We have recently discovered that a GDSL lipase/esterase (TcGLIP) in Tanacetum cinerariifolium catalyzed acyltransferase activity to form an ester bond in the natural insecticide, pyrethrin. TcGLIP contained Ser40 in Block I, Gly64 in Block II, Asn168 in Block III and Asp318 and His321 in Block V, suggesting underlying hydrolase activity, although little is known about their role in acyltransferase activity. We expressed TcGLIP here in Esherichia coli as a fusion with maltose-binding protein (MBP), part of the fusion being cleaved with a protease to obtain MBP-free TcGLIP. A kinetic analysis revealed that the MBP moiety scarcely influenced the kinetic parameters. The effects on acyltransferase activity of mutations of Gly64, Asn168, Asp318 and His321 were investigated by using MBP-fused TcGLIP. Mutations of these amino acids markedly reduced the acyltransferase activity, suggesting their critical role in the production of pyrethrins.

抄録全体を表示

Eight analogues of (3R,11Z,19Z)-CH503 (3-acetoxy-11,19-octacosadien-1-ol), the anti-aphrodisiac pheromone of male Drosophila melanogaster, were synthesized for a bioassay. These were the enantiomers of 3-acetoxy-11,19-octacosadiyn-1-ol (1), 3-acetoxyoctacosan-1-ol (2), (Z)-3-acetoxy-11-octacosen-1-ol (3), and (Z)-3-acetoxy-19-octacosen-1-ol (4). None of them were pheromonally active, indicating that the two double bonds at C-11 and C-19 were necessary for bioactivity.

抄録全体を表示

A variety of insect species induce galls on host plants. Liquid chromatographic/tandem mass spectrometric analyses showed that a gall midge (Rhopalomyia yomogicola) that induces galls on Artemisia princeps contained high levels of indole-3-acetic acid and cytokinins. The gall midge larvae also synthesized indole-3-acetic acid from tryptophan. Close observation of gall tissue sections indicated that the larval chamber was surrounded by layers of cells having secondary cell walls with extensive lignin deposition, except for the part of the gall that constituted the feeding nutritive tissue which was composed of small cells negatively stained for lignin. The differences between these two types of tissue were confirmed by an expression analysis of the genes involved in the synthesis of the secondary cell wall. Phytohormones may have functioned in maintaining the feeding part of the gall as fresh nutritive tissue. Together with the results in our previous study, those presented here suggest the importance of phytohormones in gall induction.

抄録全体を表示

A new 16-membered macrolide named makinolide B (1) was isolated from Streptomyces sp. MK-19. The structure of makinolide B (1) was determined on the basis of 2D NMR experiments involving DQF-COSY, TOCSY, HSQC, and HMBC methods. Application of the paper disk diffusion method to makinolide B (1) showed weak antibacterial activity against Staphylococcus aureus at the dose of 100 µg/disk.

抄録全体を表示

Fifteen constituents, including a new compound, were isolated from an ethanolic extract of the peel of Citrus kawachiensis Hort. ex. Y. Tanaka (Japanese brand name, kawachibankan) which is one of the citrus products specific to Ehime, Japan. The new compound was characterized as 4'-dihydrophaseic acid β-glucopyranose ester (15) on the basis of spectral and chemical evidence.

抄録全体を表示

Epigallocatechin 3-gallate (EGCG) has cytotoxic effects in many cancer cells. It has been reported that A549 lung cancer cells are markedly resistant to cell death induced by EGCG. In the present study, the effects of EGCG on A549 lung cancer cell growth and angiogenesis were studied. We found that EGCG dose-dependently suppressed A549 cell growth, while A549 cells were markedly resistant to cell death in vitro. Next we found that EGCG increased endostatin expression and suppressed vascular endothelial growth factor (VEGF) expression. We further studied to determine whether EGCG would suppress A549 tumor growth in nude mouse and angiogenesis. EGCG in drinking water significantly suppressed A549 tumor growth in nude mice. Histological analysis revealed that the number of CD34 positive vessels had a tendency to decrease in the tumor. In sum, EGCG had anti-proliferative effects of A549 on tumor growth and showed a tendency to suppress angiogenesis.

抄録全体を表示

If genetically modified organisms (GMOs) are spread through the natural environment, it might affect the natural environment. To help prevent the spread of GMOs, we examined whether it is possible to introduce conditional lethality by excising centromeric DNA from a chromosome by site-specific recombination in Saccharomyces cerevisiae as model organism. First, we constructed haploid cells in which excision of the centromeric DNA from chromosome IV can occur due to recombinase induced by galactose. By this excision, cell death can occur. In diploid cells, cell death can also occur by excision from both homologous chromosomes IV. Furthermore, cell death can occur in the case of chromosome V. A small number of surviving cells appeared with excision of centromeric DNA, and the diploid showed greater viability than the haploid in both chromosomes IV and V. The surviving cells appeared mainly due to deletion of a recombination target site (RS) from the chromosome.

