Amyloid fibril formation by human stefin B in vitro: immunogold labelling and comparison to stefin A

The mechanism by which proteins form amyloid fibrils is of high interest to the scientific community as its understanding could resolve questions relevant to conformational diseases. The structural and energetic basis of the process is still largely unknown. The main controversial issue is the co-existence of several protein conformations. Three models for the mechanism of protein fibrillogenesis have been proposed which need to be tested by experiments. In this report, amyloid fibrils grown from human stefin B (type I cystatin) are described. This physiologically relevant protein readily forms fibrils in vitro, in contrast to the homologue--human stefin A--which forms fibrils under extreme conditions only. In order to specifically label stefin B fibrils in vitro, rabbit polyclonal antibody and mouse monoclonal antibody A6/2 against human stefin B were used for immunogold labelling. Samples were examined by transmission electron microscopy. Fibrils of stefin B were strongly labelled using polyclonal antibody and Protein A gold, whereas no positive reaction was observed with monoclonal antibody A6/2.     

A possible origin of the torocyte-like shape of vesicles derived from the transverse tubule in the triad junction of skeletal muscle

The origin of characteristic torocyte-like shape of vesicles derived from transverse tubule in triad junction of skeletal muscles is studied theoretically. Two possible mechanisms are suggested. The first is the minimization of membrane bending energy where the special intermediate molecular structures in the central region of the vesicle is assumed to protect the opposing bilayers to come in the direct contact. The second mechanism is based on the assumption that the characteristic shape of the vesicles may be explained by non-homogenous lateral distribution of anisotropic membrane components.     

Evolutionary and ecological traps for brown bears Ursus arctos in human‐modified landscapes

相似文献   

Ostreopexin: A hemopexin fold protein from the oyster mushroom, Pleurotus ostreatus

Proteins with hemopexin repeats are widespread in viruses, prokaryotes and eukaryotes. We report here for the first time the existence of a protein in fungi with the four-bladed β-propeller fold that is typical for hemopexin-like proteins. This protein was isolated from the edible basidiomycetous fungus Pleurotus ostreatus and is named ostreopexin. It binds to Ni^2 +-NTA-agarose, and is structurally and functionally very similar to PA2 albumins isolated from legume seeds and the hemopexin fold protein from rice. Like these plant proteins, ostreopexin shows reversible binding to hemin with moderate affinity, but does not bind to polyamines. We suggest that ostreopexin participates in intracellular management of metal (II or III)-chelates.     

Use of GPS location clusters analysis to study predation, feeding, and maternal behavior of the Eurasian lynx

Analysis of global positioning system (GPS) location clusters (GLCs) is becoming increasingly popular in studies of carnivore ecology. While promising, this application of GPS technology is still poorly developed for most species. We applied this method to study predation and maternal behavior of the Eurasian lynx (Lynx lynx) in the Dinaric Mountains. Low population densities, rugged terrain, dense vegetation, and administrative borders make studies of this endangered population using traditional methods and a limited budget very challenging. We used the geographic information system and linear mixed-effects models to understand the movement of lynx during the consumption process and denning period and estimate lynx kill rates. A total of 99 % of kills were found at GLCs longer than 30 h and with minimum two locations within 300 m. We confirmed 86 % of potential kills and all potential dens that were searched for in the field. High success in predicting kill and den sites showed that the Eurasian lynx is a suitable species for application of the GLC analysis methods. Comparison of field-confirmed kills with model predictions showed the possibility for remote estimation of approximate kill rates in Eurasian lynx. Movements of the lynx were primarily affected by daytime period, time since the last kill/den translocation, lynx category, and their interactions. Based on the empirical data, we programmed simulations of lynx movements and elaborated recommendations for more efficient field procedures and study designs (GPS schedules) for future studies. We believe that our findings and approach will also benefit studies of other species with similar behavior.     

