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PubMed1998

Expression of genes from Rahnella aquatilis that are necessary for mineral phosphate solubilization in Escherichia coli.

Rahnella aquatilis is a Gram-negative bacterium that can fix atmospheric nitrogen and also has the ability to solubilize mineral phosphate. We have cloned the genes that confer the mineral phosphate solubilizing (Mps) trait from this organism by mobilizing a cosmid library of R. aquatilis into Escherichia coli HB101. A 7.0-kb EcoRI fragment from a cosmid, when transferred into E. coli strains HB101 and DH5 alpha, conferred the ability to solubilize hydroxyapatite and the production of gluconic acid to E. coli. The relative amounts of soluble phosphate and gluconic acid produced by the cloned 7.0-kb EcoRI fragment in E. coli were significantly higher than those of R. aquatilis. Nucleotide sequence analysis revealed two complete open reading frames (ORF1 and ORF2) and a partial ORF, ORF1 and ORF2 encoded proteins of molecular mass 10 kDa and 44 kDa. The 44-kDa protein showed extensive sequence similarity to pqqE of Erwinia herbicola, Klebsiella pneumoniae and A. Acinetobacter calcoaceticus. The 10-kDa protein revealed strong similarity to the pqqD of K. pneumoniae and A. calcoaceticus.

Chemotactic Factors↗
PubMed2004

Treatment of the common cold with troxerutin.

The rutosides are naturally occurring flavonoids that have documented effects on capillary permeability and edema. The purpose of this study was to assess the effect of troxerutin, one of the rutosides, on the symptoms of the common cold. Ninety-four volunteers with common cold symptoms were recruited for participation in the study. Volunteers were randomized to either active treatment (n=49) with troxerutin (50 mg) and Zn gluconate (25 mg) or control treatment (n=45) with 10 mg Zn gluconate. Symptoms were assessed by subjective symptom score prior to treatment and then daily for the next 4 days. The total symptom score over the 4 days of study treatment was 27.7+/-2.0 (mean+/-SEM) and 33.0+/-2.6 in the active and control groups, respectively (p=0.10, unpaired t-test). The total daily symptom score on day 1 was reduced by 11% compared to baseline in the active group and by 1% in the control group (p=0.03). Evaluation of the effect of treatment on individual symptoms revealed a significant effect on rhinorrhea. The total rhinorrhea score over the course of the study was 3.7+/-0.4 in the active group compared to 5.1+/-0.5 in the control group (p=0.025, unpaired t-test). Daily rhinorrhea scores were significantly lower in the active group on study days 1 and 3. Based on this preliminary study, the possibility that the rutosides might provide a safe and effective treatment for rhinorrhea in the common cold deserves systematic evaluation.

Administration, Oral↗
PubMed1980

The influence of various counter ions on the interaction of chlorhexidine with the hydrophilic contact lens polymer, poly(2-hydroxyethyl methacrylate).

The commercially available salts of chlorhexidine were found to interact with the hydrogel poly (2-hydroxyethyl methacrylate) to different extents, the affinity for the polymer decreasing in the order acetate, gluconate, chloride. The uptakes of the acetate and gluconate salts were almost entirely irreversible over the concentration range studied. The influence of some alternative counter ions, namely amino acids and dicarboxylic acid salts, was examined and found to generate a three-fold variation in the extent of the chlorhexidine interaction. Uptakes were greatest for counter ions that were either hydrophobic in character or bore a net negative charge. Only two compounds, glycine and monosodium oxalate, were successful in reducing the extent of sorption below that observed for chlorhexidine hydrochloride.

Acetates↗
PubMed1974

The sodium-transporting compartment of the epithelium of frog skin.

