The immune system and the commensal bacteria in the intestine, which together form the intestinal symbiotic system, greatly contribute to regulation of allergy. Of the various types of cells constituting the intestinal immune system, this review focuses on epithelial cells and mast cells and the interaction of these cells with commensals. Mast cells express the high affinity IgE receptor FcεRI which is essential to the induction of allergic inflammatory reactions. The molecular mechanisms of transcriptional regulation of genes encoding FcεRI have been clarified. On the other hand, the expression of the molecules involved in microbe recognition is regulated in a specific manner in intestinal epithelial cells, which are continuously exposed to the commensals inhabiting the intestinal lumen, to prevent excessive inflammatory reactions. Microbial components directly regulate the functions of mast cells through Toll-like receptors. These aspects provide targets for the regulation of allergy based on the maintenance of the intestinal symbiotic system.

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The volatile components of fresh leaves from 15 citrus species were investigated by headspace SPME with a GC-MS analysis. Three indigenous Taiwan citrus species, Citrus taiwanica, C. tachibana and C. depressa, were the major subjects. Eighty volatile organic compounds were detected as indicators of the genetic relationship. Linalool was the most abundant compound, and citronellal, geranial, neral, limonene and trans-β-ocimene were the major volatile compounds in fresh leaves. Linalool (56.37%) and myrcene (7.21%) were predominant in C. tawanica. An aldehyde-rich profile with citronellal (24.54%) contributed most to the aroma of leaves in C. tachibana, while Citrus depressa exhibited a high linalool/citronellal composition (23.56%/12.51%). The qualitative and quantitative patterns of the volatiles revealed that C. taiwanica was linked with sour orange, and either C. tachibana or C. depressa belonged to the mandarin group with C. tankan. Dendrograms also showed that the volatile patterns were related to the genetic classification.

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An extract of the edible parts of Hosta sieboldiana suppressed the growth of P388 mouse leukemic cells. A new steroidal saponin was isolated from Hosta sieboldiana in this study, and the structure was determined by a spectroscopic analysis to be (25R)-3β-(α-L-rhamnopyranosyl-(1→4)-β-D-glucopyranosyl-(1→3)-[β-D-glucopyranosyl-(1→2)]-β-D-glucopyranosyl-(1→4)-β-D-galactopyranosyl)-5α-spirostan-2α-ol.

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Tea catechins in a buffer at pH 7 with N₂ replacing O₂ epimerized rapidly at 80 °C with less than 10% of oxidative side reactions and gave catechin epimers in a 50–63% yield. The epimerization of catechins with three hydroxyl groups was faster than with two groups, and of galloyl-free catechins was faster than catechins with a galloyl ester.

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Methanolic extracts of edible plants and seaweeds were tested for their inhibitory activity against Jack bean urease. Quercetin-4′-O-β-D-glucopyranoside was isolated from Allium cepa as a urease inhibitor with an IC₅₀ value of 190 μM-. Quercetin and two quercetin glycosides, avicularin and guaijaverin, were isolated from Psidium guajava as urease inhibitors with respective IC₅₀ values of 80 μM-, 140 μM-, and 120 μM-.

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Aspernolide A and butyrolactones I and II showed inhibitory activities against soybean lipoxygenase. All of them also had DPPH (2,2-diphenyl-1-picrylhydrazyl) radical-scavenging activity. An analysis of the mechanism for radical scavenging allowed us to deduce that aspernolide A was converted to a quinone methide by a reaction with two molecules of the DPPH radical.

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Genome sequencing has revealed many pairs of proteins termed two-component systems (TCSs) in bacteria. Each pair consists of a sensor or histidine kinase (HK) and an effector or response regulator (RR). The HK is usually a membrane-spanning protein that senses specific environmental parameters and communicates this information to the cytoplasmic RR protein through phosphotransfer reactions to cope with a variety of environmental stresses, including osmotic pressure, nitrogen lack, phosphoric acid lack, and the presence of oxygen. Furthermore, some proteins have been identified as hybrid kinases composed of HK and RR. We identified the domain structures of 360 bacteria and 43 archaea by domain search against the PFAM database using HMMER. We then classified 8,573 HK, 10,807 RR, and 2,477 hybrid kinases. In addition, we identified specific domains among phylogenic clusters based on differences in domain structure of TCS genes applying the Signal-to-Noise ratio.

