Comparative study of two thioredoxin peroxidases from disk abalone (Haliotis discus discus): cloning, recombinant protein purification, characterization of antioxidant activities and expression analysis

Thioredoxin peroxidase (TPx), also named peroxiredoxin (Prx), is an important peroxidase, which can protect organisms against various oxidative stresses. Two TPxs were isolated from a disk abalone (Haliotis discus discus) cDNA library, named as AbTPx1 and AbTPx2, respectively. AbTPx1 and AbTPx2 consist of 1315 and 1045 bp full-length cDNA with 753 and 597 bp open reading frames encoding 251 and 199 amino acids, respectively. The TPx signature motif 1 (FYPLDFTFVCPTEI) and motif 2 (GEVCPA) were conserved in both AbTPx1 and AbTPx2 amino acid sequences. Purified recombinant abalone TPx fusion proteins catalyzed the reduction of H2O2 and butyl hydroperoxide in peroxidase assays. Furthermore, both AbTPx fusion proteins were shown to protect super-coiled DNA from damage by metal-catalyzed oxidation (MCO) in vitro. Escherichia coli cells transformed with AbTPx1 and AbTPx2 coding sequences in pMAL-c2x showed resistance to H2O2 at 0.8 mM concentration by in vivo H2O2 tolerance assay. AbTPx1 and AbTPx2 mRNA were constitutively expressed in gill, mantle, abductor muscle and digestive tract in a tissue specific manner. Additionally, both TPxs mRNA were up-regulated in gill and digestive tract tissues against H2O2 at 3h post injection. The results indicate that AbTPx1 and AbTPx2 gene expressions are induced by oxidative stress and their respective proteins function in the detoxification of different ROS molecules to maintain efficient antioxidant defense in disk abalone.     

Biological activity of water-soluble nanostructures of dihydroquercetin with cyclodextrins

The biological properties of dihydroquercetin (DHQ) modified by including it into the ring of β-cyclodextrin (β-CD) to give it more water-soluble properties have been investigated. It was shown that the peroral administration of the DHQ/β-CD complex provides a long increase of DHQ concentration in rat blood (up to 7.5 h), and, unlike pure DHQ, the complex does not accumulate in the liver. As DHQ is released from the complex, it penetrates into liposome membranes, changing their thermodynamic characteristics. DHQ decreases the specific heat absorption, enthalpies, and temperature maximum of lipid melting and increases the transition half-width. This property is used to estimate the stability of the DHQ/β-CD complex. It was shown that complex DHQ/β-CD is not stable, and DHQ molecules slowly leave the complex in water environment. Seven and a half hours after the peroral injection of drugs, DHQ was found in the blood plasma of rats to which water-soluble complex DHQ/βCD was injected and in the liver of rats to which free DHQ was injected. Thus, DHQ/βCD not only is a more water-soluble complex but also it slowly releases DHQ, supporting long a low concentration of the free form of DHQ and providing the penetration of DHQ into the blood stream. After several weeks of feeding old mice with antioxidants, the activity of mitochondrial enzymes was restored to the level observed in young animals.     

Maintenance of CMV-specific CD8+ T cell responses and the relationship of IL-27 to IFN-γ levels with aging

We investigated whether healthy young (age ? 40) and elderly (age ? 65) people infected with cytomegalovirus (CMV) had similar levels of CD8⁺ T cell cytokine production and proliferation in response to an immunodominant CMV pp65 peptide pool given the role of CD8⁺ T cells in controlling viral infection and the association of CMV with immunosenescence. Plus, we determined the effects of aging and CMV-infectious status on plasma levels of IL-27, an innate immune cytokine with pro- and anti-inflammatory properties, as well as on its relationship to IFN-γ in that IL-27 can promote the production of IFN-γ. The results of our study show that young and elderly people had similar levels of CD8⁺ T cell proliferation, and IFN-γ and TNF-α production in response to CMV pp65 peptides. Plasma levels of IL-27 were similar between the two groups although CMV-infected young and elderly people had a trend toward increased levels of IL-27. Regardless of aging and CMV-infectious status, plasma levels of IL-27 correlated highly with plasma levels of IFN-γ. These findings suggest the maintenance of CMV pp65-specific CD8⁺ T cell proliferation and cytokine production with aging as well as the sustaining of circulatory IL-27 levels and its biological link to IFN-γ in young and elderly people irrespective of CMV infection.     