1. The abdominal frog skin was mounted between two chambers containing Ringer with 1 mM-Na on the outside and 115 mM-Na on the inside. When the Na concentration of the outer solution ([Na](o)) is instantaneously raised from 1 to 50 mM, the short circuit current (I) increases to a new value in less than a second, and becomes essentially time-independent. Only in a few experiments was it observed to increase further, although at a much slower rate.2. At a time t after this increase, the addition of 10(-4)M amiloride to the outer solution produces an exponential decrease of I. The area under this exponential curve is generally taken to reflect the existence of a Na- transporting compartment (NaTC).3. The amount of Na represented by NaTC is a function of t: it increases from 1.7 x 10(-9) mole. cm(-2), at t = 10 sec, to 22.8 x 10(-9) mole. cm(-2) at t = 10 min.4. In view of the fact that (a) I is not a function of the size of the ;NaTC' and (b) that whereas I reaches a steady value in a fraction of a second the size of NaTC keeps increasing for minutes, it is proposed that the ;NaTC' represents an amount of Na which is not located along the main route of transepithelial transport.5. On the assumption that the NaTC is located in a cellular compartment and that, in order to accumulate in this compartment Na should be accompanied by a permeable anion, a series of experiments were performed with Ringer in which Cl(-) was replaced by gluconate. It was observed as expected, that NaTC in gluconate is 164 times smaller than in Cl(-), but I only decreases to one half its value in Cl(-) Ringer.

Amiloride↗
PubMed1985

Apical membrane potassium and chloride permeabilities in surface cells of rabbit descending colon epithelium.

The apical membranes of surface cells in the rabbit descending colon possess a significant ionic conductance in parallel to amiloride-blockable Na+ channels. The identity of the ion(s) responsible for the amiloride-insensitive conductance is unknown. The purpose of the present paper was to assess the permeability and net driving forces for K+ and Cl- across this membrane using conventional and ion-sensitive micro-electrode techniques. Intracellular Cl- activity (aiCl) averaged 23 +/- 2 mM with an equilibrium potential (ECl) of -38 +/- 2 mV. This value is less than previous estimates of the electromotive force (e.m.f.) of the amiloride-insensitive pathway (ca. -50 mV). Consequently, Cl- alone cannot account for the amiloride-insensitive conductance. Replacement of Cl- by gluconate in the serosal solution decreased aiCl to 17 +/- 2.8 mM. aiCl was lowered to approximately 1 mM by replacement in the mucosal bath or by replacement in both solutions. The results indicate a low Cl- conductance in the basolateral membrane, in agreement with previous electrophysiological studies of this epithelium. In contrast to Cl-, the chemical driving force for K+ was large enough to support the e.m.f. of the amiloride-insensitive pathway (K+ equilibrium potential, EK = -66 mV). The basolateral membrane potential (Vbl), EK and the intracellular K+ activity (aiK) were decreased in parallel following inhibition of the basolateral Na-K pump, providing evidence that Vbl is largely due to a K+ diffusion potential. In the presence of serosal 10(-4) M-ouabain, aiK appeared to remain above equilibrium and more than doubled after addition of Ba2+ to the serosal bath. Replacement of the mucosal bathing solution with KCl or gluconate Ringer solution largely restored Vbl and the transepithelial potential (VT) in tissues which had been previously treated with ouabain. The restoration of VT was decreased and the transepithelial resistance (RT) was increased by addition of tetraethylammonium to the mucosal bath. The above results suggest that there are at least four routes for ion movement across the apical membrane of rabbit colon surface epithelial cells. These are: (1) an amiloride-sensitive Na+ channel, (2) a K+ conductance, (3) electroneutral uptake of Cl- from lumen to cell interior and (4) an active K+ transport mechanism, also from lumen to cell interior.

Animals↗
PubMed1988

Electrical transients produced by the toad urinary bladder in response to altered medium osmolality.