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A recombinant putative acid phosphatase from Thermus thermophilus was expressed and purified from Escherichia coli. The recombinant phosphatase displayed activities in a broad range of temperature, from 40 to 90 °C, with optimal temperature at 70 °C. In addition, the recombinant enzyme had activities in a wide range of pH, from 3.6 to 9.1, with optimal pH at 6 in acetate buffer and with optimal pH at 6.5 in Hepes buffer. Furthermore, it showed significant thermal stability and still possessed 44% residual activity after 70 °C treatment for 15 min. Moreover, the recombinant phosphatase showed broad substrates specificities for monophosphate esters, p-nitrophenyl phosphate (pNPP) being the most preferred substrate, and it was able to resist inhibition by sodium tartrate. Additionally, the recombinant protein formed stable oligomer under partially denatured conditions and required calcium ions for enzymic activity.

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The development and progression of esophageal cancer is associated with multiple alterations in the genome, including loss of the tumor suppressor phosphatase and tensin homolog deleted from the chromosome 10 (PTEN) gene. The purpose of this study was to determine the effects of adenovirus-mediated MMAC/PTEN expression on the growth and survival of human esophageal cancer cells in vitro and in vivo. We found that compared to control cells, overexpression of PTEN significantly suppressed growth and induced apoptosis in esophageal cancer cell lines Eca-109 and TE-1 via downregulation of Bcl-2 expression and changes in cell-cycle progression. Adenovirus PTEN also inhibited the growth of subcutaneous tumor xenografts by significantly reducing tumor size in vivo. Thus our results confirm the proposed functional role of MMAC/PTEN as a regulator of esophageal cancer progression in vivo and in vitro. PTEN might be an important biological marker and potential therapeutic target in the treatment of human esophageal cancer.

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AP-1-like transcription factors play crucial roles in oxidative stress responses in yeast and filamentous fungi. The deletion of an AP-1-like transcription factor gene, nap-1, in Neurospora crassa slightly increased its sensitivity to oxidative stressors, including menadione. Microarray and quantitative real-time reverse transcriptase-PCR analyses were employed to identify menadione-inducible genes (migs) and the roles of NAP-1 in their regulation. N. crassa migs include three putative glutathione S-transferase genes and two NADH:flavin oxidoreductase genes, orthologs of OYE2 and OYE3, both of which play roles in menadione tolerance in Saccharomyces cerevisiae. Menadione induced nuclear localization of NAP-1, and oxidative upregulation of many of migs were NAP-1 dependent. Genes for a thioredoxin, a glutathione reductase, and a glutathione peroxidase were slightly upregulated by the chemical only in the wild-type strain, suggesting that NAP-1 is involved in their oxidative induction and probably dose not contribute to high-level constitutive expressions of such genes.

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Hyperpigmentations are a serious concern addressed by both the medical community and the cosmetic industry through the development of agents that block melanin biosynthesis. In this study, we found that 2-amino-3H-phenoxazin-3-one (APO), isolated from extracts of the edible mushroom Agaricus bisporus Imbach, exhibited potent inhibitory effects on melanogenesis in B16 cells, a murine melanoma cell line. APO inhibited melanin biosynthesis at 1,000 times lower concentrations (IC₅₀=1.31±0.08 μM) than kojic acid (IC₅₀=1.31±0.13 mM), without causing cellular toxicity. APO did not directly inhibit the enzyme activity of tyrosinase, the rate-limiting melanogenic enzyme. Further study showed that APO inhibited the protein expression of tyrosinase and microphthalmia-associated transcription factor (MITF), a melanogenic transcription factor that regulates the expression of tyrosinase. These results suggest that APO is a promising depigmenting agent with both therapeutic and cosmetic value in preventing melanogenesis.

