 2 clinical trial. Much still remains to be learned about how O. Formigenes establishes and maintains gut colonization and the precise mechanisms by which it modifies stone risk.
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Oxalobacter Formigenes Colonization and Oxalate Dynamics in a Mouse Model.
Applied and environmental microbiology, 2015Co-Authors: Melissa L Ellis, John KnightAbstract:Animal and human studies have provided compelling evidence that colonization of the intestine with Oxalobacter Formigenes reduces urinary oxalate excretion and lowers the risk of forming calcium oxalate kidney stones. The mechanism providing protection appears to be related to the unique ability of O. Formigenes to rely on oxalate as a major source of carbon and energy for growth. However, much is not known about the factors that influence colonization and host-bacterium interactions. We have colonized mice with O. Formigenes OxCC13 and systematically investigated the impacts of diets with different levels of calcium and oxalate on O. Formigenes intestinal densities and urinary and intestinal oxalate levels. Measurement of intestinal oxalate levels in mice colonized or not colonized with O. Formigenes demonstrated the highly efficient degradation of soluble oxalate by O. Formigenes relative to other microbiota. The ratio of calcium to oxalate in diets was important in determining colonization densities and conditions where urinary oxalate and fecal oxalate excretion were modified, and the results were consistent with those from studies we have performed with colonized and noncolonized humans. The use of low-oxalate purified diets showed that 80% of animals retained O. Formigenes colonization after a 1-week dietary oxalate deprivation. Animals not colonized with O. Formigenes excreted two times more oxalate in feces than they had ingested. This nondietary source of oxalate may play an important role in the survival of O. Formigenes during periods of dietary oxalate deprivation. These studies suggest that the mouse will be a useful model to further characterize interactions between O. Formigenes and the host and factors that impact colonization.
Ross P. Holmes - One of the best experts on this subject based on the ideXlab platform.
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Proteome Dynamics of the Specialist Oxalate Degrader Oxalobacter Formigenes.
Journal of proteomics & bioinformatics, 2016Co-Authors: Melissa L Ellis, Ross P. Holmes, James A. Mobley, John KnightAbstract:Oxalobacter Formigenes is a unique intestinal organism that relies on oxalate degradation to meet most of its energy and carbon needs. A lack of colonization is a risk factor for calcium oxalate kidney stone disease. The release of the genome sequence of O. Formigenes has provided an opportunity to increase our understanding of the biology of O. Formigenes. This study used mass spectrometry based shotgun proteomics to examine changes in protein levels associated with the transition of growth from log to stationary phase. Of the 1867 unique protein coding genes in the genome of O. Formigenes strain OxCC13, 1822 proteins were detected, which is at the lower end of the range of 1500-7500 proteins found in free-living bacteria. From the protein datasets presented here it is clear that O. Formigenes contains a repertoire of metabolic pathways expected of an intestinal microbe that permit it to survive and adapt to new environments. Although further experimental testing is needed to confirm the physiological and regulatory processes that mediate adaptation with nutrient shifts, the O. Formigenes protein datasets presented here can be used as a reference for studying proteome dynamics under different conditions and have significant potential for hypothesis development.
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Role of Oxalobacter Formigenes Colonization in Calcium Oxalate Kidney Stone Disease
The Role of Bacteria in Urology, 2016Co-Authors: John Knight, Ross P. HolmesAbstract:Oxalobacter Formigenes is part of the bacterial flora in the large intestine of humans and many other mammalian species. It is unique in that it requires oxalate both as an energy and carbon source. A lack of colonization with O. Formigenes is a risk factor for idiopathic recurrent calcium oxalate stone disease. Protection against calcium oxalate stone disease appears to be due to the oxalate degradation that occurs in the gut as measurements of 24 h urinary oxalate indicate that O. Formigenes colonized calcium oxalate stone formers excrete less oxalate compared to non-colonized individuals when ingesting standardized diets. There is also some evidence that suggests a possible mechanism involving intestinal oxalate secretion triggered by the bacterium itself, as O. Formigenes colonization appears to lower plasma oxalate. Whether high oral doses of this organism can promote sufficient intestinal oxalate secretion to diminish the oxalate burden on the kidney in individuals with Primary Hyperoxaluria is currently being tested by OxThera, Inc. in a phase 2 clinical trial. Much still remains to be learned about how O. Formigenes establishes and maintains gut colonization and the precise mechanisms by which it modifies stone risk.
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The genetic composition of Oxalobacter Formigenes and its relationship to colonization and calcium oxalate stone disease
Urolithiasis, 2013Co-Authors: John Knight, Dean G. Assimos, Rajendar Deora, Ross P. HolmesAbstract:Oxalobacter Formigenes is a unique intestinal organism that relies on oxalate degradation to meet most of its energy and carbon needs. A lack of colonization is a risk factor for calcium oxalate stone disease. Protection against calcium oxalate stone disease appears to be due to the oxalate degradation that occurs in the gut on low calcium diets with a possible further contribution from intestinal oxalate secretion. Much remains to be learned about how the organism establishes and maintains gut colonization and the precise mechanisms by which it modifies stone risk. The sequencing and annotation of the genomes of a Group 1 and a Group 2 strain of O. Formigenes should provide the informatic tools required for the identification of the genes and pathways associated with colonization and survival. In this review we have identified genes that may be involved and where appropriate suggested how they may be important in calcium oxalate stone disease. Elaborating the functional roles of these genes should accelerate our understanding of the organism and clarify its role in preventing stone formation.
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Sensitivity of human strains of Oxalobacter Formigenes to commonly prescribed antibiotics.
Urology, 2012Co-Authors: Jessica N. Lange, Kyle Wood, John Knight, Ross P. Holmes, Hayes Wong, Richard Otto, Patrick W. Mufarrij, Haluk Akpinar, Dean G. AssimosAbstract:Objective To determine the sensitivity of 4 strains of Oxalobacter Formigenes (Oxf) found in humans—HC1, Va3, CC13, and OxK—to varying concentrations of commonly prescribed antibiotics. Oxf gut colonization has been associated with a decreased risk of forming recurrent calcium oxalate kidney stones. Methods For each strain and each antibiotic concentration, 100 μL of an overnight culture and 100 μL of the appropriate antibiotic were added to a 7-mL vial of oxalate culture medium containing 20 mM oxalate. On the fourth day, vials were visually examined for growth, and a calcium oxalate precipitation test was performed to determine whether Oxf grew in the presence of the antibiotic. Results All 4 Oxf strains were resistant to amoxicillin, amoxicillin/clavulanate, ceftriaxone, cephalexin, and vancomycin, and they were all sensitive to azithromycin, ciprofloxacin, clarithromycin, clindamycin, doxycycline, gentamicin, levofloxacin, metronidazole, and tetracycline. One strain, CC13, was resistant to nitrofurantoin, and the others were sensitive. Differences in minimum inhibitory concentration between strains were demonstrated. Conclusion Four human strains of Oxf are sensitive to a number of antibiotics commonly used in clinical practice; however, minimum inhibitory concentrations differ between strains.
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Oral antibiotic treatment of Helicobacter pylori leads to persistently reduced intestinal colonization rates with Oxalobacter Formigenes.
Journal of endourology, 2011Co-Authors: Viktoria Kharlamb, Juquan Jiang, Ross P. Holmes, Jennifer Schelker, Fritz Francois, David S. GoldfarbAbstract:Abstract Background and Purpose: Oxalobacter Formigenes (OF) may play a protective role in preventing calcium oxalate stones. This is the first prospective study to evaluate the effect of antibioti...