1. The effects of changes in media osmolality on the transepithelial current through the toad urinary bladder under voltage-clamp conditions have been studied. Over the limited range (+/- 24 mosmol/kg H2O) used in these investigations, changes in the osmolality of the mucosal bathing fluid produced no changes in transepithelial current. 2. Changes in osmolality of the serosal fluid greatly affected the transepithelial current with a decrease (increase) in osmolality producing a sustained increase (decrease) in current. 3. The changes in steady-state current were approximately proportional to the magnitude of the osmotic steps and were reproducible and reversible if the osmolalities of the solutions were confined to a domain of 220-260 mosmol/kg H2O. 4. Amiloride, which was used to block all active current, also eliminated the electrical responses to an osmotic pulse, indicating that the responses were of cellular origin. 5. The effects of substituting gluconate for medium chloride were examined. Similar responses were observed, indicating that they were not due to changes in a plasma membrane chloride conductance. 6. The transient currents observed during the changes from one steady state to the other often contained an oscillatory component, the amplitude and the degree of damping of which varied between bladders. The amplitude of the oscillations, but not their frequency, could be varied by altering the magnitude of the osmotic pulse and by changing the imposed transepithelial voltage. Decreasing the electrical potential of the mucosal solution with respect to that of the serosal solution decreased the amplitude of the oscillations, as did increased serosal potassium or substitution of gluconate for serosal chloride. The period of the oscillations always remained within the range of 9-12 min. 7. The results suggest that two major processes are initiated by an osmotic step in the serosal bathing medium. The first involves the establishment of new ion gradients and the second, alterations in sodium pump activity. In addition, there is evidence for a voltage-dependent sodium conductance in the apical membrane.

Action Potentials↗
PubMed1989

The effect of anions on bound acetylcholine in frog sartorius muscle.

1. Frog sartorius muscles were treated with an irreversible cholinesterase inhibitor and then incubated in isotonic potassium propionate solution (isotonic KPr). Total and bound, presumably vesicular, acetylcholine (ACh) in the tissue and ACh in the medium were assayed by mass fragmentography, miniature end-plate potentials (MEPPs) were recorded and the end-plates were investigated by electron microscopy. 2. Incubation in isotonic KPr for 30 min stimulated ACh release and concomitantly decreased total and bound ACh. Nerve stimulation for 30 min by trains of impulses (0.1 s trains of 100 Hz, 1 train s-1) in normal-potassium propionate-containing solution had the same effects. 3. When the tissue was incubated in normal-K+ Ringer solution for 3 h, following chemical or electric stimulation, bound ACh recovered to about 75% of the initial value, provided that Cl- ions were present in the medium. In the presence of propionate instead of Cl- ions almost no recovery of bound ACh took place. There was also recovery of bound ACh in the presence of either NO3- or gluconate ions. In NO3- it was the same as in Cl-, but in gluconate it was less than found in Cl- -containing medium. 4. Recovery of total ACh, in contrast to bound ACh, took place even in the presence of propionate ions, showing that extracellular Cl- is not required for the synthesis of ACh. 5. In terminals recovered in normal Ringer solution, many synaptic vesicles were found, but terminals 'recovered' in propionate solution were depleted of vesicles. 6. From these and other results it is concluded that the recycling of synaptic vesicles normally requires the presence of extracellular chloride.

Acetylcholine↗
PubMed1990

Intracellular pH regulation in the embryonic chicken lens epithelium.

1. The intracellular pH (pHi) of embryonic lens epithelia was measured by the emission ratio technique using the fluorescent pH probe carboxy-seminaphthorhodafluor-1 (Snarf-1). 2. In artificial aqueous humour solutions (AAH) containing HCO3-, pHi was 7.45, a value more alkaline than that of the bathing medium (pH = 7.3). In HCO3- -free AAH, pHi was 7.29. 3. Acetazolamide, an inhibitor of carbonic anhydrase, had no effect on resting pHi. 4. The pHi could be manipulated experimentally by changing the external pH (pHo) of HEPES-buffered AAH, the addition or withdrawal of CO2-HCO3-, or by perfusion with the weak bases NH4Cl and procaine. 5. The pHi change induced by withdrawal of 5 mM-procaine was used to calculate a value for the intrinsic cytoplasmic buffering capacity (beta i) of 16.5 mM. 6. The addition of amiloride (1 mM) or treatment with low-Na+ AAH solutions led to a decrease in pHi of 0.23 over the 10 min exposure. In addition, these treatments inhibited pHi recovery from NH4(+)-induced acidosis. These observations are consistent with the presence of amiloride-sensitive N(+)-H+ antiport. 7. Addition of exogenous antiport activity in the form of 50 microM-monensin caused an increase in pHi of 0.24. 8. In HCO3(-)-containing media, replacing Cl- by gluconate or isothionate led to an immediate, reversible increase in pHi which could be completely inhibited by 2 mM-4-acetamido-4'-isothiocyanato-stillbene-2,2'-disulphonic acid (SITS). This indicates the presence of Cl(-)-HCO3- exchange in this tissue. 9. Under HCO3(-)-free conditions, replacement of Cl- by gluconate or isothionate caused a small transient acidification followed, 5 min later, by a large sustained alkalinization. The delayed increase in pHi could be completely blocked by 1 mM-amiloride and may reflect volume-sensitive stimulation of the Na(+)-H+ antiporter as cell volume (estimated by cell height measurements) was shown to decrease significantly during this period.

