scholarly journals Active transport of charged substrates by a proton/sugar co-transport system. Amino-sugar uptake in the yeast Rhodotorula gracilis

1981 ◽  
Vol 194 (2) ◽  
pp. 433-441 ◽  
Author(s):  
C Niemietz ◽  
R Hauer ◽  
M Höfer
Keyword(s):  
Membrane Potential ◽  
Amino Sugar ◽  
Protonated Form ◽  
Sugar Uptake ◽  
Amino Sugars ◽  
Maximal Velocity ◽  
Deprotonated Form ◽  
Half Saturation Constant ◽  
Carbonyl Cyanide ◽  
Rhodotorula Gracilis

1. In the yeast Rhodotorula gracilis several amino sugars were actively transported. Glucosamine, which is largely protonated at physiological pH (pK 7.75) was used as a model substrate. At pH 6.75 its half-saturation constant was 1 mM and the maximal velocity was 50 nmol/min per mg dry wt. 2. Amino sugars were taken up via the monosaccharide carrier. The transport of glucosamine was strongly restricted by monosaccharides. D-Xylose inhibited competitively the uptake of glucosamine. The inhibition constant was 1 mM. Cells preloaded with D-xylose showed exchange transport on subsequent addition of glucosamine. 3. Transport of glucosamine was energized by the membrane potential. Uncoupling agents such as carbonyl cyanide m-chlorophenyl-hydrazone and the lipophilic cation TPP+ (tetraphenylphosphonium ion) at concentrations that depolarized the membrane potential inhibited the uptake of glucosamine. Conversely the transport of glucosamine partly dissipated the membrane potential, which was monitored by radioactively labelled lipophilic cations. 4. The translocated charges were electrically compensated by the extrusion of protons and K+ (1 glucosamine molecule/0.85 H+ + 0.15 K+). 5. An increase of the pH in the range 4.75-8.75 lead to a decrease of the half-saturation constant from 5 mM to 1 mM and to an optimum of the maximal velocity at pH 6.75. We suggest that this fair constancy is due to the carrier not distinguishing between the protonated form of glucosamine (pH less than 7.75) and the deprotonated form (pH greater than 7.75). The increase of V(T) (maximal transport velocity) between pH 4.75 and 6.75 is due to the increase of the membrane potential: the decrease between pH 6.75 and 8.75 is due to the deprotonization of the carrier.

scholarly journals The Hexose-Proton Cotransport System of Chlorella

1974 ◽  
Vol 64 (5) ◽  
pp. 568-581 ◽  
Author(s):  
Ewald Komor ◽  
Widmar Tanner
Keyword(s):  
Sugar Transport ◽  
Protonated Form ◽  
Weak Acid ◽  
Sugar Uptake ◽  
Sugar Accumulation ◽  
Maximal Velocity ◽  
Uptake System ◽  
High Affinity ◽  
Ph Values ◽  
Proton Concentration

The proton concentration in the medium affects the maximal velocity of sugar uptake with a Km of 0.3 mM (high affinity uptake). By decreasing the proton concentration a decrease in high affinity sugar uptake is observed, in parallel the activity of a low affinity uptake system (Km of 50 mM) rises. Both systems add up to 100%. The existence of the carrier in two conformational states (protonated and unprotonated) has been proposed therefore, the protonated form with high affinity to 6-deoxyglucose, the unprotonated form with low affinity. A plot of extrapolated Vmax values at low substrate concentration versus proton concentration results in a Km for protons of 0.14 µM, i.e. half-maximal protonation of the carrier is achieved at pH 6.85. The stoichiometry of protons cotransported per 6-deoxyglucose is close to 1 at pH 6.0–6.5. At higher pH values the stoichiometry continuously decreases; at pH 8.0 only one proton is cotransported per four molecules of sugar. Whereas the translocation of the protonated carrier is strictly dependent on sugar this coupling is less strict for the unprotonated form. Therefore at alkaline pH a considerable net efflux of accumulated sugar can occur. The dependence of sugar accumulation on pH has been measured. The decrease in accumulation with higher pH values can quantitatively be explained by the decrease in the amount of protonated carrier. The properties of the unprotonated carrier resemble strikingly the properties of carrier at the inner side of the membrane. The inside pH of Chlorella was measured with the weak acid 5,5-dimethyl-2, 4-oxazolidinedion (DMO). At an outside pH of 6.5 the internal pH was found to be 7.2. To explain the extent of sugar accumulation it has to be assumed that the membrane potential also contributes to active sugar transport in this alga.

