Cycloartane glycosides from the rhizomes of Curculigo orchioides

Cycloartane glycosides (1-9) were isolated from rhizomes of Curculigo orchioides (Hypoxidaceae), and this structures were determined by spectroscopic analysis and a few chemical transformations. Cytotoxic activity of glycosides (1-9) and their common aglycone (1a) against HL-60 human promyelocytic leukemia cells was also examined.     

The binding of a thyroid hormone metabolite, 3-monoiodo-L-thyronine, to bovine serum albumin as measured by circular dichroism

The interaction between a thyroid hormone metabolite, 3-monoiodo-L-thyronine (3-T1) and bovine serum albumin (BSA) was investigated by using the CD method. An enhanced CD band was observed at the absorption wavelength region of 3-T1 around 293 nm suggesting the binding of 3-T1 to the BSA molecule. The ellipticity at 293 nm was measured at various molar ratios of 3-T1 to BSA, and the apparent binding constant and the maximum number of binding sites could be estimated as Kapp = 8.85 +/- 1.07 X 10(4) M-1 and n = 23.8 +/- 0.9 respectively. The CD of a mixture of BSA, 3-T1 and thyroxine (T4) was also studied at various pH's. The pH profile of the two characteristic CD bands at 293 nm and 320 nm, attributed to bound 3-T1 and T4, suggested that the optimum binding condition of 3-T1 was attained at alkaline pH of around 9, while that of T4 was attained over a wide pH range between 5-10. A significant role of the ionized 4'-hydroxyl group of 3-T1 in the binding reaction with BSA is also suggested.     

Plant cell wall degradation by saprophytic Bacillus subtilis strains: gene clusters responsible for rhamnogalacturonan depolymerization

Plant cell wall degradation is a premier event when Bacillus subtilis, a typical saprophytic bacterium, invades plants. Here we show the degradation system of rhamnogalacturonan type I (RG-I), a component of pectin from the plant cell wall, in B. subtilis strain 168. Strain 168 cells showed a significant growth on plant cell wall polysaccharides such as pectin, polygalacturonan, and RG-I as a carbon source. DNA microarray analysis indicated that three gene clusters (yesOPQRSTUVWXYZ, ytePQRST, and ybcMOPST-ybdABDE) are inducibly expressed in strain 168 cells grown on RG-I. Cells of an industrially important bacterium, B. subtilis strain natto, fermenting soybeans also express the gene cluster including the yes series during the assimilation of soybean used as a carbon source. Among proteins encoded in the yes cluster, YesW and YesX were found to be novel types of RG lyases releasing disaccharide from RG-I. Genetic and enzymatic properties of YesW and YesX suggest that strain 168 cells secrete YesW, which catalyzes the initial cleavage of the RG-I main chain, and the resultant oligosaccharides are converted to disaccharides through the extracellular exotype YesX reaction. The disaccharide is finally degraded into its constituent monosaccharides through the reaction of intracellular unsaturated galacturonyl hydrolases YesR and YteR. This enzymatic route for RG-I degradation in strain 168 differs significantly from that in plant-pathogenic fungus Aspergillus aculeatus. This is, to our knowledge, the first report on the bacterial system for complete RG-I main chain degradation.     

Synthesis of UV-curable chitosan derivatives and palladium (II) adsorption behavior on their UV-exposed films

Novel chitosan derivatives with UV-curable functional groups, such as 3-methoxy-4-(2-hydroxy-3-methacryloyloxypropoxy)benzyl, 3,4-bis(2-hydroxy-3-methacryloyloxypropoxy)benzyl, 3-methoxy-4-methacryloyloxybenzyl, and 3,5-dimethacryloyloxybenzyl groups, were prepared. Introduction of photosensitive functional groups to chitosan was accomplished by reductive N-alkylation via Schiff’s bases using corresponding photosensitive aldehydes. Compared to starting chitosan, UV-curable chitosan derivatives showed better solubility in several organic solvents, such as DMSO and 70% methacrylic acid. The solubility of these compounds increased with an increase in the degree of substitution of the N-alkyl side chains. After UV irradiation for 20 s under a high-pressure mercury lamp at a distance of 15 cm from the samples, acidic methanol solutions of these derivatives were transformed to gels in the presence of photo-initiator, and their dried films adsorbed palladium (II) at pH 1.1 and pH 5.3. The UV-curable chitosan derivatives were successfully used as coating materials for electroless plating on non-conductive substances.     

Efficient induction of formate hydrogen lyase of aerobically grown <Emphasis Type="Italic">Escherichia coli</Emphasis> in a three-step biohydrogen production process

A three-step biohydrogen production process characterized by efficient anaerobic induction of the formate hydrogen lyase (FHL) of aerobically grown Escherichia coli was established. Using E. coli strain SR13 (fhlA ⁺⁺, ΔhycA) at a cell density of 8.2 g/l medium in this process, a specific hydrogen productivity (28.0 ± 5.0 mmol h⁻¹ g⁻¹ dry cell) of one order of magnitude lower than we previously reported was realized after 8 h of anaerobic incubation. The reduced productivity was attributed partly to the inhibitory effects of accumulated metabolites on FHL induction. To avoid this inhibition, strain SR14 (SR13 ΔldhA ΔfrdBC) was constructed and used to the effect that specific hydrogen productivity increased 1.3-fold to 37.4 ± 6.9 mmol h⁻¹ g⁻¹. Furthermore, a maximum hydrogen production rate of 144.2 mmol h⁻¹ g⁻¹ was realized when a metabolite excretion system that achieved a dilution rate of 2.0 h⁻¹ was implemented. These results demonstrate that by avoiding anaerobic cultivation altogether, more economical harvesting of hydrogen-producing cells for use in our biohydrogen process was made possible.     