抄録全体を表示

The higher plant ADP-glucose (ADPG) pyrophosphorylase (AGPase), composed of two small subunits and two large subunits (LSs), produces ADPG, the sole substrate for starch biosynthesis from α-D-glucose 1-phosphate and ATP. This enzyme controls a key step in starch synthesis as its catalytic activity is activated by 3-phosphoglycerate (3-PGA) and inhibited by orthophosphate (Pi). Previously, two mutations in the LS of potato AGPase (PLS), PLS-E38K and PLS-G101N, were found to increase sensitivity to 3-PGA activation and tolerance to Pi inhibition. In the present study, the double mutated enzyme (PLS-E38K/G101N) was evaluated. In a complementation assay of ADPG synthesis in an Escherichia coli mutant defective in the synthesis of ADPG, expression of PLS-E38K/G101N mediated higher glycogen production than wild-type potato AGPase (PLS-WT) and the single mutant enzymes, PLS-E38K and PLS-G101N, individually. Purified PLS-E38K/G101N showed higher sensitivity to 3-PGA activation and tolerance to Pi inhibition than PLS-E38K or PLS-G101N. Moreover, the enzyme activities of PLS-E38K, PLS-G101N, and PLS-E38K/G101N were more readily stimulated by other major phosphate-ester metabolites, such as fructose 6-phosphate, fructose 2,6-bisphosphate, and ribose 5-phosphate, than was that of PLS-WT. Hence, although the specific enzyme activities of the LS mutants toward 3-PGA were impaired to some extent by the mutations, our results suggest that their enhanced allosteric regulatory properties and the broadened effector selectivity gained by the same mutations not only offset the lowered enzyme catalytic turnover rates but also increase the net performance of potato AGPase in vivo in view of increased glycogen production in bacterial cells.

抄録全体を表示

In this study, Agrobacterium tumefaciens-mediated transformation (ATMT) was used in breeding industrial strains for the purpose of improving α-transglucosidase production. Firstly, an efficient ATMT system for Asperillus niger was established by optimization of several influencing factors, in which transformation efficiency was improved up to 14-fold compared with the initial conditions. Furthermore, binary vector pBI-Glu containing an α-transglucosidase expression cassette was constructed and transferred into Agrobacterium tumefaciens LBA4404 in order to infect A. niger. By the efficient ATMT method, the gene for α-transglucosidase, driven by strong promoter P_glaA (the glucoamylase gene promoter), had a high expression level in A. niger A-8 (25.02 U/mL). The optimized ATMT system was found to be effective and suitable for A. niger, and should be a useful tool for studying the function of A. niger genes and for industrial breeding of this strain.

抄録全体を表示

α-Amylases (EC 3.2.1.1) hydrolyze internal α-1,4-glucosidic linkages of starch and related glucans. Bacillus sp. AAH-31 produces an alkalophilic thermophilic α-amylase (AmyL) of higher molecular mass, 91 kDa, than typical bacterial α-amylases. In this study, the AmyL gene was cloned to determine its primary structure, and the recombinant enzyme, produced in Escherichia coli, was characterized. AmyL shows no hydrolytic activity towards pullulan, but the central region of AmyL (Gly395-Asp684) was similar to neopullulanase-like α-amylases. In contrast to known neopullulanase-like α-amylases, the N-terminal region (Gln29-Phe102) of AmyL was similar to carbohydrate-binding module family 20 (CBM20), which is involved in the binding of enzymes to starch granules. Recombinant AmyL showed more than 95% of its maximum activity in a pH range of 8.2–10.5, and was stable below 65 °C and from pH 6.4 to 11.9. The k_cat values for soluble starch, γ-cyclodextrin, and maltotriose were 103 s⁻¹, 67.6 s⁻¹, and 5.33 s⁻¹, respectively, and the K_m values were 0.100 mg/mL, 0.348 mM, and 2.06 mM, respectively. Recombinant AmyL did not bind to starch granules. But the substitution of Trp45 and Trp84, conserved in site 1 of CBM20, with Ala reduced affinity to soluble starch, while the mutations did not affect affinity for oligosaccharides. Substitution of Trp61, conserved in site 2 of CBM20, with Ala enhanced hydrolytic activity towards soluble starch, indicating that site 2 of AmyL does not contribute to binding to soluble long-chain substrates.

