Viability and formation of conjugated dienes in plasma membrane lipids of Saccharomyces cerevisiae,Schizosaccharomyces pombe,Rhodotorula glutinis and Candida albicans exposed to hydrophilic,amphiphilic and hydrophobic pro-oxidants

Effects of four lipid peroxidation-inducing pro-oxidants-amphiphilictert-butyl hydroperoxide (TBHP), hydrophobic 1,1′-azobis(4-cyclohexanecarbonitrile) (ACHN), hydrophilic Fe¹¹ and 2,2′-azobis(2-amidinopropane) dihydrochloride (AAPH)-on cell growth and on generation of peroxidation products in isolated plasma membrane lipids were determined in four yeast species (S. cerevisiae, S. pombe, R. glutinis andC. albicans) differing in their plasma membrane lipid composition. TBHP and ACHN inhibited cell growth most strongly, Fe¹¹ and AAPH exerted inhibitory action for about 2 h, with subsequent cell growth resumption.S. cerevisiae strain SP4 was doped during growth with unsaturated linoleic (18∶2) and linolenic (18∶3) acids to change its resistance to lipid peroxidation. Its plasma membranes then contained some 30% of these acids as compared with some 1.3% of 18∶2 acid found in undopedS. cerevisiae, while the content of (16∶1) and (18∶1) acids was lower than in undopedS. cerevisiae. The presence of linoleic and linolenic acids inS. cerevisiae cells lowered cell survival and increased the sensitivity to pro-oxidants. Peroxidationgenerated conjugated dienes (CD) were measured in pure TBHP- and ACHN-exposed fatty acids used as standards. The CD level depended on the extent of unsaturation and the pro-oxidant used. The TBHP-induced CD production in a mixture of oleic acid and its ester was somewhat lower than in free acid and ester alone. In lipids isolated from the yeast plasma membranes, the CD production was time-dependent and decreased after a 5–15-min pro-oxidant exposure. ACHN was less active than TBHP. The most oxidizable were lipids fromS. cerevisiae plasma membranes doped with linoleic and linolenic acids and fromC. albicans with indigenous linolenic acid.     

Kinetic properties of DM-nitrophen binding to calcium and magnesium

Caged-Ca(2+) compounds such as nitrophenyl-EGTA (NP-EGTA) and DM-nitrophen (DMn) are extremely useful in biological research, but their use in live cells is hampered by cytoplasmic [Mg(2+)]. We determined the properties of Ca(2+) release from NP-EGTA and DMn by using Oregon green BAPTA-5N to measure changes in [Ca(2+)] after ultraviolet flash photolysis in vitro, with or without Mg(2+) present. A large fraction (65%) of NP-EGTA, which has a negligible Mg(2+) affinity, uncages with a time constant of 10.3 ms, resulting in relatively slow increases in [Ca(2+)]. Uncaging of DMn is considerably faster, but DMn has a significant affinity for Mg(2+) to complicate the uncaging process. With experimentally determined values for the Ca(2+) and Mg(2+) binding/unbinding rates of DMn and NP-EGTA, we built a mathematical model to assess the utility of NP-EGTA and DMn in rapid Ca(2+)-uncaging experiments in the presence of Mg(2+). We discuss the advantages and disadvantages of using each compound under different conditions. To determine the kinetics of Ca(2+) binding to biologically relevant Ca(2+) buffers, such as Ca(2+)-binding proteins, the use of DMn is preferable even in the presence of Mg(2+).     

Chemical structure of wall teichoic acid isolated from Enterococcus faecium strain U0317

Wall teichoic acid (WTA) was isolated from Enterococcus faecium strain U0317 and structurally characterized using ¹H, ¹³C, and ³¹P NMR spectroscopy, including two-dimensional COSY, TOCSY, ROESY, HMQC, and HMBC experiments. Further compositional determination was undertaken using classical chemical methods and HF treatment followed by GLC and GLC–MS analyses. The repeating unit of WTA consisted of two residues of 2-acetamido-2-deoxy-d-galactose, glycerol (Gro), and phosphate, and has the structure shown below:→6)-α-d-GalpNAc-(1→3)-β-d-GalpNAc-(1→2)-Gro-(3→P→     

Hoc protein regulates the biological effects of T4 phage in mammals

We previously investigated the biological, non-antibacterial effects of bacteriophage T4 in mammals (binding to cancer cells in vitro and attenuating tumour growth and metastases in vivo); we selected the phage mutant HAP1 that was significantly more effective than T4. In this study we describe a non-sense mutation in the hoc gene that differentiates bacteriophage HAP1 and its parental strain T4. We found no substantial effects of the mutation on the mutant morphology, and its effects on electrophoretic mobility and hydrodynamic size were moderate. Only the high ionic strength of the environment resulted in a size difference of about 10 nm between T4 and HAP1. We compared the antimetastatic activity of the T2 phage, which does not express protein Hoc, with those of T4 and HAP1 (B16 melanoma lung colonies). We found that HAP1 and T2 decreased metastases with equal effect, more strongly than did T4. We also investigated concentrations of T4 and HAP1 in the murine blood, tumour (B16), spleen, liver, or muscle. We found that HAP1 was rapidly cleared from the organism, most probably by the liver. Although HAP1 was previously defined to bind cancer cells more effectively (than T4), its rapid elimination precluded its higher concentration in tumours. Maria Zembala and Janusz Boratynski contributed equally to this work.     