Determining the critical nucleus and mechanism of fibril elongation of the Alzheimer's Abeta(1-40) peptide

We use a coarse-grained protein model to characterize the critical nucleus, structural stability, and fibril elongation propensity of Aβ_1-40 oligomers for the C_2x and C_2z quaternary forms proposed by solid-state NMR. By estimating equilibrium populations of structurally stable and unstable protofibrils, we determine the shift in the dominant population from free monomer to ordered fibril at a critical nucleus of ten chains for the C_2x and C_2z forms. We find that a minimum assembly of 16 monomer chains is necessary to mimic a mature fibril, and show that its structural stability correlates with a plateau in the hydrophobic residue density and a decrease in the likelihood of losing hydrophobic interactions by rotating the fibril subunits. While Aβ_1-40 protofibrils show similar structural stability for both C_2x and C_2z quaternary structures, we find that the fibril elongation propensity is greater for the C_2z form relative to the C_2x form. We attribute the increased propensity for elongation of the C_2z form as being due to a stagger in the interdigitation of the N-terminal and C-terminal β-strands, resulting in structural asymmetry in the presented fibril ends that decreases the amount of incorrect addition to the N terminus on one end. We show that because different combinations of stagger and quaternary structure affect the structural symmetry of the fibril end, we propose that differences in quaternary structures will affect directional growth patterns and possibly different morphologies in the mature fiber.     

Suppression of viral replication by stress-inducible GADD34 protein via the mammalian serine/threonine protein kinase mTOR pathway

GADD34 is a protein that is induced by a variety of stressors, including DNA damage, heat shock, nutrient deprivation, energy depletion, and endoplasmic reticulum stress. Here, we demonstrated that GADD34 induced by vesicular stomatitis virus (VSV) infection suppressed viral replication in wild-type (WT) mouse embryo fibroblasts (MEFs), whereas replication was enhanced in GADD34-deficient (GADD34-KO) MEFs. Enhanced viral replication in GADD34-KO MEFs was reduced by retroviral gene rescue of GADD34. The level of VSV protein expression in GADD34-KO MEFs was significantly higher than that in WT MEFs. Neither phosphorylation of eIF2alpha nor cellular protein synthesis was correlated with viral replication in GADD34-KO MEFs. On the other hand, phosphorylation of S6 and 4EBP1, proteins downstream of mTOR, was suppressed by VSV infection in WT MEFs but not in GADD34-KO MEFs. GADD34 was able to associate with TSC1/2 and dephosphorylate TSC2 at Thr1462. VSV replication was higher in TSC2-null cells than in TSC2-expressing cells, and constitutively active Akt enhanced VSV replication. On the other hand, rapamycin, an mTOR inhibitor, significantly suppressed VSV replication in GADD34-KO MEFs. These findings demonstrate that GADD34 induced by VSV infection suppresses viral replication via mTOR pathway inhibition, indicating that cross talk between stress-inducible GADD34 and the mTOR signaling pathway plays a critical role in antiviral defense.     

In Situ Activity and Spatial Organization of Anaerobic Ammonium-Oxidizing (Anammox) Bacteria in Biofilms

We investigated autotrophic anaerobic ammonium-oxidizing (anammox) biofilms for their spatial organization, community composition, and in situ activities by using molecular biological techniques combined with microelectrodes. Results of phylogenetic analysis and fluorescence in situ hybridization (FISH) revealed that “Brocadia”-like anammox bacteria that hybridized with the Amx820 probe dominated, with 60 to 92% of total bacteria in the upper part (<1,000 μm) of the biofilm, where high anammox activity was mainly detected with microelectrodes. The relative abundance of anammox bacteria decreased along the flow direction of the reactor. FISH results also indicated that Nitrosomonas-, Nitrosospira-, and Nitrosococcus-like aerobic ammonia-oxidizing bacteria (AOB) and Nitrospira-like nitrite-oxidizing bacteria (NOB) coexisted with anammox bacteria and accounted for 13 to 21% of total bacteria in the biofilms. Microelectrode measurements at three points along the anammox reactor revealed that the NH₄⁺ and NO₂⁻ consumption rates decreased from 0.68 and 0.64 μmol cm⁻² h⁻¹ at P2 (the second port, 170 mm from the inlet port) to 0.30 and 0.35 μmol cm⁻² h⁻¹ at P3 (the third port, 205 mm from the inlet port), respectively. No anammox activity was detected at P4 (the fourth port, 240 mm from the inlet port), even though sufficient amounts of NH₄⁺ and NO₂⁻ and a high abundance of anammox bacteria were still present. This result could be explained by the inhibitory effect of organic compounds derived from biomass decay and/or produced by anammox and coexisting bacteria in the upper parts of the biofilm and in the upstream part of the reactor. The anammox activities in the biofilm determined by microelectrodes reflected the overall reactor performance. The several groups of aerobic AOB lineages, Nitrospira-like NOB, and Betaproteobacteria coexisting in the anammox biofilm might consume a trace amount of O₂ or organic compounds, which consequently established suitable microenvironments for anammox bacteria.