scholarly journals Evidence for a proton/sugar symport in the yeast Rhodotorula gracilis (glutinis)

1978 ◽  
Vol 172 (1) ◽  
pp. 15-22 ◽  
Author(s):  
M Höfer ◽  
P C Misra
Keyword(s):  
Sugar Uptake ◽  
Sugar Accumulation ◽  
Electrochemical Gradient ◽  
Apparent Dissociation Constant ◽  
Physiological Conditions ◽  
Half Saturation Constant ◽  
Monosaccharide Transport ◽  
Rhodotorula Gracilis ◽  
Proton Uptake ◽  
Effective Carrier

1. The uptake of monosaccharides and polyols in the obligatory aerobic yeast Rhodotorula gracilis (glutinis) was accompanied by proton uptake. 2. The half-saturation constant of transport, KT, depended on pH, changing from about 2mM at pH 4.5 to 80mM at pH8.5 for D-xylose; this change of the effective carrier affinity was reversible. 3. The apparent dissociation constant of the monosaccharide carrier was estimated at pKa 6.75. 4. At pH8.5, when the pH gradient across the cell membrane vanished, no sugar accumulation was demonstrable. 5. The half-saturation constants of sugar uptake and H+ co-transport were very similar to each other, the latter obviously being controlled by the former. 6. The H+/sugar stoicheiometry remained constant under various physiological conditions; it amounted to one H+ ion per sugar molecule taken up. 7. The data are interpreted as a strong piece of evidence in favour of the active monosaccharide transport in R. gracilis (glutinis) being an H+-symport energized by the electrochemical gradient of H+ across the plasma membrane of the yeast.

scholarly journals Divergent Accumulation of Microbial Residues and Amino Sugars in Loess Soil after Six Years of Different Inorganic Nitrogen Enrichment Scenarios

2021 ◽  
Vol 11 (13) ◽  
pp. 5788
Author(s):  
Dominic Kwadwo Anning ◽  
Zhilong Li ◽  
Huizhen Qiu ◽  
Delei Deng ◽  
Chunhong Zhang ◽  
...  
Keyword(s):  
Plant Biomass ◽  
Inorganic Nitrogen ◽  
Amino Sugar ◽  
N Fertilization ◽  
Biochemical Properties ◽  
Sugar Accumulation ◽  
Amino Sugars ◽  
N Enrichment ◽  
Over Time ◽  
Microbial Residues

Amino sugars are key microbial biomarkers for determining the contribution of microbial residues in soil organic matter (SOM). However, it remains largely unclear as to what extent inorganic nitrogen (N) fertilization can lead to the significant degradation of SOM in alkaline agricultural soils. A six-year field experiment was conducted from 2013 to 2018 to evaluate the effects of chronic N enrichment on microbial residues, amino sugars, and soil biochemical properties under four nitrogen (urea, 46% N) fertilization scenarios: 0 (no-N, control), 75 (low-N), 225 (medium-N), and 375 (high-N) kg N ha−1. The results showed that chronic N enrichment stimulated microbial residues and amino sugar accumulation over time. The medium-N treatment increased the concentration of muramic acid (15.77%), glucosamine (13.55%), galactosamine (18.84%), bacterial residues (16.88%), fungal residues (11.31%), and total microbial residues (12.57%) compared to the control in 2018; however, these concentrations were comparable to the high-N treatment concentrations. The ratio of glucosamine to galactosamine and of glucosamine to muramic acid decreased over time due to a larger increase in bacterial residues as compared to fungal residues. Microbial biomass, soil organic carbon, and aboveground plant biomass positively correlated with microbial residues and amino sugar components. Chronic N enrichment improved the soil biochemical properties and aboveground plant biomass, which stimulated microbial residues and amino sugar accumulation over time.

Monitoring of mitochondrial membrane potential in isolated perfused rat heart

1984 ◽  
Vol 247 (4) ◽  
pp. H508-H516
Author(s):  
R. A. Kauppinen ◽  
I. E. Hassinen
Keyword(s):  
Membrane Potential ◽  
Mitochondrial Membrane Potential ◽  
Mitochondrial Membrane ◽  
Optical Methods ◽  
Perfused Heart ◽  
Rat Hearts ◽  
Carbonyl Cyanide ◽  
Perfused Rat Hearts ◽  
Beating Rate ◽  
Isolated Perfused Rat Hearts