抄録全体を表示

Chlamydomonas reinhardtii cells are surrounded by a mixture of hydroxyprolin-rich glycoproteins consisting of L-arabinose, D-galactose, D-glucose, and D-mannose residues. The L-arabinose residue is thought to be attached by a transfer of UDP-L-arabinofuranose (UDP-Araf), which is produced from UDP-L-arabinopyranose (UDP-Arap) by UDP-arabinopyranose mutase (UAM). UAM was purified from the cytosol to determine the involvement of C. reinhardtii UAM (CrUAM) in glycoprotein synthesis. CrUAM was purified 94-fold to electrophoretic homogeneity by hydrophobic and size-exclusion chromatography. CrUAM catalyzed the reversible conversion between UDP-Arap and UDP-Araf and exhibited autoglycosylation activity when UDP-D-[¹⁴C]glucose was added as substrate. Compared to the properties of native and recombinant CrUAM overexpressed in Escherichia coli, native CrUAM showed a higher affinity for UDP-Arap than recombinant CrUAM did. This increased affinity for UDP-Arap might have been caused by post-translational modifications that occur in eukaryotes but not in prokaryotes.

抄録全体を表示

We purified both the type I subunit and type II subunit of porcine hair keratin and compared their ability to form a uniform film of reconstituted keratin on a culture plate, and their effect on a model of neural cells. We observed the surface of the keratin-immobilized plate using a scanning electron microscope (SEM) and measured water contact angles to characterize the surface. We cultured PC12 cells on plates on which crude keratin, the type I subunit, or the type II subunit were immobilized. The water contact angles were slightly different from each other. The cells proliferated well on all three keratin-immobilized plates. The type II subunit showed a tendency to inhibit the differentiation of PC12 cells significantly as an extension of the cell shapes and neurite outgrowth in comparison with the crude extract and the type I subunit. The type I subunit and the type II subunit showed slight differences in cell differentiation, but not in cell proliferation.

抄録全体を表示

Most zinc metalloproteinases have the consensus zinc-binding motif sequence HEXXH, in which two histidine residues chelate a catalytic zinc ion. The zinc-binding motif sequence of thermolysin, H¹⁴²ELTH¹⁴⁶, belongs to this motif sequence, while that of dipeptidyl peptidase III (DPP III), H⁴⁵⁰ELLGH⁴⁵⁵, belongs to the motif sequence HEXXXH. In this study, we examined effects of conversion of HEXXH to HEXXXH in thermolysin on its activity and stability. Thermolysin variants bearing H¹⁴²ELLGH¹⁴⁶ or H¹⁴²ELTGH¹⁴⁶ (designated T145LG and T145TG respectively) were constructed by site-directed mutagenesis and were produced in Escherichia coli cells by co-expressing the mature and pro domains separately. They did not exhibit hydrolyzing activity for casein or N-[3-(2-furyl)acryloyl]-glycyl-L-leucine amide, but exhibited binding ability to a substrate analog glycyl-D-phenylalanine (Gly-D-Phe). The apparent denaturing temperatures based on the ellipticity at 222 nm of T145LG and T145TG were 85 ± 1 °C and 86 ± 1 °C respectively, almost the same as that of wild-type thermolysin (85 ± 1 °C). These results indicate that conversion of HEXXH to HEXXXH abolishes thermolysin activity, but does not affect its binding ability to Gly-D-Phe or its stability. Our results are in contrast to ones reported previously, that DPP III variants bearing H⁴⁵⁰ELGH⁴⁵⁵ exhibit activity.

抄録全体を表示

Cell-surface expression of the discoidin domain receptor (DDR) tyrosine kinase family in high molecular mass form was controlled sensitively by the glucose concentration through a post-translational N-glycosylation process. Cycloheximide time-course experiments revealed that the high-molecular-mass forms of DDR1 and DDR2 were significantly less stable than control receptor tyrosine kinases. Site-directed mutational analysis of the consensus N-glycosylation sites of the DDRs revealed that mutations of asparagine 213 of DDR2 and asparagine 211 of DDR1, a conserved N-glycosylation site among vertebrate DDRs, inhibited the generation of the high-molecular-mass isoform. Taken together, these results suggest a mechanism to control the activity of the DDR family by regulating its cell-surface expression. Due to low stability, the steady-state population of functional DDR proteins in the cell surface depends sensitively on its maturation process via post-translational N-glycosylation, which is controlled by the glucose supply and the presence of a conserved N-glycosylation site.