Autophagy protein Rubicon mediates phagocytic NADPH oxidase activation in response to microbial infection or TLR stimulation

Phagocytosis and autophagy are two important and related arms of the host's first-line defense against microbial invasion. Rubicon is a RUN domain containing cysteine-rich protein that functions as part of a Beclin-1-Vps34-containing autophagy complex. We report that Rubicon is also an essential, positive regulator of the NADPH oxidase complex. Upon microbial infection or Toll-like-receptor 2 (TLR2) activation, Rubicon interacts with the p22phox subunit of the NADPH oxidase complex, facilitating its phagosomal trafficking to induce a burst of reactive oxygen species (ROS) and inflammatory cytokines. Consequently, ectopic expression or depletion of Rubicon profoundly affected ROS, inflammatory cytokine production, and subsequent antimicrobial activity. Rubicon's actions in autophagy and in the NADPH oxidase complex are functionally and genetically separable, indicating that Rubicon functions in two ancient innate immune machineries, autophagy and phagocytosis, depending on the environmental stimulus. Rubicon may thus be pivotal to generating an optimal intracellular immune response against microbial infection.     

Vertical distribution of bacterial community is associated with the degree of soil organic matter decomposition in the active layer of moist acidic tundra

The increasing temperature in Arctic tundra deepens the active layer, which is the upper layer of permafrost soil that experiences repeated thawing and freezing. The increasing of soil temperature and the deepening of active layer seem to affect soil microbial communities. Therefore, information on soil microbial communities at various soil depths is essential to understand their potential responses to climate change in the active layer soil. We investigated the community structure of soil bacteria in the active layer from moist acidic tundra in Council, Alaska. We also interpreted their relationship with some relevant soil physicochemical characteristics along soil depth with a fine scale (5 cm depth interval). The bacterial community structure was found to change along soil depth. The relative abundances of Acidobacteria, Gammaproteobacteria, Planctomycetes, and candidate phylum WPS-2 rapidly decreased with soil depth, while those of Bacteroidetes, Chloroflexi, Gemmatimonadetes, and candidate AD3 rapidly increased. A structural shift was also found in the soil bacterial communities around 20 cm depth, where two organic (upper Oi and lower Oa) horizons are subdivided. The quality and the decomposition degree of organic matter might have influenced the bacterial community structure. Besides the organic matter quality, the vertical distribution of bacterial communities was also found to be related to soil pH and total phosphorus content. This study showed the vertical change of bacterial community in the active layer with a fine scale resolution and the possible influence of the quality of soil organic matter on shaping bacterial community structure.     

Optimization of expression,purification, and NMR measurement for structural studies of transmembrane region of amyloid β protein

The amyloid precursor protein (APP) is an integral transmembrane protein which has been suggested to play a central role in the pathogenesis of Alzheimer’s disease. Despite the enormous amount of research conducted on amyloid protein, the precise mechanism of its toxic effect is not yet fully understood. To better understand the mechanism and function of amyloid protein, it is critical to elucidate the three-dimensional structure of the single transmembrane spanning region of human APP (hAPP-TM). Unfortunately, it is difficult to prepare the peptide sample because hAPP-TM is a membrane-bound protein that transverses the lipid bilayer of the cell membrane. Generally, the preparation of a transmembrane peptide is very difficult and time-consuming. In fact, high yield production of transmembrane peptides has been limited by experimental difficulties related to insufficient yields and the low solubility of such peptides. In this study, we describe experimental processes developed to optimize the expression, purification, and NMR measurement conditions for hAPP-TM transmembrane peptide.     

Distribution of the Ussur Brown Katydid,Paratlanticus ussuriensis,in South Korea

This study was conducted to monitor the abundance and distribution of the ussur brown katydid, Paratlanticus ussuriensis in South Korea, and to gather basic information about potential outbreak areas of this species. P. ussuriensis samples were collected from April to July in 2009 and 2010 using the transect method at the city or gun level. P. ussuriensis were identified in 86 cities or guns, and eight guns were determined to be potential risk areas of P. ussuriensis outbreaks. The current distribution of P. ussuriensis is well matched with the distribution of broadleaf trees. Their distribution and population density are expected to increase due to the expanding distribution of Quercus spp., the major food source of P. ussuriensis.