Development and Use of a Kinetical and Real-Time Monitoring System to Analyze the Replication of Hepatitis C Virus

In microbiological research, it is important to understand the time course of each step in a pathogen’s lifecycle and changes in the host cell environment induced by infection. This study is the first to develop a real-time monitoring system that kinetically detects luminescence reporter activity over time without sampling cells or culture supernatants for analyzing the virus replication. Subgenomic replicon experiments with hepatitis C virus (HCV) showed that transient translation and genome replication can be detected separately, with the first peak of translation observed at 3–4 h and replication beginning around 20 h after viral RNA introduction into cells. From the bioluminescence data set measured every 30 min (48 measurements per day), the initial rates of translation and replication were calculated, and their capacity levels were expressed as the sums of the measured signals in each process, which correspond to the areas on the kinetics graphs. The comparison of various HuH-7-derived cell lines showed that the bioluminescence profile differs among cell lines, suggesting that both translation and replication capacities potentially influence differences in HCV susceptibility. The effects of RNA mutations within the 5′ UTR of the replicon on viral translation and replication were further analyzed in the system developed, confirming that mutations to the miR-122 binding sites primarily reduce replication activity rather than translation. The newly developed real-time monitoring system should be applied to the studies of various viruses and contribute to the analysis of transitions and progression of each process of their life cycle.     

Identification of Multiple Domains of Entamoeba histolytica Intermediate Subunit Lectin-1 with Hemolytic and Cytotoxic Activities

Galactose and N-acetyl-D-galactosamine-inhibitable lectin of Entamoeba histolytica have roles in the pathogenicity of intestinal amoebiasis. Igl1, the intermediate subunit lectin-1 of E. histolytica, has been shown to have both hemolytic and cytotoxic activities that reside in the C-terminus of the protein. To identify the amino acid regions responsible for these activities, recombinant proteins were prepared and used in hemolytic and cytotoxic assays. The results revealed that Igl1 has multiple domains with hemolytic and cytotoxic activities and that amino acids 787-846, 968-1028 and 1029-1088 are involved in these activities. The hemolytic activities of the fragments were partly inhibited by mannose, galactose and N-acetylgalactosamine, and glucose showed lower or negligible inhibitory effects for the activities. This is the first report of a protozoan protein with hemolytic and cytotoxic activities in multiple domains.     

Studies on Pyrodextrinization of Corn Starch

During pyrodextrinization of corn starch in the absence of any added catalyst under different experimental conditions, the changes of dextrinization might have two or threes stages and remarkable transformation would begin to occur at 190°C. At higher temperatures, many non-carbohydrates are found. At lower temperatures, side chains of amylopectin-like substances might be cleaved to produce the linear parts.     

Cooling Process and Cast Structure of Zr-Al-Ni-Cu–Based Bulk Metallic Glasses Produced in Various Atmospheres

Cooling process in the production of Zr₆₅Al_7.5Ni₁₀Cu_17.5-based bulk metallic glasses (BMGs) was investigated in various chamber atmospheres in Cu mold casting. Two different cooling modes, consisting of the direct heat transfer between the melt and Cu mold in the high temperature and indirect transfer via cavity in the low-temperature regions, are suggested. In the later case, the cooling effect should depend on the chamber atmosphere, which results in the formation of glassy structure with large relaxation enthalpy casting under the ambient Ar and He atmospheres due to a good thermal conductivity. The less relaxed BMGs produced by an improved cooling effect are also expected to contain a large amount of free volume for significant deformability. Actually, it is clarified that the compressive plastic deformation is improved with an increase of the relaxation enthalpy. The present study indicates a necessity of development of the glassy structure, i.e., relaxation state, and provides a new technique of the formation of less relaxed glassy structure for the improvement of plasticity in BMGs.     

Improvement of Stainless Steel Making Operation Adapting Instable Raw Material Price

Sumikin Iron and Steel Corporation (SISC) is the main manufacturing base of seamless tube materials in Sumitomo Metals group, and stainless steel making plant is in charge of semi‐manufactured special steel products such as stainless and high alloy steel. Total manufacturing costs of these special steels are heavily dependent on the price of raw material. Therefore, reduction of its usage and optimizing operation process in order to absorb influence of its unstable price are important issues. In this report, we introduce three cases as following: yield improvement in order to reduce raw material usage, optimization of the steel dust usage, and active use of low‐price raw material through dividing refining process.     

