Selective activators of protein phosphatase 5 target the auto-inhibitory mechanism

Protein phosphatase 5 (PP5) is an evolutionary conserved serine/threonine phosphatase. Its dephosphorylation activity modulates a diverse set of cellular factors including protein kinases and the microtubule-associated tau protein involved in neurodegenerative disorders. It is auto-regulated by its heat-shock protein (Hsp90)-interacting tetratricopeptide repeat (TPR) domain and its C-terminal α-helix. In the present study, we report the identification of five specific PP5 activators [PP5 small-molecule activators (P5SAs)] that enhance the phosphatase activity up to 8-fold. The compounds are allosteric modulators accelerating efficiently the turnover rate of PP5, but do barely affect substrate binding or the interaction between PP5 and the chaperone Hsp90. Enzymatic studies imply that the compounds bind to the phosphatase domain of PP5. For the most promising compound crystallographic comparisons of the apo PP5 and the PP5–P5SA-2 complex indicate a relaxation of the auto-inhibited state of PP5. Residual electron density and mutation analyses in PP5 suggest activator binding to a pocket in the phosphatase/TPR domain interface, which may exert regulatory functions. These compounds thus may expose regulatory mechanisms in the PP5 enzyme and serve to develop optimized activators based on these scaffolds.     

Isavuconazole and Liposomal Amphotericin B as Successful Combination Therapy of Refractory Invasive Candidiasis in a Liver Transplant Recipient: A Case Report and Literature Review

Invasive fungal infections in liver transplant recipients are associated with elevated morbidity and mortality and pose a challenge to the treating physicians. Despite of lacking clinical data, the use of antifungal combination therapy is often considered to improve response rates in an immunocompromised patient population. We herein report a case of refractory invasive candidiasis in a liver transplant recipient treated successfully with a combination of isavuconazole und high-dose liposomal amphotericin B. The antimycotic combination treatment was able to clear a bloodstream infection with C. glabrata and led to regression of bilomas among tolerable side effects. The use of the above-mentioned antifungal combination therapy in a liver transplant recipient has not been reported previously. This case highlights the efficacy and safety of antifungal combination therapy in immunocompromised patients with refractory invasive candidiasis.

     

Levels of heavy metals and their binding protein metallothionein in type 2 diabetics with kidney disease

Hyperglycemia, a major metabolic disturbance present in diabetes, promotes oxidative stress. Activation of antioxidant defense is an important mechanism to prevent cell damage. Levels of heavy metals and their binding proteins can contribute to oxidative stress. Antiradical capacity and levels of metallothionein (MT), metals (zinc and copper), and selected antioxidants (bilirubin, cysteine, and glutathione) were determined in 70 type 2 diabetes mellitus (T2DM) subjects and 80 healthy subjects of Caucasian origin. Single nucleotide polymorphism (rs28366003) in MT gene was detected. Antiradical capacity, conjugated bilirubin, and copper were significantly increased in diabetics, whereas MT and glutathione were decreased. Genotype AA of rs28366003 was associated with higher zinc levels in the diabetic group. The studied parameters were not influenced by renal function. This is the first study comprehensively investigating differences in MT and metals relevant to oxidative stress in T2DM. Ascertained differences indicate increased oxidative stress in T2DM accompanied by abnormalities in non‐enzymatic antioxidant defense systems.     

Development of Orodispersible Films Containing Benzydamine Hydrochloride Using a Modified Solvent Casting Method

