Data on nucleoid-associated proteins isolated from Mycoplasma gallisepticum after intracellular infection.

Mycoplasma gallisepticum (M. gallisepticum) belongs to the class of Mollicutes. It causes chronic respiratory disease in avian species. It is characterized by lack of cell wall and reduced genome size. As a result of genome reduction, M. gallisepticum has a limited variety of DNA-binding proteins (DBP) and transcription factors. Consequently, the diversity of DNA-binding proteins and transcription factors (TF) in M. gallisepticum is limited in comparison with related bacteria such as Bacillus subtilis. Studies have shown, however, that mycoplasmas demonstrate a wide range of differential expression of genes in response to various stress factors, which promotes effective adaptation to unfavorable conditions. We assume that in the case of mycoplasmas, which are characterized by a combination of the reduction of known gene expression regulation systems and a high adaptive potential, the coordination of gene expression can be provided due to local changes in the structure and spatial organization of the chromosome. The study of the dynamic changes of the proteomic profile of M. gallisepticum nucleoid may assist in revealing its mechanisms of functioning, regulation of chromosome organization and stress adaptation including its changes upon invasion of the host cells.

The response of two polar monocyte subsets to inflammation.

Macrophages are a central component of innate immunity that play an important role in the defense of the organism. Macrophages are highly plastic and are activated by interaction with other cells and environmental factors. In this work, we study the effect of lipopolysaccharide on macrophages derived from the two most polar (CD14+ and CD16+ monocytes) as well as the intermediate subset of blood monocytes from healthy donors and assess what happens to the subset most prone to polarization on the transcriptomic and proteomic level. It has been shown that, according to primary pro-inflammatory polarization markers, their cytokine profile, and their phagocytic activity, macrophages derived from CD14+ monocytes exhibit higher sensitivity to inducers of pro-inflammatory polarization. Flow cytometry analysis revealed increased levels of CD86, while secretome analysis demonstrated significant increase of pro-inflammatory and anti-inflammatory cytokines observed in CD14+-derived macrophages, as compared to CD16+-derived macrophages in conditioned media. Assessment of the transcriptome and proteome of CD14+-derived macrophages with further bioinformatic analysis identified the most significant differences after polarization towards the pro-inflammatory phenotype. Immune-, membrane-, IFN-γ-, cytokine-, and defense-associated pathways were found significantly prevalent, while downregulated pathways were represented by RNA binding-, housekeeping-, exocytosis-, intracellular transport-, peptide and amide metabolic-related signaling. This data could be useful for macrophage-based cell therapeutics of cancer, as it provides additional background for the manipulation of donor monocytes intended for back transplantation.

The effect of phytoglobin overexpression on the plant proteome during nonhost response of barley (Hordeum vulgare) to wheat powdery mildew (Blumeria graminis f. sp. tritici).

Nonhost resistance, a resistance of plant species against all nonadapted pathogens, is considered the most durable and efficient immune system in plants. To increase our understanding of the response of barley plants to infection by powdery mildew, Blumeria graminis f. sp. tritici, we used quantitative proteomic analysis (LC-MS/MS). We compared the response of two genotypes of barley cultivar Golden Promise, wild type (WT) and plants with overexpression of phytoglobin (previously hemoglobin) class 1 (HO), which has previously been shown to significantly weaken nonhost resistance. A total of 8804 proteins were identified and quantified, out of which the abundance of 1044 proteins changed significantly in at least one of the four comparisons ('i' stands for 'inoculated')- HO/WT and HOi/WTi (giving genotype differences), and WTi/WT and HOi/HO (giving treatment differences). Among these differentially abundant proteins (DAP) were proteins related to structural organization, disease/defense, metabolism, transporters, signal transduction and protein synthesis. We demonstrate that quantitative changes in the proteome can explain physiological changes observed during the infection process such as progression of the mildew infection in HO plants that was correlated with changes in proteins taking part in papillae formation and preinvasion resistance. Overexpression of phytoglobins led to modification in signal transduction prominently by dramatically reducing the number of kinases induced, but also in the turnover of other signaling molecules such as phytohormones, polyamines and Ca2+. Thus, quantitative proteomics broaden our understanding of the role NO and phytoglobins play in barley during nonhost resistance against powdery mildew.

Helitrons and Retrotransposons Are Co-localized in Bos taurus Genomes.