抄録全体を表示

A Δ6-desaturase gene was isolated from Microula sikkimensis. Sequence analysis indicated that the gene, designated MsD6DES, had an open reading frame of 1,357 bp and encoded 448 amino acids. Heterologous expression in tobacco indicated that MsD6DES can use endogenous substrates to synthesize γ-linolenic acid (GLA, 18:3^{Δ 6,9,12}) and octadecatetraenoic acid (OTA, 18:4^{Δ 6,9,12,15}). MsD6DES transcripts were distributed in all tested tissues, with high expression levels in seeds and young leaves. The effects of temperature and wounding stresses on MsD6DES expression were analyzed. The results indicated that temperature regulates MsD6DES at the transcriptional level. MsD6DES expression increased first, reaching a maximum 4 h after low-temperature treatment. A slight increase in MsD6DES transcript levels was also observed under high temperature. We found that the response of MsD6DES to temperature stress was different from those of fungi and algae. In addition, MsD6DES was found to be wound-inducible.

抄録全体を表示

Phospholipase D (PLD)-mediated transphosphatidylation of phosphatidylcholine (PC) in a biphasic system was limited by the hydrolysis reaction. A biphasic system can produce a large amount of water. To solve this problem, a microaqueous water-immiscible organic solvent was used for the first time in the bioconversion of phosphatidylserine (PS). The transphosphatidylation among 40 µmol PC, 800 µmol L-serine, and 0.17 U/mL PLD in 2.133 mL of butyl acetate with 6.25% water (V/V) was conducted at a trans-phosphatidylation rate of 88% (mol/mol), and no hydrolytic reaction was observed. Compared to commonly used biphasic systems, this system shows a similar transphosphatidylation rate, whereas the undesirable hydrolysis of phospholipids was completely suppressed.

抄録全体を表示

1,3-β-D-Glucan phosphorylase (BGP) is an enzyme that catalyzes the reversible phosphorolysis of 1,3-β-glucosidic linkages to form α-D-glucose 1-phosphate (G1P). Here we report on the purification and characterization of BGP from Ochromonas danica (OdBGP). The purified enzyme preparation showed three bands (113, 118, and 124 kDa) on SDS-polyacrylamide gel electrophoresis. The optimum pH and temperature were 5.5 and 25 °C–30 °C. OdBGP phosphorolysed laminaritriose, larger laminarioligosaccharides, and laminarin, but not laminaribiose. In the synthesis reaction, laminarin and laminarioligosaccharides served as good acceptors, but OdBGP did not act on glucose. Kinetic analysis indicated that the phosphorolysis reaction of OdBGP follows a sequential Bi Bi mechanism. The equilibrium of the enzymatic reaction indicated that OdBGP favors the reaction in the synthetic direction. Overnight incubation of OdBGP with laminaribiose and G1P resulted in the formation of precipitates, which were probably 1,3-β-glucans.

抄録全体を表示

We investigated the effects of cAMP on the basolateral Na⁺, K⁺-ATPase and apical amiloride-sensitive Na⁺ channel in the rat distal colon. Under mucosal treatment with nystatin, isoproterenol and forskolin increased the basolateral Na⁺ pump current. Under serosal treatment with ouabain, both agents slightly increased the amiloride-sensitive Na current, but not to a significant level. We concluded that cAMP activates the basolateral Na⁺, K⁺-ATPase in the rat distal colon.

抄録全体を表示

Octaphlorethol A is a phenolic compound isolated from the marine alga Ishige foliacea. In the present study, we investigated the anti-inflammatory activity of octaphlorethol A in CpG-stimulated primary murine bone marrow-derived macrophages and dendritic cells. It exhibited anti-inflammatory activity by regulating the mitogen-activated protein kinase and NF-κB pathways.

抄録全体を表示

The β-xylosidase, which is active against plant complex type N-glycans, was purified to homogeneity from Ginkgo biloba seeds. The N-terminal amino acid sequence, G-S-A-A-G-N-R-, of the Ginkgo β-xylosidase (β-Xyl'ase Gb) was consistent with the deduced internal amino acid sequence of an Arabidopsis β-xylosidase (AtBXL1). β-Xyl'ase Gb hydrolyzed the β1-2 xylosyl residue from Xylβ1-2Manβ1-4GlcNAcβ1-4GlcNAc-PA and Xylβ1-2Manβ1-4GlcNAcβ1-4(Fucα1-3)GlcNAc-PA, but not that from Manα1-6(Manα1-3)(Xylβ1-2)Manβ1-4GlcNAcβ1-4(Fucα1-3)GlcNAc-PA.