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Sensory neurons release calcitonin-gene related peptide (CGRP) on stimulation. We have reported that topical application of capsaicin increases facial skin elasticity by increasing the production of dermal insulin-like growth factor-I (IGF-I) through stimulation of sensory neurons in mice and humans. In this study, we examined whether topical application of α-D-glucosylglycerol (GG), a compound found in Japanese traditional brewed foods such as sake (Japanese rice wine), increases the dermal production of IGF-I in mice and increases the facial skin elasticity in females. GG increased CGRP release and cAMP levels in dorsal root ganglion (DRG) neurons isolated from wild-type mice. Pretreatment with capsazepine, an inhibitor of vanilloid receptor-1 activation, and with KT5720, an inhibitor of protein kinase A, reversed GG-induced increases in CGRP release from DRG neurons. Topical application of GG increased dermal levels of IGF-I, IGF-I mRNA, and collagen in wild-type mice, but not in CGRP-knockout mice. Topical application of GG increased cheek-skin elasticity in 13 female volunteers. These observations strongly suggest that GG increases the production of IGF-I in the skin through sensory neuron stimulation, thereby increasing skin elasticity.

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In the green sulfur bacterium Chlorobaculum tepidum, three sulfur oxidizing enzyme system (Sox) proteins, SoxAXK, SoxYZ, and SoxB (the core TOMES, thiosulfate oxidizing multi-enzyme system) are essential to in vitro thiosulfate oxidation. We purified monomeric flavoprotein SoxF from this bacterium, which had sulfide dehydrogenase activity. SoxF enhanced the thiosulfate oxidation activity of the purified core TOMES with various cytochromes as electron acceptors to different degrees without any change in the affinity for thiosulfate. The apparent reaction rates with 50 μM- C. tepidum cytochrome c-554 were slightly higher than with horse-heart cytochrome c, and the addition of 0.5 μM- SoxF increased the rate by 92%. The rates with 50 μM- horse-heart cytochrome c and 50 μM- horse-heart cytochrome c plus 0.5 μM- cytochrome c-554 were increased by SoxF by 31% and 120% respectively. We conclude that SoxF mediates electron transfer between the components of core TOMES and externally added cytochromes.

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Anoectochilus formosanus is used in traditional folk medicine as an hepatoprotective agent. The purpose of this study was to investigate the effects of a standardized aqueous extract of A. formosanus (SAEAF) on thioacetamide (TAA)-induced liver fibrosis. An in vitro study showed that the inhibitive effect of kinsenoside, a major component of SAEAF, on tumor necrosis factor α (TNF-α) secretion from Kupffer cells might be derived at least partly from downregulation of LPS-receptor Toll-like receptor 4 (TLR4) signaling. Hepatic fibrosis was produced by TAA (200 mg/kg, i.p.) 3 times per week for 12 weeks. Mice in the three TAA groups were treated daily with distilled water and SAEAF (1.0, 0.2 g/kg) via gastrogavage throughout the experimental period. The mice that received the SAEAF treatment had significantly reduced plasma alanine aminotransferase activity, relative liver weights, and hepatic hydroxyproline contents. A histological examination also confirmed that SAEAF reduced the degree of fibrosis caused by TAA treatment. RT-PCR analysis showed that SAEAF treatment reduced mRNA expression of collagen (α1)(I), lipopolysaccharide-binding protein, CD14, TLR4, and TNF receptor 1. An immunohistochemical examination also indicated that SAEAF reduced the number of CD68-positive cells (macrophages). In conclusion, oral administration of SAEAF significantly reduced TAA-induced hepatic fibrosis in mice, probably through inhibition of hepatic Kupffer cell activation.

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Trichoderonic acids A (1) and B (2), novel terpenoids, and (+)-heptelidic acid (3) isolated from cultures of a fungus, Trichoderma virens, and their structures were identified by spectroscopic analysis. These compounds selectively and competitively inhibited the activities of mammalian DNA polymerases β, λ (pols β, λ), and terminal deoxynucleotidyl transferase (TdT) in family X of pols, and compound 2 was a stronger inhibitor than compound 1 or 3. On the other hand, compounds 1–3 did not influence the activities of the other families (A-, B-, and the Y-families) of the mammalian pols tested, and showed no effect on the activities of plant pol α, fish pol δ, prokaryotic pol, or the other DNA metabolic enzymes tested. Compound 2 suppressed the growth of two human cancer cell lines, cervix carcinoma cells (HeLa) and breast carcinoma cells (MCF-7). The results suggest that these compounds identified inhibition among the families of mammalian pols.