The aim of this study was to develop benzydamine hydrochloride-loaded orodispersible films using the modification of a solvent casting method. An innovative approach was developed when the drying process of a small-scale production was used based on a heated inert base for casting the film. During this process, two types of film-forming maltodextrins for rapid drug delivery were used. They were plasticized with two different polyols (xylitol and sorbitol). Superdisintegrant Kollidon® CL-F was tested as an excipient that can induce faster disintegration of the prepared films. The influence of the formulation parameters (dextrose equivalent of film-forming maltodextrins, a type of plasticizer, and the presence of superdisintegrant) on the disintegration time, mechanical properties, and moisture content of films was statistically evaluated using a multivariate data analysis. Orodispersible films containing maltodextrin with lower dextrose equivalent value showed better mechanical properties (tensile strength ranged from 886.6?±?30.2 to 1484.2?±?226.9 N cm^?2), lower moisture content (0.5?±?0.0 to 1.2?±?0.2%), and shorter disintegration time (17.6?±?2.9 to 27.8?±?2.8 s). Films plasticized with xylitol showed shorter disintegration time (17.6?±?2.9 to 29.2?±?3.8 s) than films containing sorbitol (23.8?±?2.9 to 31.7?±?3.9 s). With the addition of superdisintegrant Kollidon® CL-F, a significant influence on disintegration time was not observed. The modified solvent casting method shows great promise in a small-scale laboratory production of orodispersible films, e.g., in a pharmacy lab.     

Study on metal-triggered callose deposition in roots of maize and soybean

Callose plays important roles in a variety of processes of plant development, and/or in a response to a range of biotic and abiotic stresses. In the current work we have studied and compared the effect of lead, cadmium and arsenic on accumulation of newly formed callose deposits in the roots of maize and soybean. We observed formation of characteristic callose deposits in the root cell walls, probably associated with plasmodesmata, depending on the type of metal and the plant species investigated. Further, the callose turnover was analysed by measuring of total callose content as well as activities of total β-(1,3)-glucanases in roots. The latter enzymes are responsible for callose depletion, and their possible role during metal stress has previously been proposed. However, neither of these biochemical values appeared to be sufficiently reliable for scoring the altered callose turnover (including local deposits) in plant tissue. The microscopical observations are discussed in light of the biochemical data obtained.     

Cysteine cathepsins: from structure, function and regulation to new frontiers

It is more than 50 years since the lysosome was discovered. Since then its hydrolytic machinery, including proteases and other hydrolases, has been fairly well identified and characterized. Among these are the cysteine cathepsins, members of the family of papain-like cysteine proteases. They have unique reactive-site properties and an uneven tissue-specific expression pattern. In living organisms their activity is a delicate balance of expression, targeting, zymogen activation, inhibition by protein inhibitors and degradation. The specificity of their substrate binding sites, small-molecule inhibitor repertoire and crystal structures are providing new tools for research and development. Their unique reactive-site properties have made it possible to confine the targets simply by the use of appropriate reactive groups. The epoxysuccinyls still dominate the field, but now nitriles seem to be the most appropriate "warhead". The view of cysteine cathepsins as lysosomal proteases is changing as there is now clear evidence of their localization in other cellular compartments. Besides being involved in protein turnover, they build an important part of the endosomal antigen presentation. Together with the growing number of non-endosomal roles of cysteine cathepsins is growing also the knowledge of their involvement in diseases such as cancer and rheumatoid arthritis, among others. Finally, cysteine cathepsins are important regulators and signaling molecules of an unimaginable number of biological processes. The current challenge is to identify their endogenous substrates, in order to gain an insight into the mechanisms of substrate degradation and processing. In this review, some of the remarkable advances that have taken place in the past decade are presented. This article is part of a Special Issue entitled: Proteolysis 50 years after the discovery of lysosome.     

The Hemagglutinin of Canine Distemper Virus Determines Tropism and Cytopathogenicity

