This review summarizes the main results obtained in the fields of general and molecular microbiology and microbial genetics at the Institute of Microbiology of the Academy of Sciences of the Czech Republic (AS CR) [formerly Czechoslovak Academy of Sciences (CAS)] over more than 50 years. Contribution of the founder of the Institute, academician Ivan Málek, to the introduction of these topics into the scientific program of the Institute of Microbiology and to further development of these studies is also included.

• MeSH
• Academies and Institutes history   MeSH
• History, 20th Century MeSH
• Genetics, Microbial history   MeSH
• Molecular Biology history   MeSH
• Check Tag
• History, 20th Century MeSH
• Publication type
• Journal Article MeSH
• Historical Article MeSH
• Research Support, Non-U.S. Gov't MeSH
• Review MeSH
• Geographicals
• Czech Republic MeSH

The rate and extent of uptake of the fluorescent probe diS-C3(3) reporting on membrane potential in S. cerevisiae is affected by the strain under study, cell-growth phase, starvation and by the concentration of glucose both in the growth medium and in the monitored cell suspension under non-growth conditions. Killer toxin K1 brings about changes in membrane potential. In all types of cells tested, viz. in glucose-supplied stationary or exponential cells of the killer-sensitive strain S6/1 or a conventional strain RXII, or in glucose-free exponential cells of both strains, both active and heat-inactivated toxin slow down the potential-dependent uptake of diS-C3(3) into the cells. This may reflect "clogging" of pores in the cell wall that hinders, but does not prevent, probe passage to the plasma membrane and its equilibration. The clogging effect of heat-inactivated toxin is stronger than that exerted by active toxin. In susceptible cells, i.e. in exponential-phase glucose-supplied cells of the sensitive strain S6/1, this phase of probe uptake retardation is followed by an irreversible red shift in probe fluorescence maximum lambda max indicating damage to membrane integrity and cell permeabilization. A similar fast red shift in lambda max signifying lethal cell damage was found in heat-killed or nystatin-treated cells.

• MeSH
• Fluorescent Dyes metabolism   MeSH
• Fungal Proteins pharmacology   MeSH
• Carbocyanines metabolism   MeSH
• Killer Factors, Yeast MeSH
• Membrane Potentials drug effects   MeSH
• Mycotoxins pharmacology   MeSH
• Nystatin pharmacology   MeSH
• Saccharomyces cerevisiae physiology   MeSH
• Publication type
• Journal Article MeSH
• Research Support, Non-U.S. Gov't MeSH
• Names of Substances
• 3,3'-dipropylthiacarbocyanine MeSH Browser
• Fluorescent Dyes MeSH
• Fungal Proteins MeSH
• K1 killer toxin MeSH Browser
• Carbocyanines MeSH
• Killer Factors, Yeast MeSH
• Mycotoxins MeSH
• Nystatin MeSH

Three fluorescent probes, tetramethyl rhodamine ethyl ester (TMRE), 3,3'-dipropylthiacarbocyanine iodide (diS-C3(3)) and 3,3'-dipropyloxacarbocyanine iodide (diO-C3(3)), were tested for their suitability as fluorescent indicators of membrane potential in Saccharomyces cerevisiae in studies performed by flow cytometry. For all these dyes the intensity of fluorescence of stained cells increased with probe concentration in the range of 60-3000 nmol/L. The optimum staining period was 15-20 min for diS-C3(3). Depolarization of cells by increased extracellular potassium level and by valinomycin elicited with all probes a drop in fluorescence intensity. In some yeast batches this depolarization was accompanied by a separation of subpopulations with different fluorescence properties.

• MeSH
• Artifacts MeSH
• Benzothiazoles MeSH
• Time Factors MeSH
• Fluorescence MeSH
• Fluorescent Dyes * MeSH
• Fluorometry * MeSH
• Indicators and Reagents MeSH
• Carbocyanines * MeSH
• Membrane Potentials * MeSH
• Organometallic Compounds MeSH
• Flow Cytometry methods   MeSH
• Pyridinium Compounds analysis   metabolism   MeSH
• Scattering, Radiation MeSH
• Saccharomyces cerevisiae physiology   MeSH
• Publication type
• Journal Article MeSH
• Research Support, Non-U.S. Gov't MeSH
• Names of Substances
• 1-(3-sulfonatopropyl)-4-(beta)(2-(di-n-butylamino)-6-naphthylvinyl)pyridinium betaine MeSH Browser
• 3,3'-dipropyl-2,2'-thiadicarbocyanine MeSH Browser
• 3,3'-dipropyloxadicarbocyanine MeSH Browser
• 3,3'-dipropylthiacarbocyanine MeSH Browser
• Benzothiazoles MeSH
• Fluorescent Dyes * MeSH
• Indicators and Reagents MeSH
• Carbocyanines * MeSH
• Organometallic Compounds MeSH
• Pyridinium Compounds MeSH
• tetramethyl rhodamine ethyl ester MeSH Browser

• MeSH
• Genetic Techniques MeSH
• Hybridization, Genetic MeSH
• Killer Factors, Yeast MeSH
• Mutation MeSH
• Mycotoxins biosynthesis   genetics   pharmacology   MeSH
• Industry MeSH
• Saccharomyces cerevisiae Proteins MeSH
• Saccharomyces cerevisiae drug effects   genetics   metabolism   MeSH
• Transformation, Genetic MeSH
• Publication type
• Journal Article MeSH
• Research Support, Non-U.S. Gov't MeSH
• Review MeSH
• Names of Substances
• KHR1 protein, S cerevisiae MeSH Browser
• Killer Factors, Yeast MeSH
• Mycotoxins MeSH
• Saccharomyces cerevisiae Proteins MeSH

The minimum period between the addition of killer toxin K1 to sensitive yeast cells and the appearance of first cells stained with bromocresol purple indicating membrane damage, is approximately 20 min. The length of this lag phase depends strongly on toxin concentration, extending sharply at toxin levels lower than 60 lethal units (LU) per cell (about one-tenth of the toxin concentration necessary for saturating all surface receptors). As the binding of the toxin to the cell is virtually complete within 1 min, the rest of the lag phase reflects processes different from actual binding, e.g. combination of several toxin molecules to form a membrane ion channel or pore.