Numerical computation of two-loop box diagrams with masses

A new approach is presented to evaluate multi-loop integrals, which appear in the calculation of cross-sections in high-energy physics. It relies on a fully numerical method and is applicable to a wide class of integrals with various mass configurations. As an example, the computation of two-loop planar and non-planar box diagrams is shown. The results are confirmed by comparisons with other techniques, including the reduction method, and by a consistency check using the dispersion relation.     

The terminal protein of the linear DNA plasmid pGKL2 shares an N-terminal domain of the plasmid-encoded DNA polymerase

The 36K protein attached at the 5′ end of the linear DNA plasmid pGKL2 from the yeast Kluyveromyces lactis was first purified and characterized. The terminal protein was purified from cells (1 kg wet weight) by ammonium sulphate precipitation and two rounds of centrifugation to equilibrium in CsCl gradients. The pGKL2 was present only in the post-microsomal supernatant. Approximately 10 mg of the purified pGKL2 was recovered and digested with DNase I. The terminal protein (final ca. 0·8 mg) was homogeneous by electrophoresis and we determined the N-terminal amino acid sequence up to ten residues, showing that it existed in the cryptic N-terminal domain of pGKL2-ORF2 (DNA polymerase) sequence.     

Crystalline Sodium Selenocarboxylates – synthesis and characterization

A series of sodium selenocarboxylates 2 were isolated from the reaction of diacyl selenides with sodium ethanolate and characterized. A convenient preparation of the sodium salts 2 by the direct reaction of acyl chlorides with sodium selenide was also established. The salts are colourless to slightly pale yellow crystals and labile towards moisture. They readily react with alkyl iodides at room temperature to give the corresponding Se-alkyl esters 3 .     

Fabrication of CaCO3-Coated Vesicles by Biomineralization and Their Application as Carriers of Drug Delivery Systems

We fabricated CaCO₃-coated vesicles as drug carriers that release their cargo under a weakly acidic condition. We designed and synthesized a peptide lipid containing the Val-His-Val-Glu-Val-Ser sequence as the hydrophilic part, and with two palmitoyl groups at the N-terminal as the anchor groups of the lipid bilayer membrane. Vesicles embedded with the peptide lipids were prepared. The CaCO₃ coating of the vesicle surface was performed by the mineralization induced by the embedded peptide lipid. The peptide lipid produced the mineral source, CO₃²⁻, for CaCO₃ mineralization through the hydrolysis of urea. We investigated the structure of the obtained CaCO₃-coated vesicles using transmission electron microscopy (TEM). The vesicles retained the spherical shapes, even in vacuo. Furthermore, the vesicles had inner spaces that acted as the drug cargo, as observed by the TEM tomographic analysis. The thickness of the CaCO₃ shell was estimated as ca. 20 nm. CaCO₃-coated vesicles containing hydrophobic or hydrophilic drugs were prepared, and the drug release properties were examined under various pH conditions. The mineralized CaCO₃ shell of the vesicle surface was dissolved under a weakly acidic condition, pH 6.0, such as in the neighborhood of cancer tissues. The degradation of the CaCO₃ shell induced an effective release of the drugs. Such behavior suggests potential of the CaCO₃-coated vesicles as carriers for cancer therapies.     

Effects of processing and cooking on the levels of pesticide residues in soybean samples

The effects of processing and cooking on the levels of pesticide residues in soybean samples were investigated for 14 pesticides in pre-harvest samples. On soaking, the transfer ratios (%, total pesticide residue amount in product/that in soybean) of soaked soybean were greater than 60% for most of the pesticides investigated. The transfer ratio of soymilk ranged from 37% to 92%, and that of tofu ranged from 7% to 63%. The processing factor (Pf, the concentration (mg/kg) of pesticide in product/that in soybean) of tofu ranged from 0.026 to 0.28. These values varied among pesticides. There was a high correlation between the log P(ow) and the transfer ratio of tofu. The test described here should be useful to obtain the transfer ratios of pesticide residues in processing and/or cooking steps.