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A Kotyk

Publications and source records attributed to A Kotyk.

At least 73 records · Page 4Linked to original sources
PubMed1988

Distinct affinity and effector residues in the binding site for a regulatory ligand. The mitochondrial uncoupling protein as a model.

A hypothesis concerning two distinct classes of amino acid residues in some regulatory binding sites is proposed. The "affinity residues" are those that are unable to transduce the ligand information signal but are responsible for overcoming the barrier for the attachment of a ligand to its binding site while the "effector residues" transfer the binding signal to the other functional part of the protein, which then undergoes a non-equilibrium energetic cycle induced by interaction with the ligand. As an example, the purine nucleotide inhibition of H+ transport through the uncoupling protein of brown adipose tissue mitochondria is discussed; there is a concentration range in which the nucleotide is bound but does not inhibit H+ transport. This is interpreted in terms of inaccessibility of the effector residues inducing H+ transport inhibition below a certain threshold concentration.

Adenosine Triphosphate↗
PubMed1988

Effect of ethanol on the specific transport system for L-lysine in Saccharomyces cerevisiae.

Preincubation of resting cells of Saccharomyces cerevisiae double mutant can1 gap1 (with a single transport system for L-lysine) with metabolic substrates stimulated subsequent uptake of lysine. While in the wild type the stimulation is connected primarily with carrier protein synthesis (delayed, cycloheximide-inhibitable effect) in the mutant an immediate tapping of an energy source (antimycin-inhibited) is practically solely involved.

Amino Acids↗
PubMed1988

Uptake of L-lysine by a double mutant of Saccharomyces cerevisiae.

A gap1 can1 mutant of Saccharomyces cerevisiae with a single lysine transport system remaining was used to study detailed kinetics of this transport. Its half-saturation constant was 78 mumol per litre, its maximum rate of transport was 0.29 mumol L-lysine per g dry matter per minute, both parameters being lower by more than an order of magnitude in comparison with the GAP system. The pH optimum lay at very acid values of about 3, the temperature dependence without any transition point showed an activation energy of 48 kJ/mol. The transport was inhibited by common metabolic inhibitors (3'-chlorophenylhydrazonomalononitrile, antimycin, 2-deoxy-D-glucose, sodium arsenate) as well as by a membrane-active one (uranyl nitrate). The specificity of the system was extremely high, none of the natural amino acids acting as competitor to L-lysine. The maximum accumulation ratio attained (at about 5 mg dry matter per mL) was 100: 1-120: 1, in agreement with the measured protonmotive force under the assumption of 1 H+ ion being transported with 1 lysine molecule. The ratio decreased with increasing external concentration of lysine to as little as 4: 1 at 1 mmol lysine per litre. It also decreased with increasing suspension density and it was at extremely low suspension densities (0.2 mg dry matter per mL) that ratios of as much as 500: 1 were reached. Application of group-specific inhibitors showed that the active site of the carrier contains an essential histidine residue.

Glucose↗
PubMed1988

Effect of chloroquine on membrane permeability in yeast--release of cellular coproporphyrin.

1. The influx and efflux of labelled substances with and without chloroquine was studied in yeast cells. 2. The uptake of delta-aminolevulinic acid by Saccharomyces cerevisiae is characterized by a KT of 3-4 mM and Jmax of 1.0-1.2 mumol min-1 g dry weight-1. 3. A method for loading yeast with labelled coproporphyrin is suggested. 4. The uptake of sorbitol and coproporphyrin was slightly stimulated, while the uptake of 6-deoxyglucose was slightly, that of 2-aminoisobutyric acid and leucine strongly inhibited by chloroquine. 5. The efflux of coproporphyrin, 2-aminoisobutyric acid and sorbitol was stimulated while that of leucine was not influenced by chloroquine. 6. The result showed that chloroquine influenced directly but nonspecifically the membrane permeability, apparently mainly that of the vacuolar membrane.

Aminoisobutyric Acids↗
PubMed1987

Effects of yeast suspension density on the accumulation ratio of transported solutes.

The previously described effect of cell suspension density on metabolic and transport phenomena in yeast, apparently caused by inhibition by dissolved carbon dioxide, is also observed with the accumulation ratio of both sugars and amino acids where not only a kinetic but also a energetic factor comes into play. Unlike all previously measured metabolic and transport parameters, the dependence of the accumulation ratio on suspension density is not monotonic but shows a pronounced maximum in the range of 4-8 mg dry wt/ml, depending on yeast species and on cultivation conditions. In Rhodotorula gracilis and in Lodderomyces elongisporus it is not due to CO2 but is semiquantitatively related to the proton-motive force across the plasma membrane as well as to the intracellular ATP content. It is observed both in oxygen and in argon, over a wide range of pH values and of temperatures, but it is suppressed by metabolic inhibitors. It is expressed only in a range of transported solute concentrations between about 0.1 and 10 mM.

Amino Acids↗
PubMed1985

Conditions of activation of yeast plasma membrane ATPase.