4-Acetamido-4'-isothiocyanatostilbene-2,2'-disulfo↗
PubMed1994

Kinetic separation of charge movement components in intact frog skeletal muscle.

1. Procedures for a complete charge movement separation employed a combination of its steady-state inactivation and activation properties in intact frog skeletal muscle fibres in gluconate-containing solutions. 2. Holding potential shifts from -70 to -50 mV reduced the total charge available between -90 and -20 mV from 16.76 +/- 1.70 nC microF-1 (mean +/- S.E.M.; n = 4 fibres) to 9.25 +/- 1.43 nC microF-1 without significant loss of tetracaine-resistant charge (q beta). 3. The steady-state and kinetic properties of tetracaine-sensitive charge (q gamma) persisted through holding potential changes from -90 to -70 mV in the presence of gluconate and generally resembled activation properties established hitherto in sulphate-containing solutions. 4. Further holding potential displacement to -50 mV abolished q gamma charge movements and depressed the charge-voltage curve. 5. Test voltage steps applied from a -70 mV prepulse level gave rapid monotonic q beta decays and similarly depressed activation functions in 2 mM tetracaine unchanged by holding potential shifts between -70 and -50 mV. 6. The isolated 'on' q gamma charge movements, I(t), always included early transients that preceded any prolonged charging phases and which increased with depolarization. They decayed to stable baselines in the absence of prolonged time-dependent or inward-current phases and yielded integrals, Q(t), that monotonically increased with test voltage. 7. 'Off' steps always elicited rapid monotonic q gamma decays that fully returned the 'on' charge. 8. 'On' and 'off' q gamma currents, I(t), following voltage steps from fixed conditioning to varying test levels mapped onto topologically distinct higher-order phase-plane trajectories, I(Q), that steeply varied with test voltage. 9. In contrast, voltage steps to fixed test potentials of either -70 or -20 mV elicited identical q gamma phase-plane trajectories independent of prepulse history. 10. The q gamma current thus reflects an independent, capacitative process driven uniquely by higher-order dependences upon charge distribution, Q(t), and test voltage, V(t), autonomous of prepulse history or time, t.

Animals↗
PubMed2002

Vesicle size and transmitter release at the frog neuromuscular junction when quantal acetylcholine content is increased or decreased.

We investigated whether the synaptic vesicles at the neuromuscular junction change size when their acetylcholine (ACh) content is altered. The size of the miniature endplate potential (MEPP) increased 3- or 4-fold in preparations pre-treated in a hypertonic solution in which the anion was gluconate. We measured the dimensions of synaptic vesicles in such preparations and in controls. The size of the vesicles and size distribution were indistinguishable. Quanta contained about half of the usual amount of ACh in preparations stimulated in the presence of hemicholinium-3, an inhibitor of choline uptake, or in NH(4)(+), which diminishes the proton gradient for ACh uptake into the vesicles. Neither treatment changed the size of the synaptic vesicles. ACh content and vesicle size were both decreased in preparations stimulated in (-)-vesamicol, an inhibitor of ACh uptake in vesicles. Since the other inhibitors decreased ACh content by a similar amount without altering vesicle size, (-)-vesamicol may decrease vesicle size by acting on another target. We also found that a hypertonic solution in which the anion was aspartate increased quantal size similar to gluconate. Both anions have high hydration energy and a large volume. When these treatments increased quantal size the mean 20-80 % rise time of MEPPs recorded with an extracellular electrode was 170 micros. In the controls it was 97 micros. Perhaps some of the added ACh is bound within the vesicles, which slows the rise. Our major conclusion is that ACh content can change notably without any change in the size of the synaptic vesicles.