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Several physical and psychological stresses frequently become triggers for gastrointestinal disorders such as ulcer. In this study, we tried to identify serum proteins as potential biomarkers for the evaluation of stress-induced gastric ulcer. By proteomic analysis using rats with gastric ulcer induced by water immersion and restraint (WIR) stress as an animal model, we found quantitative changes in several serum proteins, including creatine kinase muscle M chain (CK-M) and apolipoprotein A-IV (ApoA4) in the stressed rats. On western blotting and enzyme-linked immunosorbent assay (ELISA), we confirmed that serum CK-M was remarkably increased by WIR stress. However, ApoA4 appeared to be decreased by fasting, but not WIR stress, which is usually applied prior to WIR stress. The findings suggest that these two serum proteins might be useful as biomarkers, CK-M for stress-induced gastric ulcer and ApoA4 for starvation.

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Transcription and complementation experiments were carried out to analyze the regulatory hierarchy of two Streptomyces antibiotic regulatory protein (SARP) genes, srrY and srrZ, in the γ-butyrolactone (GB)-dependent regulatory cascade in Streptomyces rochei 7434AN4. The srrY gene was transcribed in the srrZ mutant, while the srrZ gene was not in the srrY mutant. The SrrY protein was specifically bound to the promoter region of srrZ, where a possible SARP binding sequence was identified 26 bp upstream of the −10 sequence. Deletion of the repeat sequences from this region abolished its SrrY binding activity. In addition, complementation of srrZ restored lankamycin production in the srrY mutant. All of these results confirm that the SARP gene srrY directly regulates expression of the second SARP gene srrZ in a positive manner. This study gave the first confirmation of direct regulation of two SARP genes in the GB-dependent regulatory cascade in Streptomyces.

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Matriptase is a type II transmembrane serine protease. Paradoxically, activation of this protease is thought to require its physiological inhibitor, hepatocyte growth factor activator inhibitor type-1 (HAI-1). In the present study, however, we obtained evidence in a stable transfection experiment using Madin–Darby canine kidney cells that matriptase activation does not require HAI-1.

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In order to establish a basis for transformation technology in the petroleum plant Euphorbia tirucalli, the callus of the plant was infected with Agrobacterium, washed with distilled water, sterilized with distilled water containing 100 mg/l of carbenicillin, selected on solidified B5 medium containing 13 mg/l of G418 and 100 mg/l of carbenicillin, and then on solidified B5 medium containing 25 mg/l of G418 and 100 mg/l of carbenicillin for the transgenic calli, and then the callus lines were subcultured successively on solidified B5 medium containing 50 mg/l of G418. We performed PCR analysis of sterilized G418-resistant callus line DNA and concluded that the gene introduced was integrated into the callus genome.

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The free flavin-dependent Fenton reaction was detected in cell-free extracts of Chlorella. The corresponding enzyme was purified to homogeneity, and its N-terminal sequence was highly homologous to those of aldo-keto reductase family enzymes. The purified enzyme displayed aldehyde reductase activity in the presence of NADPH. Additionally, it showed ferric reductase activity and drove the Fenton reaction in the presence of free FAD and NADH.

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The utility of the homogeneous assay for fluorescence concentrated on membrane (HAFCOM) in the analysis of the substrate specificity of protease was investigated using tobacco etch virus (TEV) protease. The V_max of TEV protease against variants of a substrate was obtained by a simple procedure. It was considered that HAFCOM was more accurate than other endpoint measurements of protease assay.

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TransDSL is an RNA ligase ribozyme whose enzyme unit joins two RNA fragments constituting a substrate. The enzyme unit recognizes the substrate by means of two clamp modules. We constructed active variants by replacing the original clamp module with various types of interactions. Such flexible modularity would be advantageous in the application of this ribozyme in nanobiotechnology.