Estimation of bacteriological spoilage of pork cutlets by electronic nose

The utility of chemosensor array (EN) signals of head-space volatiles of aerobically stored pork cutlets as a non-invasive technique for monitoring their microbiological load was studied during storage at 4, 8 and 12 degrees C, respectively. The bacteriological quality of the meat samples was determined by standard total aerobic plate counts (TAPC) and colony count of selectively estimated Pseudomonas (PS) spp., the predominant aerobic spoilage bacteria. Statistical analysis of the electronic nose measurements were principal component analysis (PCA), and canonical discriminant analysis (CDA). Partial least squares (PLS) regression was used to model correlation between microbial loads and EN signal responses, the degree of bacteriological spoilage, independently of the temperature of the refrigerated storage. Sensor selection techniques were applied to reduce the dimensionality and more robust calibration models were computed by determining few individual sensors having the smallest cross correlations and highest correlations with the reference data. Correlations between the predicted and "real" values were given on cross-validated data from both data reduced models and for full calibrations using the 23 sensor elements. At the same time, sensorial quality of the raw cutlets was noted subjectively on faultiness of the odour and colour, and drip formation of the samples. These preliminary studies indicated that the electronic nose technique has a potential to detect bacteriological spoilage earlier or at the same time as olfactory quality deterioration.     

Preapoptotic chromatin changes induced by ultraviolet B irradiation in human erythroleukemia K562 cells

Exponentially growing human erythroleukemia K562 cells were permeabilized and the dose dependent decrease of DNA synthesis rate was measured after ultraviolet (UV B, 290 nm) irradiation. Cells were able to overcome 2 and 5 J/m2 UV doses, partial recovery was observed at 15 J/m2, while at high (25 J/m2) UV dose replicative DNA synthesis remained suppressed. K562 cells were subjected to synchronization prior to and after UV irradiation (24 J/m2) and 18 fractions were collected by centrifugal elutriation. Cell cycle analysis by flow cytometry did not show early apoptotic cells after UV irradiation. The gradual increase in DNA content typical for non-irradiated cells was contrasted by an early S phase block between 2.2 and 2.4 C-values after UV irradiation. Cell cycle dependent chromatin changes after ultraviolet irradiation were seen as a fine fibrillary network covering the mainly fibrous chromatin structures and incompletely folded primitive chromosomes. Based on observations after UV irradiation and on earlier results with cadmium treatment and gamma irradiation, we confirm that typical chromatin changes characteristic to genotoxic agents can be recognized and classified.     

Structure of the ovaries and oogenesis in Cixius nervosus (Cixiidae), Javesella pellucida and Conomelus anceps (Delphacidae) (Insecta, Hemiptera, Fulgoromorpha)

Ovary organization in representatives of two families of Fulgoromorpha, Cixiidae (Cixius nervosus) and Delphacidae (Javesella pellucida and Conomelus anceps), was examined by light and transmission electron microscopy. Ovaries of studied fulgoromorphans consist of telotrophic ovarioles. From apex to base individual ovarioles have four well defined regions: a terminal filament, tropharium (trophic chamber), vitellarium and pedicel (ovariolar stalk). Tropharia are not differentiated into distinct zones and consist of syncytial lobes containing multiple trophocyte nuclei embedded in a common cytoplasm. Lobes are radially arranged around a branched, cell-free trophic core. Early previtellogenic (arrested) oocytes and prefollicular cells are located at the base of the tropharium. The vitellarium houses linearly arranged developing oocytes each of which is connected to the trophic core by a broad nutritive cord. Each oocyte is surrounded by a single layer of follicular cells that become binucleate at the beginning of vitellogenesis.     

Physiological and Molecular Background of Maize Cold-Tolerance Enhancement with S-methylmethionine Salicylate

Low temperature is amongst the most influential abiotic stress factors, having deep impact on plant growth, yield and productivity. Studies on beneficial effects of certain biologically active substances, S-methylmethionine (SMM) and salicylic acid (SA) have provided a lot of valuable information regarding their role to counteract harmful effects of environmental stresses such as chilling. To obtain a more complex and stable defence compound with an extended range of stress-protective effect, the new derivative S-methylmethionine salicylate (MMS) was synthesised from the natural, biologically active substances SMM and SA. Since both original materials have complex stress-protective roles, the new compound was expected to combine the effects of original substances and to stabilise the unstable SMM in the new compound, thus providing an extended stress tolerance. Photosynthetic efficiency and accumulation of stress-related metabolites (polyamines and flavonoids) were measured in chilled and control plants, with and without MMS pretreatment, and expression changes of several genes involved in the cold stress response were analysed by quantitative real-time PCR (RT-qPCR) and a detailed microarray study. Our data show how the MMS combines the effect of SMM and SA on molecular level, causing numerous changes in the gene expression pattern and metabolite content. MMS gives rise to a better physiological condition, thus it could provide an alternative, environmental friendly way to enhance the plants defence mechanisms against stressors. As MMS is more stable than SMM, it promises easier, more long-lasting and more cost-effective usage in agriculture, with a complementing effect of SA.