Acetylcholine↗
PubMed2002

Energy generation by extracellular aldose oxidation in N(2)-fixing Gluconacetobacter diazotrophicus.

Gluconacetobacter diazotrophicus PAL3 was grown in a chemostat with N(2) and mixtures of xylose and gluconate. Xylose was oxidized to xylonate, which was accumulated in the culture supernatants. Biomass yields and carbon from gluconate incorporated into biomass increased with the rate of xylose oxidation. By using metabolic balances it is demonstrated that extracellular xylose oxidation led N(2)-fixing G. diazotrophicus cultures to increase the efficiency of energy generation.

Acetobacteraceae↗
PubMed1967

Computer simulation of fermentation systems.

Results of batch fermentation of gluconic acid by Pseudomonas ovalis were graphically analyzed to obtain a kinetic model to represent the data. Since gluconic acid was produced by the hydrolysis of a lactone intermediate, the model was necessarily represented by a set of kinetic equations. A computer simulation technique involving the use of the MIDAS program was developed to solve the system of nonlinear equations and to check the appropriateness of the model. Since the maximal specific growth rate and the rate constant for the production of the lactone intermediate varied with time, function generators were used to simulate these system parameters. The merit of using the MIDAS program was considered in relation to analysis and model testing in microbiological processes of similar types.

Computers↗
PubMed1973

Pathways of carbohydrate metabolism in Microcyclus species.

Radiorespirometric and enzymatic studies were conducted to determine primary and secondary pathways of carbohydrate catabolism in Microcyclus aquaticus and M. flavus. M. aquaticus catabolizes both glucose and gluconate mainly via the Entner-Doudoroff and pentose phosphate pathways with some concurrent participation of the Embden-Meyerhof pathway. M. flavus, however, oxidizes glucose mainly via the Embden-Meyerhof pathway and gluconate via the Entner-Doudoroff pathway with some simultaneous operation of the pentose phosphate pathway. Both of the organisms showed evidence of the tricarboxylic acid cycle as a secondary pathway for the oxidation of carbohydrates.

Acetates↗
PubMed1973

Abnormal septation and inhibition of sporulation by accumulation of L- -glycerophosphate in Bacillus subtilis mutants.

Accumulation of l-alpha-glycerophosphate, in cells of Bacillus subtilis mutants lacking the nicotinamide adenine dinucleotide-independent glycerophosphate dehydrogenase activity, suppresses both growth and sporulation. After growth has stopped, the cells slowly develop one and later more asymmetric septa that are thicker than normal prespore septa and apparently contain too much cell wall material to allow further membrane development into forespores or spores. l-Malate prevents accumulation of glycerophosphate and restores sporulation of the mutant. Glucose or gluconate cannot resotre sporulation, because they still effect glycerophosphate accumulation via de novo synthesis. If that accumulation is blocked in a double mutant, which is unable to make glycerophosphate from or to metabolize it into Embden-Meyerhof compounds, then nonsuppressing amounts of glucose or gluconate can restore sporulation.

Bacillus subtilis↗
PubMed1973

Taxonomy of marine bacteria: Beneckea parahaemolytica and Beneckea alginolytica.

A collection of 169 strains, including 91 obtained from cases of gastroenteritis and 41 from localized tissue infections and infections of the eye and ear, was submitted to an extensive nutritional, physiological, and morphological characterization. The nutritional and physiological data obtained from these strains, as well as data for strains of other species of the genus Beneckea, were submitted to a numerical analysis which grouped the strains into clusters on the basis of phenotypic similarity. Strains from cases of gastroenteritis formed a group of three clusters which linked at a similarity value of 68%. These three clusters could not, however, be separated from each other by universally positive or negative traits, and on the basis of their overall phenotypic similarity were assigned to a single species, B. parahaemolytica. The majority of the strains from human, nonenteric sources segregated into two distinct clusters, one designated B. alginolytica and the other unassigned with respect to species (group C-2). B. parahaemolytica, B. alginolytica, and group C-2 could be readily distinguished from one another as well as from the remaining species of the genus Beneckea by multiple, unrelated, phenotypic traits. Activities of selected enzymes of glucose and gluconate catabolism in cell-free extracts of B. parahaemolytica, B. alginolytica, and group C-2 suggested that these organisms utilized glucose primarily via the Embden-Meyerhof pathway and gluconate primarily via the Entner-Doudoroff pathway. Similar results were observed in the other members of the genus Beneckea.