BACKGROUND: DNA transposons helitrons are mobile genetic elements responsible for major movements of the genetic material within and across different genomes. This ability makes helitrons suitable candidate elements for the development of new approaches of multilocus genotyping of live-stock animals, along with the well-known microsatellite loci. OBJECTIVE: We aimed to estimate the informativeness of helitron and microsatellite markers in assessing the consolidation and the "gene pool" standards of two commercial dairy cattle breeds (Ayrshire breed and holsteinized Black-and-White cattle) and one local breed of Kalmyk cattle, and to reveal any inter-breed difference in the organization of genomic regions flanked by helitrons in the studied cattle breeds. METHOD: We used the combination of two highly-polymorphic genomic elements - helitrons and trinu-cleotide microsatellites (AGC)6G and (GAG)6C, respectively - for genome scanning of the sampled groups of cattle. Also, we pyrosequenced the genomic regions flanked by the inverted repeats of 3'-end of Heligloria family of helitron fragments. RESULTS: Generally, the both combinations of markers generated polymorphic spectra, based on which certain interbreed differentiation could be observed. The analysis of the identified interspersed repeats suggests that in factory and local cattle the genomic regions flanked by helitron fragments are shaped differently and contain different superfamilies of transposable elements, especially retrotransposons. CONCLUSION: Despite the well-known fact of retrotransposon-dependent microsatellite expansion, our data suggest that, in the cattle genome, the DNA transposons and microsatellites can also be found in close neighbourhood, and that helitrons and retrotransposons may form domains of increased variability - targets for factors of artificial selection.

[Influence of sociohygienic factors on the shaping of the nutritional status in children and teenagers].

There was revealed the structure of deteriorations in the nutritional status of schoolchildren in the city: the most of students has normal nutritional status, but there was noted the high prevalence of excessive body weight and obesity among children and teenagers. Risk factors for development of deteriorations of the nutrition state were detected as follows: irrational food regimen, qualitative compartment offood, factors of educational environment, lifestyle. The main role in system of control of the nutritional status in children is referred to the correction of socio-hygienic factors which prove to be the priority ones in the shaping of the nutritional status in students. As the main condition determining the nutrition state of the up-to-date schoolchildren and the quality of their life in the whole the social cultural level of children and adolescents must be regarded as a result of the hygienic education and training in fundamentals of healthy lifestyle. Priority protective factors of the gain in the part of schoolchildren with normal nutritional status (optimalfood regimen, optimal dietary habits, sufficient level of physical activity) laidfrom the child age in conditions of the family, sufficient level of the physical activity and the implementation of the other element of hygienically expedient day regimen served as the base for the elaboration of the system of the control of nutritional status. Algorithm of the control of the nutritional status in the students of educational institutions includes the creation of healthcare educational environment, optimization of nutrition and physical activity, the shaping of the culture of healthy lifestyle, health-improving measures for children with disorders of nutritional status and their psychological pedagogical supports at the stage of the correction of the nutritional status, improvement of the medical service for the early detection of deviations of nutritional status with the estimation of the efficiency of the system ofpreventive and health-improving measures.

Silicon Crystals Formation Using Silicatein-Like Cathepsin of Marine Sponge Latrunculia oparinae.

The cDNA fragment encoding the catalytic domain of the new silicatein-like cathepsin enzyme LoCath was expressed in a strain Top10 of Escherichia coli, extracted and purified via nickel-affinity chromatography. Recombinant enzyme performed silica-polymerizing activity when mixed with water-soluble silica precursor-tetrakis-(2-hydroxyethyl)-orthosilicate. Scanning electron microscopy revealed hexagonal, octahedral and ß-tridimit crystals. Energy dispersion fluorescence X-ray spectrometry analysis showed that all these crystals consist of pure silicon oxide. It is the first report about the ability of marine sponge's cathepsin to polymerize silicon, as well as about the structure and composition of the silicon oxide crystal formed by recombinant cathepsin. Further study of the catalytic activity of silicatein and cathepsin will help to understand the biosilification processes in vivo, and will create basis for biotechnological use of recombinant proteins for silicon polymerization.

[Novel Water-Soluble Substrate for Silicateins].

