Radical Intermediates in the Redox Reactions of Tetrazolium Salts in Aprotic Solvents (Cyclovoltammetric, EPR and UV-VIS study)

Tetrazolium Blue (TBCl₂) and Nitrotetrazolium Blue (NTBCl₂) cathodically reduced in non aqueous solvents form radicals with the center of unpaired electron on the tetrazolyl ring (TBH₂, NTBH₂) as detected by EPR spectroscopy. After prolonged reduction, formazans (TBH₂, NTBH₂) are formed and are then further reduced to the nitro-centered anion radical (from NTBH₂) and the azogroup-centered anion radical (from TBH₂). The first cathodic peak in the cyclovoltammetric study in the region from -0.3 to -0.6 V vs. SCE (saturated calomel electrode) is irreversible and indicates an adsorption and diffusion process on the platinum and mercury electrodes. Formation of TBH⁺ and NTBH⁺ is assumed. The second peak, in the region from -0.8 to -1.3 V vs. SCE, is nearly reversible and coupled with the formation of TBH- and NTBH- radicals. UV-VIS spectra measured during the reduction show isosbestic points at the conversions: TB⁺⁺ → TBH⁺, NTB⁺⁺ - NTBH⁺; further, TBH⁺ → TBH₂ and NTBH⁺ → NTBH₂. The characteristic colours of the solutions observed can be used to characterise the reduction state of tetrazolium salts.     

Non-classical Immunity Controls Microbiota Impact on Skin Immunity and Tissue Repair

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Long-term and residual melanotropin-stimulated tyrosinase activity in S91 melanoma cells is density dependent

Summary Cell density is a factor that affects the capacity of Cloudman S91 melanoma cells to respond to melanotropins in monolayer culture. Continuous exposure of melanoma cells to α-melanotropin or its potent analog [Nle⁴,D-Phe⁷]-α-MSH, resulted in maximal stimulation of tyrosinase after 2 d of treatment, but the magnitude of stimulation decreased thereafter despite the continued presence of the melanotropins. However, when melanoma cells continually exposed to melanotropins were subcultured to an initial low cell density and maintained in contact with α-MSH or [Nle⁴,D-Phe⁷]-α-MSH (long-term culture), tyrosinase activity was rapidly restored and greatly enhanced. Also, when cells were seeded at initial densities ranging from 0.2 to 3.2×10⁶ cells/flask, and exposed for 24 h to 10⁻⁷ M α-MSH, only the cultures seeded at low densities (0.2 and 0.4×10⁶ cells/flask) exhibited maximal tyrosinase activity during the 24 h exposure to the melanotropins. Therefore, tyrosinase activity was primarily affected by cell density rather than by the duration of time the cells were in culture or by continuous exposure to melanotropin. Other flasks of various cell densities were treated with 10⁻⁷ M α-MSH or [Nle⁴,D-Phe⁷]-α-MSH for 24 h, followed byremoval of the melanotropins from the culture medium. The magnitude and duration of theresidual stimulation of melanoma tyrosinase activity by melanotropins were also found to be dependent on the initial cell density. These results reveal that there is a limited range of optimal cell densities at which melanoma cells can respond to melanotropins and express increased tyrosinase activity.     

The landscape of viral proteomics and its potential to impact human health

Introduction: Advances in mass spectrometry-based proteomic technologies are enhancing studies of viral pathogenesis. Identification and quantification of host and viral proteins and modifications in cells and extracellular fluids during infection provides useful information about pathogenesis, and will be critical for directing clinical interventions and diagnostics.

Areas covered: Herein we review and discuss a broad range of global proteomic studies conducted during viral infection, including those of cellular responses, protein modifications, virion packaging, and serum proteomics. We focus on viruses that impact human health and focus on experimental designs that reveal disease processes and surrogate markers.

Expert commentary: Global proteomics is an important component of systems-level studies that aim to define how the interaction of humans and viruses leads to disease. Viral-community resource centers and strategies from other fields (e.g., cancer) will facilitate data sharing and platform-integration for systems-level analyses, and should provide recommended standards and assays for experimental designs and validation.     