Optical methods were tested for measuring the membrane potential changes of mitochondria in isolated perfused rat hearts. Safranin was found to be rapidly taken up by the Langendorff-perfused heart, and after loading with the dye there was practically no washout of the stain during perfusion with Krebs-Ringer bicarbonate solution. Staining with safranin induced the appearance of an intense absorption band in the reflectance spectrum of the heart, but the absorbance spectrum changes were not useful for monitoring the mitochondrial membrane potential changes because of interference by endogenous hemoproteins. The fluorescence intensity, however, responded in a manner which indicated that its changes originated from dye attached to the mitochondria. A decrease of the fluorescence was found on energizing the mitochondria by decreasing the cellular energy consumption by arrest induced by 18 mM K+ or by decreasing the beating rate of an electrically paced heart from 5 Hz to the endogenous ventricular frequency of 1.5 Hz. In hearts arrested by Ca2+ depletion, 18 mM K+ did not affect the safranin fluorescence. This was taken to indicate that under these conditions the safranin fluorescence was not sensitive to the plasma membrane potential. The uncoupler carbonyl cyanide m-chlorophenylhydrazone induced an intense enhancement of safranin fluorescence in the perfused heart, demonstrating that the probe is sensitive to mitochondrial membrane potential.(ABSTRACT TRUNCATED AT 250 WORDS)

Comprehensive assessment of precursors, diagenesis, and reactivity to water treatment of dissolved and colloidal organic matter

2004 ◽  
Vol 4 (4) ◽  
pp. 1-9 ◽  
Author(s):  
J.A. Leenheer
Keyword(s):  
Organic Matter ◽  
Water Treatment ◽  
Condensed Tannins ◽  
Drinking Water Treatment ◽  
Amino Sugar ◽  
Amino Sugars ◽  
Research Approach ◽  
Soil Aquifer Treatment ◽  
Treatment Processes ◽  
High Yields

A comprehensive isolation, fractionation, and characterization research approach was developed for dissolved and colloidal organic matter (DOM) in water, and it was applied to various surface- and groundwaters to assess DOM precursors, DOM diagenesis, and DOM reactivity to water treatment processes. Major precursors for natural DOM are amino sugars, condensed tannins, and terpenoids. Amino sugar colloids derived from bacterial cell walls are incompletely removed by drinking water treatment and foul reverse osmosis membranes, but are nearly quantitatively removed by soil/aquifer treatment. When chlorinated, amino sugars produce low yields of regulated disinfection by-products (DBPs) but they produce significant chlorine demand that is likely caused by chlorination of free amino groups. Condensed tannins are major precursors for ,blackwater- DOM such as that found in the Suwannee River. This DOM produces high yields of DBPs upon chorination, and is efficiently removed by coagulation/flocculation treatment. Terpenoid-derived DOM appears to be biologically refractory, infiltrates readily into groundwater with little removal by soil/aquifer treatment, gives low DBP-yields upon chlorination and is poorly removed by coagulation/flocculation treatments. Peptides derived from proteins are major components of the base DOM fraction (10% or less of the mass of DOM), and this fraction produces large yields of haloacetonitriles upon chorination.

Effect of membrane potential and cellular ATP on glutathione efflux from isolated rat hepatocytes

1988 ◽  
Vol 255 (4) ◽  
pp. G403-G408 ◽  
Author(s):  
J. C. Fernandez-Checa ◽  
C. Ren ◽  
T. Y. Aw ◽  
M. Ookhtens ◽  
N. Kaplowitz
Keyword(s):  
Membrane Potential ◽  
High Performance ◽  
Direct Relationship ◽  
Rat Hepatocytes ◽  
Antimycin A ◽  
Total Glutathione ◽  
Rat Hepatocyte ◽  
Isolated Rat Hepatocytes ◽  
Carbonyl Cyanide ◽  
Isolated Rat

total glutathione (GSH) efflux was studied in isolated rat hepatocyte suspensions at repleted GSH content (45-55 nmol/10(6) cells). The increase in concentrations of medium K+ in place of Na+ caused a parallel fall in membrane potential and total GSH efflux. Ouabain (1 mM) and replacement of Na+ with choline caused a gradual fall in membrane potential and GSH efflux. Hyperpolarization of hepatocytes with lipophilic anions, thiocyanate, and nitrate was associated with significantly increased efflux. Total GSH efflux was inhibited by increasing concentrations of fructose, antimycin A, and carbonyl cyanide p-trifluoromethoxyphenylhydrazone, and there was a direct relationship between the rate of efflux and cellular ATP. Changes in total GSH efflux were paralleled by changes in GSH determined by high-performance liquid chromatography. Vanadate markedly inhibited efflux but caused only a modest decrease in cellular ATP. Fructose, antimycin A, and vanadate did not affect membrane potential or cell volume under the conditions at which efflux was inhibited. These results suggest independent requirements for both membrane potential and ATP in the transport of GSH.