Bacteria↗
PubMed1974

Sugar catabolism in Aquaspirillum gracile.

Aquaspirillum (Spirillum) gracile is one of the few spirilla that cause acidification of the medium when cultured with sugars. Acidic reactions have been reported only for d-glucose, d-galactose, and l-arabinose, and the mode of attack of these sugars has not been previously investigated. The soluble portion of extracts of glucose-cultured cells of A. gracile ATCC 19624 was found by spectrophotometric methods to contain enzyme activities characteristic of the Entner-Doudoroff and Embden-Meyerhof-Parnas pathways. No activity for 6-phosphogluconate dehydrogenase (EC 1.1.1.44) was detected. Pyridine nucleotide-linked dehydrogenase activities for l-arabinose and d-galactose (EC 1.1.1.46 and EC 1.1.1.48) occurred in the soluble fraction of cells cultured with either sugar. Glucose-cultured cells contained not only glucokinase (EC 2.7.1.2) and glucose-6-phosphate dehydrogenase (EC 1.1.1.49) activities but also glucose dehydrogenase (EC 1.1.1.47) activity. Enzymes capable of oxidizing gluconate were not detectable, but gluconokinase (EC 2.7.1.12) activity was present. Paper chromatographic analysis of the spent culture supernatant media from glucose-cultured cells indicated an accumulation of gluconic acid, and this was confirmed by enzymatic methods. Evidence is presented for the production of d-galactonic and l-arabonic acids in cultures containing d-galactose or l-arabinose, respectively.

Alcohol Oxidoreductases↗
PubMed1974

Glucose uptake and phosphorylation in Pseudomonas fluorescens.

Pseudomonas fluorescens ATCC 13525 and a particulate glucose oxidase (d-glucose:oxygen oxidoreductase, EC 1.1.3.4) mutant of this organism, gox-7, were examined to determine if glucose oxidation via particulate glucose oxidase is a required first step for glucose uptake. Initial [(14)C]glucose-uptake rates in parent and gox-7 cells were qualitatively similar. Initial [(14)C]glucose-uptake product analysis revealed that glucose was accumulated via active transport and was rapidly metabolized to glucose-6-phosphate and gluconate-6-phosphate in both parent and gox-7 cells. Cell extracts contained soluble adenosine 5'-triphosphate specific kinase activity for phosphorylation of glucose. Glucose uptake was induced by glucose and not gluconate, thus, establishing independent regulation of glucose transport and glucose catabolism in p. fluorescens. The results prove that glucose oxidase was not an obligatory reaction for glucose carbon permeation in P. fluorescens. A general unifying scheme for glucose utilization in the aerobic fluorescent pseudomonads is suggested for the purpose of clarifying glucose uptake in these bacteria.

Aerobiosis↗
PubMed1975

6-Phosphogluconate dehydratase deficiency in pleiotropic carbohydrate-negative mutant strains of Pseudomonas aeruginosa.

Mutants of Pseudomonas aeruginosa, strain PAO, have been isolated that are unable to grow on mannitol, glucose, gluconate, or 2-ketogluconate, cut that exhibit wild-type growth on pyruvate, lactate, citrate, succinate, or acetate. Although some of these mutants were also unable to grow on glycerol, the mutations formed a single linkage group by quantitative transductional analysis with phage F116 on glucose minimal agar medium. Cell extracts of all mutant strains were either lacking or severely deficient in 6-phosphogluconate dehydratase activity. Glu+ transductants derived from mutant strains that retained the wild-type ability for growth at the expense of glycerol also regained the ability to grow on all C-6 compounds. Although a role for the pentose phosphate pathway in the catabolism of C6 substrates was not found, a functional Entner-Doudoroff pathway appears to be essential for the catabolism of mannitol, glucose, gluconate, and 2-ketogluconate.

Aldehyde-Lyases↗