Screening of synthetic phage display scFv libraries yields competitive ligands of human leptin receptor

Initially considered the main endogenous anorexigenic factor, fat-derived leptin turned out to be a markedly pleiotropic hormone, influencing diverse physiological processes. Moreover, hyperleptinemia in obese individuals has been linked to the onset or progression of serious disorders, such as cancer, autoimmune diseases, and atherosclerosis, and antagonizing peripheral leptin’s signalization has been shown to improve these conditions. To develop an antibody-based leptin antagonist we have devised a tailored panning procedure and screened two phage display libraries of single chain variable antibody fragments (scFvs) against recombinant leptin receptor. One of the scFvs was expressed in Escherichia coli and its interaction with leptin receptor was characterized in more detail. It was found to recognize a discontinuous epitope and to compete with leptin for receptor binding with IC50 and K_d values in the nanomolar range. The reported scFv represents a lead for development of leptin antagonists that may ultimately find use in therapy of various hyperleptinemia-related disorders.     

Life-cycle assessment of a hydrogen-based uninterruptible power supply system using renewable energy

Purpose

The paper provides an empirical assessment of an uninterruptible power supply (UPS) system based on hydrogen technologies (HT-UPS) using renewable energy sources (RES) with regard to its environmental impacts and a comparison to a UPS system based on the internal combustion engine (ICE-UPS).

Methods

For the assessment and comparison of the environmental impacts, the life-cycle assessment (LCA) method was applied, while numerical models for individual components of the UPS systems (electrolyser, storage tank, fuel cell and ICE) were developed using GaBi software. The scope of analysis was cradle-to-end of utilisation with functional unit 1 kWh of uninterrupted electricity produced. For the life-cycle inventory analysis, quantitative data was collected with on-site measurements on an experimental system, project documentation, GaBi software generic databases and literature data. The CML 2001 method was applied to evaluate the system’s environmental impacts. Energy consumption of the manufacturing phase was estimated from gross value added (GVA) and the energy intensity of the industry sector in the manufacturer’s country.

Results and discussion

In terms of global warming (GW), acidification (A), abiotic depletion (AD) and eutrophication (E), manufacturing phase of HT-UPS accounts for more than 97 % of environmental impacts. Electrolyser in all its life-cycle phases contributes above 50 % of environmental impacts to the system’s GW, A and AD. Energy return on investment (EROI) for the HT-UPS has been calculated to be 0.143 with distinction between renewable (roughly 60 %) and non-renewable energy resources inputs. HT-UPS’s life-cycle GW emissions have been calculated to be 375 g of CO₂ eq per 1 kWh of uninterruptible electric energy supplied. All these values have also been calculated for the ICE-UPS and show that in terms of GW, A and AD, the ICE-UPS has bigger environmental impacts and emits 1,190 g of CO₂ eq per 1 kWh of uninterruptible electric energy supplied. Both systems have similar operation phase energy efficiency. The ICE-UPS has a higher EROI but uses almost none RES inputs.

Conclusions

The comparison of two different technologies for providing UPS has shown that in all environmental impact categories, except eutrophication, the HT-UPS is the sounder system. Most of HT-UPS’s environmental impacts result from the manufacturing phase. On the contrary, ICE-UPS system’s environmental impacts mainly result from operational phase. Efficiency of energy conversion from electricity to hydrogen to electricity again is rather low, as is EROI, but these will likely improve as the technology matures.     

Comparing patterns of human harvest and predation by Eurasian lynx Lynx lynx on European roe deer Capreolus capreolus in a temperate forest

Human harvest is the most important mortality factor for wild ungulates in Europe and can affect several aspects of ungulate biology. There is a growing concern about possible negative side effects of human harvest. To better understand the differences between human and natural mortality, we compared the extent, age and sex structure, nutritional condition, spatial and temporal distribution of human harvest, and natural predation by the Eurasian lynx Lynx lynx on the European roe deer Capreolus capreolus, the most abundant wild ungulate in Europe. Compared to the human harvest, lynx were less likely to kill fawns and yearlings than adults, and among adult deer, lynx were more likely to kill females. The proportion of roe deer with fat-depleted bone marrow was higher among lynx prey than among harvested animals. Average lynx kill rate was estimated to 47.8 roe deer per year, and lynx predation was considerably lower than the human harvest in the same area. While human harvest increased with higher roe deer density, lynx predation was similar across the gradient of roe deer densities. Comparison with other countries indicated that differences between human harvest and natural mortality of ungulates vary considerably in different parts of Europe. Variation in hunting practices and, even more importantly, carnivore predation may have an important role in buffering unwanted side effects of harvest of wild ungulates.     