scholarly journals Structural and functional analysis of the solute-binding protein UspC from Mycobacterium tuberculosis that is specific for amino sugars

Open Biology ◽  
2016 ◽  
Vol 6 (6) ◽  
pp. 160105 ◽  
Author(s):  
Elizabeth Fullam ◽  
Ivan Prokes ◽  
Klaus Fütterer ◽  
Gurdyal S. Besra
Keyword(s):  
Mycobacterium Tuberculosis ◽  
Abc Transporters ◽  
Binding Protein ◽  
Functional Characterization ◽  
Amino Sugar ◽  
Amino Sugars ◽  
Environmental Niche ◽  
Intracellular Infection ◽  
Shift Analysis ◽  
Binding Cleft

Mycobacterium tuberculosis ( Mtb ), the aetiological agent of tuberculosis, has evolved to scavenge nutrients from the confined environment of host macrophages with mycobacterial ATP-binding cassette (ABC) transporters playing a key role in nutrient acquisition. Mtb -UspC (Rv2318) is the solute-binding protein of the essential transporter UspABC, one of four Mtb ABC transporters implicated by homology in sugar acquisition. Herein, we report the structural and functional characterization of Mtb -UspC. The 1.5 Å resolution structure of UspC reveals a two subdomain architecture that forms a highly acidic carbohydrate-substrate binding cleft. This has allowed a distinct preference of Mtb -UspC for amino sugars as determined by thermal shift analysis and solution saturation transfer difference-NMR. Taken together our data support the functional assignment of UspABC as an amino-sugar transporter. Given the limited availability of carbohydrates within the phagosomal environmental niche during Mtb intracellular infection, our studies suggest that UspABC enables Mtb to optimize the use of scarce nutrients during intracellular infection, linking essentiality of this protein to a potential role in recycling components of cell-wall peptidoglycan.

scholarly journals CO2 Uptake and Fixation by Endosymbiotic Chemoautotrophs from the Bivalve Solemya velum

2006 ◽  
Vol 73 (4) ◽  
pp. 1174-1179 ◽  
Author(s):  
Kathleen M. Scott ◽  
Colleen M. Cavanaugh
Keyword(s):  
Inorganic Carbon ◽  
Carbon Fixation ◽  
Dissolved Inorganic Carbon ◽  
Co2 Uptake ◽  
Maximal Velocity ◽  
Marine Habitats ◽  
Half Saturation Constant ◽  
Oxygen Conditions ◽  
Solemya Velum ◽  
Influence Of Ph

ABSTRACT Chemoautotrophic symbioses, in which endosymbiotic bacteria are the major source of organic carbon for the host, are found in marine habitats where sulfide and oxygen coexist. The purpose of this study was to determine the influence of pH, alternate sulfur sources, and electron acceptors on carbon fixation and to investigate which form(s) of inorganic carbon is taken up and fixed by the gamma-proteobacterial endosymbionts of the protobranch bivalve Solemya velum. Symbiont-enriched suspensions were generated by homogenization of S. velum gills, followed by velocity centrifugation to pellet the symbiont cells. Carbon fixation was measured by incubating the cells with 14C-labeled dissolved inorganic carbon. When oxygen was present, both sulfide and thiosulfate stimulated carbon fixation; however, elevated levels of either sulfide (>0.5 mM) or oxygen (1 mM) were inhibitory. In the absence of oxygen, nitrate did not enhance carbon fixation rates when sulfide was present. Symbionts fixed carbon most rapidly between pH 7.5 and 8.5. Under optimal pH, sulfide, and oxygen conditions, symbiont carbon fixation rates correlated with the concentrations of extracellular CO2 and not with HCO3 − concentrations. The half-saturation constant for carbon fixation with respect to extracellular dissolved CO2 was 28 � 3 μM, and the average maximal velocity was 50.8 � 7.1 μmol min−1 g of protein−1. The reliance of S. velum symbionts on extracellular CO2 is consistent with their intracellular lifestyle, since HCO3 − utilization would require protein-mediated transport across the bacteriocyte membrane, perisymbiont vacuole membrane, and symbiont outer and inner membranes. The use of CO2 may be a general trait shared with many symbioses with an intracellular chemoautotrophic partner.

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