In vivo characterization of thermal stabilities of Aeropyrum pernix cellular components by differential scanning calorimetry

Revival studies of Aeropyrum pernix show that the viability of cells and cell recovery after heat treatment depends on the temperature of treatment. Differential scanning calorimetry (DSC) is used to analyze the relative thermal stabilities of cellular components of A. pernix and to identify the cellular components responsible for the observed lag phase and reduced maximum growth following a heat treatment. DSC thermograms show 5 visible endothermic transitions with 2 major transitions. DSC analysis of isolated crude ribosomes aids the assignment of the 2 major peaks observed in whole-cell thermograms to denaturation of ribosomal structures. A comparison of partial and immediate full rescan thermograms of A. pernix whole cells indicates that both major peaks represent irreversible thermal transitions. A DNA peak is also identified in the whole-cell thermogram by comparison with the optical data of isolated pure DNA. DNA melting is shown to be irreversible in dilute solution, whereas it is partially reversible in whole cells, owing at least in part, to restricted volume effects. In contrast to mesophilic organisms, hyperthermophilic A. pernix ribosomes are more thermally stable than DNA, but in both organisms, irreversible changes leading to cell death occur owing to ribosomal denaturation.     

Fungal β-trefoil trypsin inhibitors cnispin and cospin demonstrate the plasticity of the β-trefoil fold

The recently identified fungal protease inhibitors cnispin, from Clitocybe nebularis, and cospin, from Coprinopsis cinerea, are both β-trefoil proteins highly specific for trypsin. The reactive site residue of cospin, Arg27, is located on the β2–β3 loop. We show here, that the reactive site residue in cnispin is Lys127, located on the β11–β12 loop. Cnispin is a substrate-like inhibitor and the β11–β12 loop is yet another β-trefoil fold loop recruited for serine protease inhibition. By site-directed mutagenesis of the P1 residues in the β2–β3 and β11–β12 loops in cospin and cnispin, protease inhibitors with different specificities for trypsin and chymotrypsin inhibition have been engineered. Double headed inhibitors of trypsin or trypsin and chymotrypsin were prepared by introducing a second specific site residue into the β2–β3 loop in cnispin and into the β11–β12 loop in cospin. These results show that β-trefoil protease inhibitors from mushrooms exhibit broad plasticity of loop utilization in protease inhibition.     

Empirical valence bond simulations of the hydride transfer step in the monoamine oxidase B catalyzed metabolism of dopamine

Monoamine oxidases (MAOs) A and B are flavoenzymes responsible for the metabolism of biogenic amines such as dopamine, serotonin and noradrenaline. In this work, we present a comprehensive study of the rate‐limiting step of dopamine degradation by MAO B, which consists in the hydride transfer from the methylene group of the substrate to the flavin moiety of the FAD prosthetic group. This article builds on our previous quantum chemical study of the same reaction using a cluster model (Vianello et al., Eur J Org Chem 2012; 7057), but now considering the full dimensionality of the hydrated enzyme with extensive configurational sampling. We show that MAO B is specifically tuned to catalyze the hydride transfer step from the substrate to the flavin moiety of the FAD prosthetic group and that it lowers the activation barrier by 12.3 kcal mol^?1 compared to the same reaction in aqueous solution, a rate enhancement of more than nine orders of magnitude. Taking into account the deprotonation of the substrate prior to the hydride transfer reaction, the activation barrier in the enzyme is calculated to be 16.1 kcal mol^?1, in excellent agreement with the experimental value of 16.5 kcal mol^?1. Additionally, we demonstrate that the protonation state of the active site residue Lys296 does not have an influence on the hydride transfer reaction. Proteins 2014; 82:3347–3355. © 2014 Wiley Periodicals, Inc.     

Extraction of Temporal Networks from Term Co-Occurrences in Online Textual Sources

A stream of unstructured news can be a valuable source of hidden relations between different entities, such as financial institutions, countries, or persons. We present an approach to continuously collect online news, recognize relevant entities in them, and extract time-varying networks. The nodes of the network are the entities, and the links are their co-occurrences. We present a method to estimate the significance of co-occurrences, and a benchmark model against which their robustness is evaluated. The approach is applied to a large set of financial news, collected over a period of two years. The entities we consider are 50 countries which issue sovereign bonds, and which are insured by Credit Default Swaps (CDS) in turn. We compare the country co-occurrence networks to the CDS networks constructed from the correlations between the CDS. The results show relatively small, but significant overlap between the networks extracted from the news and those from the CDS correlations.     

FragViz: visualization of fragmented networks

Background  

Researchers in systems biology use network visualization to summarize the results of their analysis. Such networks often include unconnected components, which popular network alignment algorithms place arbitrarily with respect to the rest of the network. This can lead to misinterpretations due to the proximity of otherwise unrelated elements.