Population densities and habitat use of the golden jackal (Canis aureus) in farmlands across the Balkan Peninsula

The main objective of this study was to analyze the habitat use and population densities of the golden jackal in four countries across lowland regions of the Balkan Peninsula, known as the core area of the species' distribution in Europe. Using indirect (acoustic) method for detecting territorial golden jackals, we surveyed jackal presence and densities on 331 monitoring sites in four countries, covering area an of 4,296 km² in total during April and May 2007–2012. We used GIS to assess landscape and environmental characteristics in a 2-km circular buffer (12.6 km²) around calling stations. Average population density of golden jackals in the study areas ranged between 0.6 and 1.1 territorial groups/10 km² (mean ± SE, 0.6?±?0.06 groups/10 km²), with several high-density areas with up to 4.8 territorial groups/10 km². Analysis of habitat use showed that for both jackal occurrence and number of jackal groups, the only significant parameter was the interaction between country and intensity of agriculture, indicating that jackals adapt their habitat selection patterns in relation to the habitat availability. We observed that selection of the more suitable habitats (shrub–herbaceous vegetation/heterogeneous agricultural vegetation) increased with lower proportion of these habitat types in the study area. Our study confirms high habitat plasticity of the golden jackal and offers explanation for its recent range expansion, which might be connected with the land use changes during the last decades in the Balkan Peninsula.     

Cobalt(II) coordination compounds with acetate and 2-aminopyridine ligands: Synthesis, characterization, structures and magnetic properties of two polymorphic forms

The synthesis and characterization of two polymorphic modifications of new cobalt coordination compound with 2-aminopyridine are reported. The modifications were prepared by the reaction of a solution of cobalt acetate tetrahydrate and 2-aminopyridine. The crystal structures of both polymorphic modifications have been determined by single-crystal X-ray diffraction analysis. The structures of both modifications are quite similar. In both of them the Co²⁺ is six coordinated by four O atoms from two bidentate chelate acetate ligands and by two N atoms from two 2-aminopyridine molecules. Acetate and 2-aminopyridine ligands are lying cis about the metal centre. The most important difference is in asymmetry of acetate ligand chelate bonding and in H-bonding network. Compounds exhibit an extensive system of intra and intermolecular hydrogen bonding. Magnetic properties of both modifications were studied between 2 K and 300 K giving the result μ_eff = 4.6 BM for modification I and μ_eff = 4.7 BM for modification II in paramagnetic region.     

Flavonoids and cinnamic acid esters as inhibitors of fungal 17beta-hydroxysteroid dehydrogenase: a synthesis, QSAR and modelling study

The 17beta-hydroxysteroid dehydrogenases (17beta-HSDs) modulate the biological potency of estrogens and androgens by interconversion of inactive 17-keto-steroids and their active 17beta-hydroxy- counterparts. We have shown previously that flavonoids are potentially useful lead compounds for developing inhibitors of 17beta-HSDs. In this paper, we describe the synthesis and biochemical evaluation of structurally analogous inhibitors, the trans-cinnamic acid esters and related compounds. Additionally, quantitative structure-activity relationship (QSAR) and modelling studies were performed to rationalize the results and to suggest further optimization. The results stress the importance of a hydrogen bond with Asn154 and hydrophobic interactions with the aromatic side chain of Tyr212 for optimal molecular recognition.     

