On the coupling of intracellular to glycolytic oscillations in yeast.

We have investigated the interplay between glycolytic oscillations and intracellular concentration in the yeast Saccharomyces cerevisiae. Intracellular concentration was measured using the fluorophore potassium-binding benzofuranisophthalate (PBFI). We found that is an essential ion for the occurrence of glycolytic oscillations and that intracellular concentration oscillates synchronously with other variables such as nicotinamide adenine dinucleotide hydride (NADH), intracellular adenosine triphosphate (ATP), and mitochondrial membrane potential.

Chaos in the peroxidase-oxidase oscillator.

The peroxidase-oxidase (PO) reaction involves the oxidation of reduced nicotinamide adenine dinucleotide by molecular oxygen. When both reactants are supplied continuously to a reaction mixture containing the enzyme and a phenolic compound, the reaction will exhibit oscillatory behavior. In fact, the reaction exhibits a zoo of dynamical behaviors ranging from simple periodic oscillations to period-doubled and mixed mode oscillations to quasiperiodicity and chaos.

Functional imaging of a model unicell: Spironucleus vortens as an anaerobic but aerotolerant flagellated protist.

Advances in optical microscopy are continually narrowing the chasm in our appreciation of biological organization between the molecular and cellular levels, but many practical problems are still limiting. Observation is always limited by the rapid dynamics of ultrastructural modifications of intracellular components, and often by cell motility: imaging of the unicellular protist parasite of ornamental fish, Spironucleus vortens, has proved challenging. Autofluorescence of nicotinamide nucleotides and flavins in the 400-580 nm region of the visible spectrum, is the most useful indicator of cellular redox state and hence vitality.

Ergosterol is mainly located in the cytoplasmic leaflet of the yeast plasma membrane.

Transbilayer lipid asymmetry is a fundamental characteristic of the eukaryotic cell plasma membrane (PM). While PM phospholipid asymmetry is well documented, the transbilayer distribution of PM sterols such as mammalian cholesterol and yeast ergosterol is not reliably known. We now report that sterols are asymmetrically distributed across the yeast PM, with the majority (~80%) located in the cytoplasmic leaflet.

An experimental study of the regulation of glycolytic oscillations in yeast.

We have studied oscillating glycolysis in the strain BY4743 and isogenic strains with deletions of genes encoding enzymes in glycolysis, mitochondrial electron transport and ATP synthesis. We found that deletion of the gene encoding the hexokinase 1 isoform does not affect the oscillations while deletion of the gene encoding the hexokinase 2 isoform results in oscillations with smaller amplitude. The latter is associated with an almost 50% decrease in hexokinase activity.

Nanoparticle embedded enzymes for improved lateral flow sensors.

In this study, combining the nanoparticle embedded sensors with lateral flow assays, a novel strategy for ensuring the quality of signalling in lateral flow assays (LFAs) was developed. A LFA for reactive oxygen species (ROS) is reported that is based on horse radish peroxidase (HRP) which is co-entrapped with Texas Red dextran inside porous polyacrylamide nanoparticles. In this system, enzymes are protected in the porous matrix of polyacrylamide which freely allows the diffusion of the analyte.

Regulation of glycolytic oscillations by mitochondrial and plasma membrane H+-ATPases.

We investigated the coupling between glycolytic and mitochondrial membrane potential oscillations in Saccharomyces cerevisiae under semianaerobic conditions. Glycolysis was measured as NADH autofluorescence, and mitochondrial membrane potential was measured using the fluorescent dye 3,3'-diethyloxacarbocyanine iodide. The responses of glycolytic and membrane potential oscillations to a number of inhibitors of glycolysis, mitochondrial electron flow, and mitochondrial and plasma membrane H(+)-ATPase were investigated.

Human myeloperoxidase catalyzes an oscillating peroxidase-oxidase reaction.

We have studied the peroxidase-oxidase reaction catalyzed by human myeloperoxidase in an open system where both substrates-molecular oxygen and NADH-are supplied continuously to the reaction mixture. The reaction shows oscillatory kinetics at pH values around 5, provided that the reaction medium in addition to the enzyme and the substrates also contains an aromatic electron mediator such as melatonin or 4-hydroxybenzoic acid and chloride ions at concentrations >1mM. The experimental findings can be simulated by a detailed model of the reaction.

Mechanism of melatonin-induced oscillations in the peroxidase-oxidase reaction.

Melatonin induces oscillations in the peroxidase-oxidase (PO) reaction catalyzed by horseradish peroxidase. We present here studies of the effect of pH, enzyme concentration, and concentration of melatonin on the oscillation frequency. We also present a mechanistic model to explain the experimentally observed changes in oscillation frequency.