The in vivo activation of the H+-ATPase of baker's yeast plasma membrane found by Serrano in 1983 was demonstrated with D-glucose aerobically and anaerobically (as well as in a respiration-deficient mutant) and, after suitable induction, with maltose, trehalose, and galactose. The activated but not the control ATPase was sensitive to oligomycin. No activation was possible in a cell-free extract with added glucose. The ATPase was not activated in yeast protoplasts which may account for the absence of glucose-stimulated secondary active transports in these wall-less cells and provide support for a microscopic coupling between ATPase activity and these transports in yeast cells.

Cell Membrane↗
PubMed1985

Absence of glucose-stimulated transport in yeast protoplasts.

Protoplasts of Saccharomyces cerevisiae prepared by snail-gut juice treatment were compared in their transport properties with intact cells. 1. Constitutive monosaccharide transport (D-xylose, 6-deoxy-D-glucose), as well as inducible transport of D-galactose, were unaltered. 2. Phosphorylation-associated transport of 2-deoxy-D-glucose was enhanced in protoplasts, possibly as a consequence of removal of the unstirred layer of the cell wall. 3. Proton-driven transports of trehalose, L-leucine, L-proline and monophosphate could not be activated by preincubation with D-glucose, apparently owing to lack of proton-solute coupling in transport. Utilization of glucose was not depressed but respiration was reduced by about 50% while acidification of the external medium after glucose addition was inhibited by more than 90%. This may be related to the inability of protoplast plasma membrane H-ATPase to be activated by glucose and hence to impaired proton-translocating capacity. Uranyl ions inhibited generally much less in protoplasts than in intact cells although their binding to protoplasts was greater (maximum 0.68 fmol per cell but 3.2 fmol per protoplast).

Biological Transport↗
PubMed1985

Transport of ethanol in baker's yeast.

Ethanol is transported into various strains of baker's yeast by simple diffusion (no effect of inhibitors and a linear concentration dependence of the initial rate of uptake and final distribution in cells). It distributes itself in 96.6 +/- 16.2% of intracellular water.

Biological Transport↗
PubMed1985

Effects of suspension density on microbial metabolic processes.

Respiration of yeasts Saccharomyces cerevisiae, Rhodotorula glutinis, Endomyces magnusii, and Candida utilis, and of bacteria Escherichia coli and Bacillus megaterium, anaerobic production of CO2 by S. cerevisiae, active transport of quinovose by R. glutinis and of L-proline and L-leucine by S. cerevisiae were highly dependent on cell suspension density. Respiration of S. cerevisiae in the presence of glucose decreased in a biphasic fashion from 140 to 40 nmol O2 per mg dry solid per min as suspension density increased from 0.01 to 2 mg/mL. Higher partial pressures of oxygen further enhanced the trend. The active transports were affected monophasically in the maximum rate of uptake which was as much as ten times greater at low than at high suspension densities. A component of the external medium is suspected to cause the decrease of metabolic functions at higher cell densities, acting as a noncompetitive inhibitor. The component was present and mutually active in suspensions of the various yeasts as well as of bacteria. Its properties and results of model experiments suggest it to be dissolved carbon dioxide.

Anaerobiosis↗
PubMed1984

Intracellular pH distribution and transmembrane pH profile of yeast cells.

The pH-dependent fluorescence excitation of fluorescein located intracellularly and in the vicinity of cells of the yeast Saccharomyces cerevisiae and Endomyces magnusii was used to obtain local pH values at a linear resolution 0.2 micron. Cells suspended in water or in a diluted (5 mM) acidic buffer had a relatively alkaline interior (about 7.0-7.5) with pH decreasing gradually toward the periphery and further out through the cell wall to the value of the bulk solution. In slightly alkaline weak buffers the cells also showed an alkaline center and a slightly acidic ring-shaped area, but the peripheral region close to the membrane was again alkaline with pH increasing toward the bulk solution. The heterogeneity of intracellular pH was reduced or nearly abolished in starved or antimycin-treated cell. Suspension of cells in strong (200 mM) buffer resulted within 15-20 min in a nearly homogeneous pH pattern throughout the cell, attaining pH values of 5.5-7.5, depending on the pH of the buffer. Addition of glucose with concomitant pH decrease of the extracellular medium did not change appreciably the intracellular pattern for 20-30 min, except with diethylstilbestrol (inhibitor of proton-extruding ATPase) when the cell became more acidic. It appears that the delta pH measurements between the cell as a whole and the bulk solution (as are used for the calculation of the electrochemical potential of protons in proton-driven transports) are not substantiated, the probable pH difference across the plasma membrane being substantially smaller than previously supposed.

Ascomycota↗
PubMed1983

Transport of nutrients in yeast protoplasts.