HWGMWSY, an unanticipated polystyrene binding peptide from random phage display libraries

Phage display is a powerful technique for the discovery of peptide ligands that bind to various targets; however, ambiguous results often appear. Peptide HWGMWSY has been isolated repeatedly in our laboratory and by other research groups dealing with different protein and nonprotein targets, which led to a hypothesis that it may be a target-unrelated peptide interacting with polystyrene plastic surfaces. We compared binding properties and amplification rate of phage clone displaying the peptide HWGMWSY, a previously confirmed plastic binding clone WHWRLPS, and a control phage clone ASVQERK. An enzyme-linked immunosorbent assay and a phage elution assay confirmed that phage clone HWGMWSY binds to polystyrene. Surface plasmon resonance measurements on the other hand excluded the possibility of binding to bovine serum albumin, a common blocking agent in phage display experiments. Amplification rates of the above-noted phage clones were not statistically different. We therefore conclude that phage clone HWGMWSY was isolated in different selection procedures as a result of its affinity to polystyrene.     

Range and local population densities of brown bear Ursus arctos in Slovenia

Solid understanding of species’ range and local population densities is important for successful wildlife management and research. Specific behavioral and ecological characteristics make brown bear Ursus arctos a difficult species to study. We present a map of range and local population densities of brown bears in Slovenia, made with the use of a new approach similar to voting classifications based on a combination of four datasets: Global Positioning System telemetry data, records of bear removals, systematic and opportunistic direct observations and signs of bear presence, and noninvasive genetic samples. Results indicate that the majority of bears in Slovenia live in Dinaric Mountains in the southern part of the country where local bear population densities exceed 40 bears/100 km². This is one of the highest population densities reported so far for this species worldwide. Population densities decrease towards the north (Alpine region) and are very low along the border with Italy and Austria where almost no females are present. This explains slow past and present expansion of this transboundary bear population into the Alps and should be considered in future bear re-colonization management strategies. Results also showed that data from observations and removals overestimate bear population densities at low values, while mortality and genetic data overestimate population densities in areas with more people. Nevertheless, all data types appeared useful for describing the general bear distribution patterns. Similar approach could be applied to studies of other charismatic or game species, for which several types of data are often available.     

Neural regulation of acetylcholinesterase-associated collagen Q in rat skeletal muscles

Acetylcholinesterase-associated collagen Q is expressed also outside of neuromuscular junctions in the slow soleus muscle, but not in fast muscles. We examined the nerve dependence of muscle collagen Q expression and mechanisms responsible for these differences. Denervation decreased extrajunctional collagen Q mRNA levels in the soleus muscles and junctional levels in fast sternomastoid muscles to about one third. Cross-innervation of denervated soleus muscles by a fast muscle nerve, or electrical stimulation by 'fast' impulse pattern, reduced their extrajunctional collagen Q mRNA levels by 70–80%. In contrast, stimulation of fast muscles by 'slow' impulse pattern had no effect on collagen Q expression. Calcineurin inhibitors tacrolimus and cyclosporin A decreased collagen Q mRNA levels in the soleus muscles to about 35%, but did not affect collagen Q expression in denervated soleus muscles or the junctional expression in fast muscles. Therefore, high extrajunctional expression of collagen Q in the soleus muscle is maintained by its tonic nerve-induced activation pattern via the activated Ca²⁺-calcineurin signaling pathway. The extrajunctional collagen Q expression in fast muscles is independent of muscle activation pattern and seems irreversibly suppressed. The junctional expression of collagen Q in fast muscles is partly nerve-dependent, but does not encompass the Ca²⁺-calcineurin signaling pathway.     

Experience obtained from box trapping and handling wildcats in Slovenia

WildcatsFelis silvestris Schreber, 1775 were captured for radio collaring as a part of a study of their spatial distribution and social organisation in southern Slovenia between 1999 and 2001. Double-door box traps, with a roof that bears easily break (bear permeable traps), have been used to capture individuals. The distances between traps were between 550 to 2200 m. They were set out on logging roads and narrow trails in the forest. Nine wildcats, one lynx Lynx lynx (Linnaeus, 1758) and one feral cat were caught as target species and 19 badgersMeles meles and one bear cubUrsus arctos as non-targets. The catching success was 1 wildcat/58 trap-days and seems to be in correlation with the lunar cycle. Overall, 7 males and 2 female wildcats were captured which might indicate sex biased trapping selection. Methodological improvements shortened the time of handling procedures. Improved field protocols as well as restraining and immobilisation procedures increased reliability and safety of drug administrations, decreased potential chances for injuries and reduced overall stress of captured animals.