Jožef Stefan Institute

About: Jožef Stefan Institute is a facility organization based out in Ljubljana, Slovenia. It is known for research contribution in the topics: Liquid crystal & Dielectric. The organization has 3828 authors who have published 12614 publications receiving 291025 citations.

Cysteine-scanning mutagenesis of an eukaryotic pore-forming toxin from sea anemone: topology in lipid membranes.

Abstract: Equinatoxin II is a cysteineless pore-forming protein from the sea anemone Actinia equina It readily creates pores in membranes containing sphingomyelin Its topology when bound in lipid membranes has been studied using cysteine-scanning mutagenesis At approximately every tenth residue, a cysteine was introduced Nineteen single cysteine mutants were produced in Escherichia coli and purified The accessibility of the thiol groups in lipid-embedded cysteine mutants was studied by reaction with biotin maleimide Most of the mutants were modified, except those with cysteines at positions 105 and 114 Mutants R144C and S160C were modified only at high concentrations of the probe Similar results were obtained if membrane-bound biotinylated mutants were tested for avidin binding, but in this case three more mutants gave a negative result: S1C, S13C and K43C Furthermore, mutants S1C, S13C, K20C, K43C and S95C reacted with biotin only after insertion into the lipid, suggesting that they were involved in major conformational changes occurring upon membrane binding These results were further confirmed by labeling the mutants with acrylodan, a polarity-sensitive fluorescent probe When labeled mutants were combined with vesicles, the following mutants exhibited blue-shifts, indicating the transfer of acrylodan into a hydrophobic environment: S13C, K20C, S105C, S114C, R120C, R144C and S160C The overall results suggest that at least two regions are embedded within the lipid membrane: the N-terminal 13‐20 region, probably forming an amphiphilic helix, and the tryptophan-rich 105‐120 region Arg144, Ser160 and residues nearby could be involved in making contacts with lipid headgroups The association with the membrane appears to be unique and different from that of bacterial pore-forming proteins and therefore equinatoxin II may serve as a model for eukaryotic channel-forming toxins

Lysosomal proteases cathepsins d, b, h, l and their inhibitors stefins a and b in head and neck cancer

Abstract: In cytosols of tumour and normal tissue of 53 patients suffering from head and neck carcinoma cathepsins D, B, H and L were measured using quantitative immunoreactive assays (ELISA). The values of cathepsins D, B and L were significantly higher in tumour tissue, whereas cathepsin H concentration was lower in tumour than in normal tissue. Median cathepsin D values were 27 pmol (tumour tissue) vs. 12 pmol (normal tissue) per mg of total protein, median cathepsin B values were 1.25 micrograms/mg (tumour tissue) vs. 0.23 micrograms/mg (normal tissue) and median cathepsin L values were 39.8 ng/mg (tumour tissue) vs. 20.0 ng/mg (normal tissue). Median cathepsin H values were 1.05 micrograms/mg and 2.20 micrograms/mg for tumour and normal tissue, respectively. Additionally, stefin A and stefin B were measured in tumour and normal tissue samples. In contrast to the cathepsins, the concentrations of these inhibitors of cysteine proteinases was not significantly different between tumour and normal samples. The concentrations of cathepsins D, B, H and L and stefins A and B measured in head and neck tumours, were independent of standard clinical and histological prognostic factors. Significant correlation of tumour tissue values was observed between cathepsins B and L and between both stefins.

Multiplex quantification of 12 European Union authorized genetically modified maize lines with droplet digital polymerase chain reaction.

Abstract: Presence of genetically modified organisms (GMO) in food and feed products is regulated in many countries. The European Union (EU) has implemented a threshold for labeling of products containing more than 0.9% of authorized GMOs per ingredient. As the number of GMOs has increased over time, standard-curve based simplex quantitative polymerase chain reaction (qPCR) analyses are no longer sufficiently cost-effective, despite widespread use of initial PCR based screenings. Newly developed GMO detection methods, also multiplex methods, are mostly focused on screening and detection but not quantification. On the basis of droplet digital PCR (ddPCR) technology, multiplex assays for quantification of all 12 EU authorized GM maize lines (per April first 2015) were developed. Because of high sequence similarity of some of the 12 GM targets, two separate multiplex assays were needed. In both assays (4-plex and 10-plex), the transgenes were labeled with one fluorescence reporter and the endogene with another (GMO co...

Hypoxia-Induced Oxidative Stress Modulation with Physical Activity

Abstract: Increased oxidative stress, defined as an imbalance between prooxidants and antioxidants, resulting in molecular damage and disruption of redox signaling, is associated with numerous pathophysiological processes and known to exacerbate chronic diseases. Prolonged systemic hypoxia, induced either by exposure to terrestrial altitude or a reduction in ambient O2 availability is known to elicit oxidative stress and thereby alter redox balance in healthy humans. The redox balance modulation is also highly dependent on the level of physical activity. For example, both high-intensity exercise and inactivity, representing the two ends of the physical activity spectrum, are known to promote oxidative stress. Numerous to-date studies indicate that hypoxia and exercise can exert additive influence upon redox balance alterations. However, recent evidence suggests that moderate physical activity can attenuate altitude/hypoxia-induced oxidative stress during long-term hypoxic exposure. The purpose of this review is to summarize recent findings on hypoxia-related oxidative stress modulation by different activity levels during prolonged hypoxic exposures and examine the potential mechanisms underlying the observed redox balance changes. The paper also explores the applicability of moderate activity as a strategy for attenuating hypoxia-related oxidative stress. Moreover, the potential of such moderate intensity activities used to counteract inactivity-related oxidative stress, often encountered in pathological, elderly and obese populations is also discussed. Finally, future research directions for investigating interactive effects of altitude/hypoxia and exercise on oxidative stress are proposed.