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The kinetics of a very large NAD-dependent glutamate dehydrogenase from Janthinobacterium lividum showed positive cooperativity toward α-ketoglutarate and NADH, and the Michaelis-Menten type toward ammonium chloride in the absence of the catalytic activator, L-aspartate. An increase in the maximum activity accompanied the decrease in the S_0.5 values for α-ketoglutarate and NADH with the addition of L-aspartate, and the kinetic response for α-ketoglutarate changed completely to a typical Michaelis-Menten type in the presence of 10 mM L-aspartate.

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The ascorbic acid (AsA) content and the mRNA levels of L-galactose-1-phosphate phosphatase (GPPase), and L-galactono-1,4-lactone dehydrogenase (GalLDH) increased by intense light, and decreased in the dark. Moreover, the promoter regions of GPPase and GalLDH contained light responsible cis-elements. These results suggest that in rice, AsA synthesis is regulated by light.

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The thermo-neutrophilic hydrogen-oxidizing bacterium Hydrogenobacter thermophilus constitutively oxidizes thiosulfate. Soluble extracts of it showed temperature-dependent thiosulfate oxidation activity with an optimum at 60 °C. A gene cluster for sox (sulfur oxidation) is presumably responsible for this activity. Among the cluster, two tandemly coded soxA genes were uniquely found. These results suggest that the Sox system is widespread even in thermo-neutrophilic environments.

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We studied the effects of α-glucosylhesperidin (G-Hsp) on the peripheral body temperature and autonomic nervous system in humans. We first conducted a survey of 97 female university students about excessive sensitivity to the cold; 74% of them replied that they were susceptible or somewhat susceptible to the cold. We subsequently conducted a three-step experiment. In the first experiment, G-Hsp (500 mg) was proven to prevent a decrease in the peripheral body temperature under an ambient temperature of 24 °C. In the second experiment, a warm beverage containing G-Hsp promoted blood circulation and kept the finger temperature higher for a longer time. We finally used a heart-rate variability analysis to study whether G-Hsp changed the autonomic nervous activity. The high-frequency (HF) component tended to be higher, while the ratio of the low-frequency (LF)/HF components tended to be lower after the G-Hsp administration. These results suggest that the mechanism for temperature control by G-Hsp might involve an effect on the autonomic nervous system.

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β-Lactoglobulin (BLG) was conjugated with the N-hydroxysuccinimide ester of the dextran-glycylglycine adduct (DG-ONSu) to reduce the immunogenicity of BLG, a major allergen of cow’s milk, and some immunological properties of the conjugate (DG-BLG) were studied. The conjugate was prepared by modifying BLG with DG-ONSu and purified in a Sephadex G-100 column. The analytical data for DG-BLG indicated that 5.2 moles of DG-ONSu with a mean molecular weight of 9,300 were covalently attached to the amino groups of the BLG molecule. Conjugation with DG-ONSu greatly decreased the reactivity of BLG with anti-BLG antibodies and suppressed their production in vivo due to its shielding action for epitope(s) on the protein’s molecular surface. It was also found that DG-BLG was resistant to proteolytic enzymes. These findings allow us to suggest that DG-ONSu could be advantageously used to suppress the hypersensitivity mediated by IgG antibodies in milk allergy.

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In screening experiments for rat intestinal α-glucosidase (sucrase and maltase) inhibitors in 325 plants cultivated in Japan’s southern island, of Tanegashima, marked inhibition against both sucrase and maltase was found in the extract of the fruit of Solanum torvum. Enzyme-assay guided fractionation of the extract led to the isolation of methyl caffeate (1) as a rat intestinal sucrase and maltase inhibitor. We examined 13 caffeoyl derivatives for sucrase- and maltase-inhibitory activities. The results showed that methyl caffeate (1) had a most favorable structure for both sucrase and maltase inhibition, except for a higher activity of methyl 3,4,5-trihydroxycinnamate (14) against sucrase. Its moderate inhibitory action against α-glucosidase provides a prospect for antidiabetic usage of S. torvum fruit.