Protoplasts were prepared with snail-gut juice from Saccharomyces cerevisiae and Candida utilis. Transport of D-xylose (A), trehalose (B), 2-deoxy-D-glucose (C), L-leucine (D), L-proline (E), inorganic phosphate (F) and H+ ions (G) was studied with special emphasis on the first species. Transport of A which is not sensitive to glucose stimulation in its synthesis was not affected by protoplast formation. Similarly unaffected were transports of B, D, E and F in their "residual" form in starved cells. However, transport of these substances after glucose-stimulated synthesis was practically fully suppressed by protoplast formation. This may be connected with the virtually complete inhibition of the proton pump (G) in protoplasts with the implication that B, D, E and F are transported in conjunction with protons but only those that are immediately produced by the proton pump (glucose consumption, oxygen utilization and membrane potential were not substantially altered in protoplasts). Transport of C was stimulated nearly two-fold in protoplasts. Uranyl ions (0.1 mM) had a pronouncedly lower inhibitory effect on all the transports studied in protoplasts.

Amino Acids↗
PubMed1983

Isolation of sulphate transport defective mutants of Candida utilis: further evidence for a common transport system for sulphate, sulphite and thiosulphate.

Selenate-resistant mutants of Candida utilis were isolated. They did not take up sulphate while incorporation of an organic sulphur source, such as L-methionine, was similar to the wild-type strain. They grew poorly on sulphate, sulphite and thiosulphate and, as expected, grew well on methionine. Sulphite reductase activities of the mutants were similar to the wild type strain. The properties of these mutants support the view of a common transport system for sulphate, sulphite and thiosulphate.

Biological Transport, Active↗
PubMed1983

Sulphate transport in Candida utilis.

Sulphate uptake by Candida utilis follows Michaelis-Menten type kinetics characterized by a Km of 1.43 mM for sulphate. The process is unidirectional, pH, temperature and energy dependent. Molybdate, selenate, thiosulphate, chromate and sulphite are competitive inhibitors. Dithionite is a mixed-type inhibitor of sulphate uptake. If cells are pre-incubated with sulphate, sulphite, thiosulphate, dithionite or sulphide, sulphate uptake is severely blocked. Inhibition by endogenous sulphate, sulphite and thiosulphate was specific for sulphate uptake. Thus, incorporation of extracellular sulphate seems to be under the control of a heterogeneous pool of sulphur compounds. These results are discussed in connection with the regulation of sulphur amino acid biosynthesis in C. utilis.

Biological Transport, Active↗
PubMed1983

Interaction of nystatin with nystatin-resistant Candida tropicalis.

Nystatin-resistant yeast Candida tropicalis was obtained after UV illumination and plating on nystatin-containing media. The mutant contained no ergosterol in the plasma membrane but bound nystatin to a degree similar to that of the wild strain (1.2 vs. 1.5 nmol per mg dry solid). Respiration of the mutant on glucose was reduced by 36% in the presence of 25 microM nystatin. This corresponded to a 25-43% decrease of the uptake of monosaccharides. Transport of amino acids was reduced by nystatin in the mutant by 44-86%, as compared with a 84-95% reduction in the wild strain. The intracellular ATP content was reduced by nystatin equally in the wild strain and in the mutant (by 43 and 47%). Nystatin appears to affect specifically membrane transport processes of nonelectrolytes while both the H+-extruding ATPase and the membrane potential are unaffected.

Biological Transport, Active↗
PubMed1982

Anilinonaphthalene sulfonate fluorescence and amino acid transport in yeast.

Fluorescence of 1-anilinonaphthalene-8-sulfonate in yeast membranes appears to be caused predominantly by binding to lipids (ANS protein:ANS lipid approximately 1 : 20) as indicated by the fluorescence lifetime, degree of polarization, and excitation spectra. It was insensitive to short-circuiting the membrane potential. Fluorescence intensity increased as cells (especially after pretreatment with energy donors such as glucose) were exposed to some amono acids, in particular, aspartic and glutamic acids. The character of fluorescence shifted to that of protein-bound ANS, suggesting an exposure of new protein sites accessible to the probe. This shift could be prevented by inhibitors of energy transduction as well as of transport. The K1/2 of the shift was at 2.5 mM aspartic acid.

Amino Acids↗
PubMed1982

Maltotriose transport and utilization in baker's and brewer's yeast.

Maltotriose is metabolized by baker's and brewer's yeast only oxidatively, with a respiratory quotient of 1.0, the QCO2Ar being, depending on the strain used, 0-11, as compared with QCO2air of 6-42 microL CO2 per h per mg dry substance. The transport appeared to proceed by facilitated diffusion (no effects of NaF, iodoacetamide and 3-chlorophenylhydrazonomalononitrile) with a KT of more than 50 mM and was inhibited by maltose greater than maltotriose greater than methyl-alpha-D-glucoside greater than maltotetraose greater than D-fructose greater than D-glucose. The transport was present constitutively in both Saccharomyces cerevisiae (baker's yeast) and in S. uvarum (brewer's yeast) and it was not significantly stimulated by preincubation with glucose or maltose. The pH optimum was 4.5-5.5, the temperature dependence yielded an activation energy of 26 kJ/mol.

Biological Transport↗