Effect of conserved intersubunit amino acid substitutions on Hfq protein structure and stability

Hfq is a thermostable RNA-binding bacterial protein that forms a uniquely shaped homohexamer. Based on sequence and structural similarity, Hfq belongs to the like-Sm (LSm) protein family. In spite of a rather high degree of homology between archaeal and eukaryotic LSm proteins, their quaternary structure is different, usually consisting of five to eight monomers. In this work, the importance of conserved intersubunit hydrogen bonds for the Hfq spatial organization was tested. The structures and stabilities for the Gln8Ala, Asn28Ala, Asp40Ala, and Tyr55Ala Hfq mutants were determined. All these proteins have the same hexamer organization, but their stability is different. Elimination of a single intersubunit hydrogen bond due to Gln8Ala, Asp40Ala, and Tyr55Ala substitutions results in decreased stability of the Hfq hexamer. Tyr55Ala Hfq as well as the earlier studied His57Ala Hfq has reduced protein thermostability, which seems to correspond to an opening of the protein hydrophobic core.     

Integrated network analysis of transcriptomic and proteomic data in psoriasis

Background  

Psoriasis is complex inflammatory skin pathology of autoimmune origin. Several cell types are perturbed in this pathology, and underlying signaling events are complex and still poorly understood.     

Pep2Path: Automated Mass Spectrometry-Guided Genome Mining of Peptidic Natural Products

Nonribosomally and ribosomally synthesized bioactive peptides constitute a source of molecules of great biomedical importance, including antibiotics such as penicillin, immunosuppressants such as cyclosporine, and cytostatics such as bleomycin. Recently, an innovative mass-spectrometry-based strategy, peptidogenomics, has been pioneered to effectively mine microbial strains for novel peptidic metabolites. Even though mass-spectrometric peptide detection can be performed quite fast, true high-throughput natural product discovery approaches have still been limited by the inability to rapidly match the identified tandem mass spectra to the gene clusters responsible for the biosynthesis of the corresponding compounds. With Pep2Path, we introduce a software package to fully automate the peptidogenomics approach through the rapid Bayesian probabilistic matching of mass spectra to their corresponding biosynthetic gene clusters. Detailed benchmarking of the method shows that the approach is powerful enough to correctly identify gene clusters even in data sets that consist of hundreds of genomes, which also makes it possible to match compounds from unsequenced organisms to closely related biosynthetic gene clusters in other genomes. Applying Pep2Path to a data set of compounds without known biosynthesis routes, we were able to identify candidate gene clusters for the biosynthesis of five important compounds. Notably, one of these clusters was detected in a genome from a different subphylum of Proteobacteria than that in which the molecule had first been identified. All in all, our approach paves the way towards high-throughput discovery of novel peptidic natural products. Pep2Path is freely available from http://pep2path.sourceforge.net/, implemented in Python, licensed under the GNU General Public License v3 and supported on MS Windows, Linux and Mac OS X.

This is a PLOS Computational Biology Software Article.

     

On the role of some conserved and nonconserved amino acid residues in the transitional state and intermediate of apomyoglobin folding

The contributions of some amino acid residues in the A, B, G, and H helices to the formation of the folding nucleus and folding intermediate of apomyoglobin were estimated. The effects of point substitutions of Ala for hydrophobic amino acid residues on the structural stability of the native (N) protein and its folding intermediate (I), as well as on the folding/unfolding rates for four mutant apomyoglobin forms, were studied. The equilibrium and kinetic studies of the folding/unfolding rates of these mutant proteins in a wide range of urea concentrations demonstrated that their native state was considerably destabilized as compared with the wild-type protein, whereas the stability of the intermediate state changed moderately. It was shown that the amino acid residues in the A, G, and H helices contributed insignificantly to the stabilization of the apomyoglobin folding nucleus in the rate-limiting I ? N transition, taking place after the formation of the intermediate, whereas the residue of the B helix was of great importance in the formation of the folding nucleus in this transition.     

Predatory cues drive colony size reduction in marine diatoms

1. Colony formation is a common feature among nonmotile marine phytoplankton. Several theories exist around the potential benefits of larger colonies.
2. Here, we test the hypothesis that predation is one of the drivers behind colony formation and chain length plasticity. We exposed cultures of Thalassiosira rotula, Chaetoceros curvisetus, and Chaetoceros affinis to copepodamides, a chemical alarm cue released by copepods and perceived as an indicator of predation threat by their prey. This was coupled with a grazing experiment, which compared copepod grazing rates on different chain lengths.
3. Our results show that T. rotula and C. curvisetus decreased their chain lengths by 79% and 49%, respectively, in response to copepodamides. Single cells and short chains were grazed at lower rates compared with long chains, and the copepodamide‐driven size shift led to 30% and 12% lower grazing in T. rotula and C. curvisetus, respectively. In contrast, C. affinis showed a slight increased chain length in response to copepodamides although nonsignificant.
4. We found that 2 of 3 studied species reduce their chain length in response to the presence of copepod grazers. Altered size structure has implications for the route of carbon in the marine food webs and carbon export to deeper strata.