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We investigated whether agaro-oligosaccharides have any immunological effects on RAW264.7 mouse macrophages and human monocytes in vitro. We demonstrate that agaro-oligosaccharides suppressed the elevated levels of nitric oxide, prostaglandin E₂, and such pro-inflammatory cytokines as tumor necrosis factor-α, interleukin-1β and interleukin-6 in lipopolysaccharide-stimulated monocytes and macrophages. We also demonstrate that those effects of agaro-oligosaccharides on activated monocytes and macrophages may have been caused by heme oxygenase-1 induction. It is therefore proposed that agaro-oligosaccharides might be a good candidate for a functional food to prevent inflammatory diseases.

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To evaluate the anticarcinogenic activity of methanol extract of Pleurospermum kamtschaticum (PKE), we assessed its apoptosis-inducing capability in HT-29 colon carcinoma cells. PKE treatment for 2 h reduced cell viability in a dose-related manner, and induced apoptotic morphological changes. Flow cytometric analysis indicated that PKE treatment at 0.05 mg/ml induced early apoptosis in 66.2% of HT-29 cells. Additionally, Bcl-2 expression was substantially reduced in PKE-treated HT-29 cells, increasing the Bax/Bcl-2 ratio. The protein levels of procaspase-9 and procaspase-3 were decreased markedly, reflecting caspase-9 and caspase-3 activation, and resulting PARP cleavage was noted in the PKE-treated HT-29 cells. Furthermore, we detected increased NAG-1 expression in the PKE-treated HT-29 cells. In an in vivo study, intraperitoneal PKE administration suppressed the formation of tumor nodules in the lungs of mice. These results indicate that PKE can serve as a beneficial supplement in the treatment and the prevention of colon cancer.

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The uric acid-lowering effect and safety of a fermented barley extract P (FBEP) prepared from barley-shochu distillery by-products were investigated in a randomized, placebo-controlled, parallel-group, double-blinded study. A total of 111 subjects with serum uric acid levels of 6.0–7.9 mg/dl were provided with either a drink containing 2 g/d of FBEP (test group) or a placebo drink. After 12 weeks, the serum uric acid levels changed by −0.21±0.56 mg/dl in the test group, showing a significant decrease in comparison to those of the placebo group (+0.02±0.54 mg/dl). Additionally, the uric acid clearance in the test group showed a tendency to increase after 12 weeks more than in the placebo group (p=0.054). No abnormalities in the physical and clinical tests were observed, and no adverse diagnostic findings were attributed to the intake of the test meal. These results demonstrated the benefits and safety of the FBEP treatment to subjects with slightly high serum uric acid or mild hyperuricemia.

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The odor-active volatiles of Kinokuni mandarin (Citrus kinokuni Hort. ex Tanaka), an original mandarin orange in Japan, were characterized by a combination of instrumental and sensory analyses and compared with those of Satsuma mandarin (Citrus unshiu Marcovitch). An aroma extract dilution analysis (AEDA) of the polar fractions of Kinokuni and Satsuma mandarin peel oils identified five odorants in common as the most odor-active volatiles: (Z)-hex-3-enal, decanal, linalool, yuzuol, and (2E)-trans-4,5-epoxydec-2-enal. In addition, seven odorants were identified solely in Kinokuni mandarin as significant contributors: octanal, dodecanal, (2E,4E)-deca-2,4-dienal, geraniol, yuzunone, (2E,7Z)-trans-4,5-epoxydeca-2,7-dienal, and thymol. The odor-active volatiles in both the non-polar components of the peel oil and an extract of the juice prepared from Kinokuni mandarin were also identified. The (S)-isomer of linalool in Kinokuni mandarin peel oil was dominant in the enantiomeric distribution (92%), whereas the (R)-isomer was dominant in Satsuma mandarin peel oil (90%).

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In this study, we investigated the effects of various temperatures between 140 to 260 °C during sub-critical water extraction of residual green tea in making a roasted green tea-like extract, a popular beverage in Japan. Each residual green tea extract was analyzed for sensory properties and antioxidative activities, and this revealed 140 °C to be the best extraction temperature. 5-Hydroxymethyl-2-furaldehyde, (+)-catechin, and (−)-epicatechin were found to be the major antioxidative compounds in the 140 °C extract, along with theanine and some important amino acids.

