Bioscience, Biotechnology, and Biochemistry最新文献

Keratinase from Nocardiopsis sp. TOA-1 (NAPase) holds significant potential for industrial and medical applications. Here, we developed a heterologous secretory expression system for NAPase in Bacillus subtilis. The recombinant enzyme exhibited catalytic properties comparable to the native enzyme, demonstrating its suitability for further protein engineering. This work provides a foundation for enhancing NAPase activity and stability, expediting its biotechnological applications.

Angiogenin (Ang), an endoribonuclease belonging to the RNase A superfamily, cleaves the anticodon-loops of tRNAs to produce tRNA half molecules. Although previous studies have demonstrated the involvement of Ang in the pathobiology of neurodegenerative disorders, the characterization of Ang-generated tRNA halves in neuronal cells remains limited. This is partly due to the technical limitations of standard RNA-seq methods, which cannot capture Ang-generated RNAs containing a 2',3'-cyclic phosphate (cP). In this report, we established an Ang treatment model using SH-SY5Y, a human neuroblastoma cell line, and demonstrated Ang-dependent accumulation of tRNA halves. By performing cP-RNA-seq, which selectively captures cP-containing RNAs, we identified Ang-generated tRNA halves and the specific cleavage positions within tRNA anticodon-loops responsible for their generation. Our results provide insights into the anticodon-loop cleavage and the selective production of a specific subset of tRNA halves by Ang.

Lipopolysaccharide (LPS) causes inflammatory cholestasis in sepsis. We investigated the role of PDZK1 in the repression of ABC transporters in LPS-induced cholestasis. Lentiviral gene transfer of PDZK1 to rats was conducted to explore its influence on cholestasis induced by LPS. And the effect of lentivirus-mediated shRNA targeting PDZK1 on ABC transporters in rat liver BRL-3A cells was evaluated. Lentiviral vector encoding rat PDZK1 was administered to rats by tail intravenous injection. Obviously elevated serum total bile acid (TBA) level and liver biochemical markers in cholestatic rats were decreased by the Lv-PDZK1 delivery. Also Lv-PDZK1 delivery stimulated the suppressed hepatic ABC transporters expression. In vitro, after the lentiviral vector LV3/PDZK1 shRNA transfection, no expression of PDZK1 and mild expression of ABC transporters were detected in BRL-3A cells by Western blotting. Our results indicate that the lentiviral-mediated hepatocyte PDZK1 expression ameliorates LPS-induced cholestasis in rats by rescuing hepatic ABC transporters expression.

Strigolactones (SLs) are allelochemicals attracting both symbiotic arbuscular mycorrhizal (AM) fungi and root parasitic plants and a novel class of phytohormones inhibiting lateral shoot branching. In general, nutrient deficiency significantly promotes SL production in roots and exudation into the rhizosphere, and also induces suppressed shoot branching phenotype in host plants of AM fungi including rice, tomato, maize, etc. Nutrient deficiencies also inhibit the shoot branching in Arabidopsis thaliana (hereafter Arabidopsis), a non-host of AM fungi, while the level of carlactone, a non-canonical SL and the SL precursor for the other SLs, was reported to be unaffected. Because Arabidopsis SLs including CL and methyl carlactonoate,, are highly unstable, relationships between SL levels and shoot branching in Arabidopsis remain elusive. Herein, we demonstrate that nitrogen deficiency increases SL levels in the basal part of shoots in Arabidopsis and lateral shoot branching appears to be strongly regulated by these SLs.

To evaluate the effects of the soluble fiber isomaltodextrin in a protein-biased diet, a 21-day protein diet trial was conducted in rats, with 60% of the calories derived from protein. The results revealed that the high-protein diet alone led to a significant increase in kidney weight. In contrast, the consumption of water with 5% isomaltodextrin dissolved in it, along with a high-protein diet, suppressed this weight gain. To elucidate this mechanism, an analysis of serum urea toxins confirmed that the concentrations of phenyl sulfate were significantly higher with high protein, and significantly lower with isomaltodextrin. The impact of a high-protein on increased phenol in cecal contents, an increased that was mitigated by the isomaltodextrin. This suggests that the inhibitory effect of isomaltodextrin on renal hypertrophy was due to the suppression of urea toxin precursor production by the gut microbiota.