We suggested to use tetrakis(2-hydroxyethyl)orthosilicate (THEOS) as a substrate for silicateins--an enzyme family playing a key role in formation of skeleton in marine sponges. We compared THEOS with tetraethylorthosilicate (TEOS)--a commonly used substrate for silicateins. These substrates were tested in reaction of amorphous silica formation in vitro catalyzed by silicatein Al from sponge Latrunculia oparinae. It was found that reaction with THEOS occurs more efficiently than with TEOS, probably due to high water solubility and higher hydrolysis rate of THEOS.

Glycosidases of marine organisms.

This review discusses the catalytic properties, activity regulation, structure, and functions of O-glycoside hydrolases from marine organisms exemplified by endo-1→3-ß-D-glucanases of marine invertebrates.

Purification and characterisation of recombinant Bacteroides fragilis toxin-2.

Fragilysin (BFT) is metalloprotease that is secreted by enterotoxigenic Bacteroides fragilis. Studying the mechanism of BFT interaction with intestinal epithelial cells requires a pure protein sample. In this study, we cloned DNA-fragments coding for the catalytic domain of fragilysin-2 and profragilysin-2 into an E. coli expression vector. Purification methods for the recombinant fragilysin-2 catalytic domain and profragilysin-2 were developed. In addition, we obtained mature active fragilysin-2 from recombinant proprotein by limited tryptic digestion. We tested the biological activity of the recombinant protein samples and revealed that E-cadherin was cleaved when HT-29 cells were treated with mature fragilysin-2 but not with profragilysin-2. Azocoll, azocasein and gelatin were not proteolytically cleaved by mature fragilysin-2. Proteins released in culture medium after HT-29 cells treatment with mature active BFT-2 were identified.

Catalytic properties and amino acid sequence of endo-1→3-ß-D-glucanase from the marine mollusk Tapes literata.

A specific 1→3-ß-D-glucanase with molecular mass 37 kDa was isolated in homogeneous state from crystalline style of the commercial marine mollusk Tapes literata. It exhibits maximal activity within the pH range from 4.5 to 7.5 at 45°C. The 1→3-ß-D-glucanase catalyzes hydrolysis of ß-1→3 bonds in glucans as an endoenzyme with retention of bond configuration, and it has transglycosylating activity. The K(m) for hydrolysis of laminaran is 0.25 mg/ml. The enzyme is classified as a glucan endo-(1→3)-ß-D-glucosidase (EC 3.2.1.39). The cDNA encoding this 1→3-ß-D-glucanase from T. literata was sequenced, and the amino acid sequence of the enzyme was determined. The endo-1→3-ß-D-glucanase from T. literata was assigned to the 16th structural family (GHF 16) of O-glycoside hydrolases.

Induction of lipid flip-flop by colicin E1 - a hallmark of proteolipidic pore formation in liposome membranes.

The addition of the channel-forming domain of colicin E1 to liposomes elicited the transmembrane diffusion (flip-flop) of lipids concomitant to the release of the fluorescent dye from liposomes. Good correlation was found between kinetic and concentration dependences of the two processes. Both the liposome leakage and the lipid flip-flop were stimulated upon alkalinization of the buffer solution after colicin binding at acidic pH. These results in combination with the analysis of the data on colicin binding to liposomes provide evidence in favor of the validity of the toroidal (proteolipidic) pore model as the mechanism of colicin channel formation.

Estrogenic activity of triterpene glycosides in yeast two-hybrid assay.

Estrogenic potency of six triterpene glycosides, Holothurin A, Holotoxin A1, Frondoside A, Cucumarioside A2-2 and Cauloside C, that are natural products and semi-synthesized Ginsenoside-Rh2, were examined with yeast two-hybrid system, including expressed genes of human estrogen receptor, hERalpha, the co-activator TIF2 and lacZ as a reporter gene. Only Ginsenoside-Rh2 exhibited significant moderate estrogenic activity in the concentration range of 10(-7) to 10(-6)M. Its effect was approximately 30% of the activity of 17beta-estradiol applied at half-effective concentration. This indicates Ginsenosides-Rh2 is a weak phytoestrogen. The sea cucumber triterpene glycosides, Holothurin A, Holotoxin A1, Cucumarioside A2-2 and Frondoside A, and plant glycoside Cauloside C had no appreciable estrogenic activity. Data obtained by yeast two-hybrid assay reflect structure-activity relationship between tested compounds and 17beta-estradiol. Only Ginsenoside-Rh2 has some similarity in chemical structure with 17beta-estradiol that might explain affinity of this glycoside to the hERalpha receptor.