Canine distemper virus (CDV) and measles virus (MV) cause severe illnesses in their respective hosts. The viruses display a characteristic cytopathic effect by forming syncytia in susceptible cells. For CDV, the proficiency of syncytium formation varies among different strains and correlates with the degree of viral attenuation. In this study, we examined the determinants for the differential fusogenicity of the wild-type CDV isolate 5804Han89 (CDV(5804)), the small- and large-plaque-forming variants of the CDV vaccine strain Onderstepoort (CDV(OS) and CDV(OL), respectively), and the MV vaccine strain Edmonston B (MV(Edm)). The cotransfection of different combinations of fusion (F) and hemagglutinin (H) genes in Vero cells indicated that the H protein is the main determinant of fusion efficiency. To verify the significance of this observation in the viral context, a reverse genetic system to generate recombinant CDVs was established. This system is based on a plasmid containing the full-length antigenomic sequence of CDV(OS). The coding regions of the H proteins of all CDV strains and MV(Edm) were introduced into the CDV and MV genetic backgrounds, and recombinant viruses rCDV-H(5804), rCDV-H(OL), rCDV-H(Edm), rMV-H(5804), rMV-H(OL), and rMV-H(OS) were recovered. Thus, the H proteins of the two morbilliviruses are interchangeable and fully functional in a heterologous complex. This is in contrast with the glycoproteins of other members of the family Paramyxoviridae, which do not function efficiently with heterologous partners. The fusogenicity, growth characteristics, and tropism of the recombinant viruses were examined and compared with those of the parental strains. All these characteristics were found to be predominantly mediated by the H protein regardless of the viral backbone used.     

Same Same but Different. Different Trigeminal Chemoreceptors Share the Same Central Pathway

Intranasal trigeminal sensations are important in everyday life of human beings, as they play a governing role in protecting the airways from harm. Trigeminal sensations arise from the binding of a ligand to various sub-types of transient receptor potential (TRP) channels located on mucosal branches of the trigeminal nerve. Which underlying neural networks are involved in the processing of various trigeminal inputs is still unknown. To target this unresolved question fourteen healthy human subjects were investigated by completing three functional magnetic resonance imaging (fMRI) scanning sessions during which three trigeminal substances, activating varying sub-types of chemoreceptors and evoking different sensations in the nose were presented: CO₂, menthol and cinnamaldehyde. We identified similar functional networks responding to all stimuli: an olfactory network, a somatosensory network and an integrative network. The processing pathway of all three stimulants was represented by the same functional networks, although CO₂ evokes painful but virtually odorless sensations, and the two other stimulants, menthol and cinnamaldehyde are perceived as mostly non painful with a clear olfactory percept. Therefore, our results suggest a common central processing pathway for trigeminal information regardless of the trigeminal chemoreceptor and sensation type.     

Constraints on the biological recovery of the Bohemian Forest lakes from acid stress

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Carbon footprint of rice production under biochar amendment – a case study in a Chinese rice cropping system

As a controversial strategy to mitigate global warming, biochar application into soil highlights the need for life cycle assessment before large‐scale practice. This study focused on the effect of biochar on carbon footprint of rice production. A field experiment was performed with three treatments: no residue amendment (Control), 6 t ha^?1 yr^?1 corn straw (CS) amendment, and 2.4 t ha^?1 yr^?1 corn straw‐derived biochar amendment (CBC). Carbon footprint was calculated by considering carbon source processes (pyrolysis energy cost, fertilizer and pesticide input, farmwork, and soil greenhouse gas emissions) and carbon sink processes (soil carbon increment and energy offset from pyrolytic gas). On average over three consecutive rice‐growing cycles from year 2011 to 2013, the CS treatment had a much higher carbon intensity of rice (0.68 kg CO₂‐C equivalent (CO₂‐C_e) kg^?1 grain) than that of Control (0.24 kg CO₂‐C_e kg^?1 grain), resulting from large soil CH₄ emissions. Biochar amendment significantly increased soil carbon pool and showed no significant effect on soil total N₂O and CH₄ emissions relative to Control; however, due to a variation in net electric energy input of biochar production based on different pyrolysis settings, carbon intensity of rice under CBC treatment ranged from 0.04 to 0.44 kg CO₂‐C_e kg^?1 grain. The results indicated that biochar strategy had the potential to significantly reduce the carbon footprint of crop production, but the energy‐efficient pyrolysis technique does matter.