• MeSH
• Cell Membrane drug effects   metabolism   MeSH
• Ion Channels drug effects   metabolism   MeSH
• Killer Factors, Yeast MeSH
• Kinetics MeSH
• Mycotoxins metabolism   pharmacology   MeSH
• Cell Membrane Permeability drug effects   MeSH
• Saccharomyces cerevisiae drug effects   metabolism   MeSH
• Publication type
• Journal Article MeSH
• Research Support, Non-U.S. Gov't MeSH
• Names of Substances
• Ion Channels MeSH
• K1 killer toxin MeSH Browser
• Killer Factors, Yeast MeSH
• Mycotoxins MeSH

A recently described new method for determination of killer toxin activity was used for kinetic measurements of K1 toxin binding. The cells of the killer sensitive strain Saccharomyces cerevisiae S6 were shown to carry two classes of toxin binding sites differing widely in their half-saturation constants and maximum binding rates. The low-affinity and high-velocity binding component (KT1 = 2.6 x 10(9) L.U./ml, Vmax1 = 0.19 s-1) probably reflects diffusion-limited binding to cell wall receptors; the high-affinity and low-velocity component (KT2 = 3.2 x 10(7) L.U./ml, Vmax2 = 0.03 s-1) presumably indicates the binding of the toxin to plasma membrane receptors. Adsorption of most of the killer toxin K1 to the surface of sensitive cells occurred within 1 min and was virtually complete within 5 min. The amount of toxin that saturated practically all cell receptors was about 600 lethal units (L.U.) per cell of S. cerevisiae S6.

• MeSH
• Adsorption MeSH
• Cell Membrane metabolism   MeSH
• Killer Factors, Yeast MeSH
• Kinetics MeSH
• Mycotoxins metabolism   MeSH
• Receptors, Cell Surface classification   metabolism   MeSH
• Saccharomyces cerevisiae metabolism   MeSH
• Publication type
• Journal Article MeSH
• Research Support, Non-U.S. Gov't MeSH
• Names of Substances
• K1 killer toxin MeSH Browser
• Killer Factors, Yeast MeSH
• Mycotoxins MeSH
• Receptors, Cell Surface MeSH

• MeSH
• Potassium pharmacology   MeSH
• Fluorescent Dyes MeSH
• Carbocyanines MeSH
• Killer Factors, Yeast MeSH
• Membrane Potentials MeSH
• Mycotoxins pharmacology   MeSH
• Saccharomyces cerevisiae physiology   MeSH
• Publication type
• Journal Article MeSH
• Research Support, Non-U.S. Gov't MeSH
• Names of Substances
• 3,3'-dipropylthiacarbocyanine MeSH Browser
• Potassium MeSH
• Fluorescent Dyes MeSH
• K1 killer toxin MeSH Browser
• Carbocyanines MeSH
• Killer Factors, Yeast MeSH
• Mycotoxins MeSH

Optimal assay conditions were established for the previously described method used to determine the activity ofSaccharomyces cerevisiae pore-forming killer toxin K1. The method is based on cell staining with bromocresol purple. Sensitive cells ofS. cerevisiae from the early exponential phase under nongrowth conditions and in the presence of glucose were the most convenient for determining the killer toxin activity. Maximum killing war reached when the suspension was buffered with 10 mM citrate-phosphate at pH 4.6.

• Publication type
• Journal Article MeSH

Optimum conditions for action of the killer toxin K1 on sensitive strain S. cerevisiae S6 were established. Maximum killing was reached in a very narrow pH range of 4.5-4.6. Maximum susceptibility to toxin was displayed by highly energized fresh cells from the early exponential phase in the presence of an external energy source (at least 200 mmol/L glucose). Further, maintenance of maximum membrane potential was necessary for killer action, as documented by decreasing toxin activity in the presence of increasing concentrations of KCl. The killing was strongly stimulated in the presence of millimolar concentrations of Ca2+ and Mg2+.

• MeSH
• Drug Resistance, Microbial MeSH
• Potassium Chloride pharmacology   MeSH
• Energy Metabolism MeSH
• Magnesium pharmacology   MeSH
• Killer Factors, Yeast MeSH
• Hydrogen-Ion Concentration MeSH
• Mycotoxins toxicity   MeSH
• Saccharomyces cerevisiae Proteins MeSH
• Saccharomyces cerevisiae drug effects   metabolism   MeSH
• Calcium pharmacology   MeSH
• Publication type
• Journal Article MeSH
• Names of Substances
• Potassium Chloride MeSH
• Magnesium MeSH
• Killer Factors, Yeast MeSH
• Mycotoxins MeSH
• Saccharomyces cerevisiae Proteins MeSH
• Calcium MeSH