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Shewanella sp. HN-41 was exploited for selenium nanoparticles synthesis from aqueous selenite compounds under anaerobic conditions. Various reaction conditions, including reaction time, initial biomass, and initial selenite concentration, were systematically investigated to determine their effects on particle size distribution and formation rate. The biomass concentration of Shewanella sp. HN-41 had no significant effect on average particle size but strongly influenced reduction rate and size distribution. Initial selenite concentration (0.01–1.0 mM) also had no significant effect on the average particle size, but affected the early growth stage mechanism of selenium particle production, which was modeled using a Michaelis Menten model. The HR-TEM and SAED patterns indicated that the synthesized selenium nanoparticles were amorphous.

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The predicted amino acid sequence of Bacillus subtilis yvdO exhibits similarity to that of the proteins belonging to the patatin family of lipolytic enzymes. In the present study, YvdO was overproduced in Escherichia coli and purified, and its enzymatic properties were determined. YvdO hydrolyzed p-nitrophenyl fatty acid esters. The enzyme was specific to middle-chain fatty acids, and its optimum pH was approximately 7.5. It maintained 86% of its initial activity after incubation for 30 min at 80 °C, and its secondary structure was retained at up to 80 °C. Free myristic acid was detected as the product of the reaction with YvdO and 1-myristoly-2-lyso-sn-glycero-3-phosphocholine, while YvdO did not hydrolyze 1,2-dimyristoly-sn-glycero-3-phosphocholine. These results suggest that YvdO is a novel thermostable lipolytic enzyme that has the ability to hydrolyze lysophospholipids.

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Endoglucanase Cel5A from Clostridium josui contains a family 17 carbohydrate-binding module (CBM) (CjCBM17) and a family 28 CBM (CjCBM28) in tandem. These two CBMs bound to non-crystalline cellulose and β-1,3-1,4-glucan. Our results indicate that the CBMs recognized different components on the cell wall of a sweet potato root. The root was cut into longitudinal sections. We used CjCBM17 and CjCBM28 fused to two different fluorescent proteins to visualize differential recognition of the plant cell wall. When they were microscopically observed, CjCBM28-fused cyan fluorescent protein (CFP) differentially bound to the root cap, but CjCBM17-fused blue fluorescent protein (BFP) did not. CjCBM17-BFP bound to the central part or the root apical meristem. These results suggest that CjCBM17 and CjCBM28 recognize different sites of the cell wall and that the cell-wall components and the polysaccharides configuration in the cell wall differ between tissues.

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Pyruvate is the key substance controlling the formation of diacetyl, acetaldehyde, and acetate during alcoholic fermentation. Here we report the breeding of a low pyruvate-producing sake yeast by isolation of a mutant resistant to ethyl α-transcyanocinnamate, an inhibitor of mitochondrial pyruvate transport. Mitochondrial function was involved in resistance to this substance and in the production of pyruvate by the mutants.

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Heat treatment is a simple technique for releasing polyphosphate (polyP) from microbial cells. We investigated the ultrastructural alterations of polyP granules (PPGs) in response to heat treatment at 70 °C using a polyP-accumulating phoU mutant of Escherichia coli as a model organism. Electron microscopic and energy-dispersive X-ray analysis suggested concurrent occurrence of PPG degradation and polyP release in the cytoplasm. PolyP is probably released by cytoplasmic extrusion through breaks in the cell envelope.

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Previously screened CoQ₁₀-overproducing Agrobacterium tumefaciens A603-35 showed a relatively high NADH/NAD⁺ ratio (1.1), as compared to parental strain C58 (0.2) when we increased the expression levels of NADH-generating enzymes. Also, the intracellular NADH/NAD⁺ ratio showed a positive correlation with the CoQ₁₀ content in A603-35. Overexpression of glyceraldehyde 3-phosphate dehydrogenase in A603-35 shifted the NADH/NAD⁺ ratio at 48 h from 0.8 to 1.2, and thus the CoQ₁₀ content in flask culture increased from 2.16 to 3.63 mg/g DCW. Due to the addition of hydroxybutyrate to the culture media, the intracellular NADH/NAD⁺ ratio in A603-35-gapA shifted from 1.2 to 1.4, which led to an increase CoQ₁₀ content (5.27 mg/g DCW).

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