Circadian rhythms are biological systems that provide approximately 24-hour cycles for the behavior and physiological functions of organisms. As diverse modern lifestyles often cause disturbances in circadian rhythms, new approaches to their regulation are required. Therefore, new compounds that affect circadian rhythms have been explored in edible mushrooms. The extract from the culture filtrate of Cyclocybe cf. erebia showed activity that advanced the circadian rhythm in a bioassay with mouse fibroblasts expressing the LUCIFERASE protein under the control of the Period2 promoter. Bioassay-guided fractionation of the extract resulted in the isolation of the compound. Spectroscopic analyses identified the compound as a phthalide derivative, and the compound was named cyclocybelide. Treatment of mouse fibroblasts with the compound shifted the circadian rhythm forward, irrespective of the timing of treatment. In addition, some phthalide derivatives with hydroxy and methoxy groups showed similar effects on circadian rhythms.

The cholesterol balance and bile acid metabolism in a mouse model of hepatic lipid accumulation induced by a diet supplemented with cholesterol and cholic acid (CA) were quantitatively evaluated. The mice were fed diets supplemented with different levels of cholesterol (0, 3, or 6 g/kg diet) and CA (0.5 g/kg diet) for six weeks. Cholesterol supplementation doubled the hepatic triglyceride concentration, regardless of the supplementation level, without inflammation or gallstone formation. Both cholesterol supplementations enhanced fecal excretion of muricholic acid. Additionally, the higher cholesterol supplementation led to an increase in fecal cholesterol excretion, accompanied by elevated expression of hepatic cholesterol exporters and a reduction in fecal bile acid excretion. In this mouse study, supplementation with 3 g cholesterol/kg diet and 0.5 g CA/kg diet was sufficient to induce hepatic lipid accumulation.

We successfully constructed a heterologous expression system for L-glutamate oxidase from the marine actinomycete Streptomyces lydicamycinicus NBRC 110027 (Sl-LGOX) in Escherichia coli BL21(DE3) as a host. This is the first example of L-glutamate oxidase from a marine microorganism. A chemically synthesized gene optimized for codon usage in E. coli was used as the inserted fragment, which was effective for enzyme expression. We expressed Sl-LGOX in the soluble fraction of E. coli BL21(DE3)/pET21b-Sl-lgox. We also succeeded in purifying recombinant Sl-LGOX (rSl-LGOX) to homogeneity from the cell-free extract of this clone via an Ni-NTA column. rSl-LGOX showed high specificity for L-Glu and was active and stable over a wide range of temperatures and pH values. In particular, it showed high specific activity and stability at an acidic pH. A variety of applications can take advantage of the unique enzymatic properties of rSl-LGOX.

Manipulation and analysis methods for large DNAs are critical for epidemiological, clinical, diagnostic, and fundamental research on bacteria, membrane vesicles, plants, yeast, and human cells. However, the physical properties of large DNAs often challenge their manipulation and analysis with high accuracy and speed using the conventional methods such as gel electrophoresis and column-based methods. This review presents the approaches that leverage micrometer- and nanometer-sized gaps within microchannels to control the dynamics and conformations of large DNAs, thereby overcoming these challenges. By designing gap structures and migration conditions based on the relationship between gap parameters and the physical characteristics of large DNAs-such as diameter and persistence length-these methods enable swifter and more precise manipulation and analysis of large DNAs, including size separation, concentration, purification, and single-molecule analysis.

Caenorhabditis elegans is a type of nematode that has significantly contributed to aging research as a multicellular animal model because of its high reproductive rate, ease of cultivation, low cost, short generation cycle, body transparency, and eukaryotic nature. Since the discovery of long-lived mutant strains of C. elegans, signaling pathways involved in lifespan have been elucidated. Some of these pathways are shared with mammals, indicating that aging research in C. elegans may be applied to other animals, including humans. Studies on the mechanisms of aging have advanced with the availability of mutants for these pathways. In recent years, C. elegans has also contributed to the discovery of antioxidants and the elucidation of the molecular mechanisms responsible for the anti-aging effects of foods and traditional medicines with lifespan as an indicator. This review summarizes the characteristics of C. elegans for life span analysis associated with functional foods.