抄録全体を表示

Gamma irradiation increased catalase activities at 0.1 kGy and decreased them at 10 kGy in Arabidopsis wild type and catalase-deficient mutants, cat3-1 and cat1 cat3. Irradiation induced DNA damage, H₂O₂ accumulation, and lipid peroxidation in both mutants as well as the wild type. Thus catalases might not be key enzymes protecting gamma irradiation-induced damage.

抄録全体を表示

A vacuolar membrane protein, Vba2p of Schizosaccharomyces pombe, is involved in basic amino acid uptake by intact cells. Here we found evidence that Vba2p mediated ATP-dependent lysine uptake by vacuolar membrane vesicles of Saccharomyces cerevisiae. Vba2p was also responsible for quinidine sensitivity, and the addition of lysine improved cell growth on quinidine-containing media. These findings should be useful for further characterization of Vba2p.

抄録全体を表示

Bunching onion [Allium fistulosum L. (Liliaceae)] secretes mucus in the cavities of its green leaves. The effects of the mucus, which is consumed as food, were examined. The mucus augmented the production of tumor necrosis factor (TNF)-α and monocyte chemotactic protein (MCP)-1 from RAW 264 cells and of interleukin (IL)-12 from J774.1 cells; however, extracts from green leaves and white sheaths did not. An oral administration of this mucus to mice augmented the immune functions of peritoneal cells by increasing TNF-α and IL-12 production and phagocytosis. It also augmented interferon (IFN)-γ production from spleen cells and natural killer (NK) activity. These results suggest that an oral administration of the A. fistulosum mucus can enhance natural immunity.

抄録全体を表示

This study shows that the ECM degradation-associated pathway, including uPA and tPA and the downstream MMP-2/-9 protein, was significantly suppressed in HA22T cells treated with a Zanthoxylum avicennae extract (YBBE). The endogenous inhibitors, including TIMP-1/-2 and PAI-1, were enhanced in HA22T cells by the YBBE treatment. The expression of MMP-2/-9 and TIMP-1/-2 was respectively assessed by using RT-PCR and a zymography assay. The mRNA levels and enzymatic activity of MMP-2/-9 were down-regulated by the YBBE treatment in a dose-dependent manner, while the TIMP-1/-2 levels were conversely markedly increased. The PP2A siRNA or PP2A inhibitor totally reversed the YBBE effects, confirming the essential role of PP2A in YBBE inhibiting the HA22T cell migration and invasion effects. Xenografted animal experiments on nude mice demonstrated similiar results to the in vitro system. Both the in vitro and in vivo models clearly demonstrate that YBBE inhibited the highly metastatic HA22T liver cancer cell migration and invasion effects through PP2A activation.

抄録全体を表示

Lactic acid bacteria have been reported to have various immune-regulating activities. We also found in the previous study that the oral administration of heat-killed Lactobacillus plantarum NRIC0380 induced CD4⁺CD25⁺Foxp3⁺ cells (Treg cells). We examine in this present study the influence of NRIC0380 on the function of intestinal dendritic cells (DCs) in vitro and in vivo. The aldehyde dehydrogenase (ALDH) activity was significantly induced in DCs obtained from the mesenteric lymph node (MLN) by culturing with NRIC0380. The oral administration of NRIC0380 also significantly increased ALDH-positive DCs in MLN. NRIC0380 significantly enhanced the production of TGF-β from MLN cells in vitro. These effects were not apparent in cells from the Peyer's patch (PP) and spleen (SPL). NRIC0380 also significantly enhanced the expression of B7-H1 on DCs of all organs in vitro. The effects of NRIC0380 on DCs, especially those located in MLN, might be involved in its function to induce Treg cells.

抄録全体を表示

We have reported an inhibitory effect of Lactobacillus gasseri OLL2809 (OLL2809) on the growth of mouse endometrial tissue in the abdominal cavity. The present study aimed to investigate the efficacy of Lactobacillus gasseri OLL2809 (OLL2809) on pre-existing endometriosis implanted on the abdominal wall in diestrus Wistar-Imamichi female rats. One week after implantation, the volume of the endometrial tissue was measured after laparotomy. OLL2809 and dienogest were administered for 4 weeks. OLL2809 significantly enhanced the decrease in the volume (p<0.01) as compared with control. Complete healing was observed in two of nine rats, but in none of the control group. Dienogest did not show significant efficacy. These findings suggest that OLL2809 is useful not only in therapy of pre-existing endometriosis but also in the prevention of the growth of endometrial tissue.

抄録全体を表示

The beneficial effects of dietary glucosylceramide on the barrier function of the skin have been increasingly reported, but the entire mechanism has not been clarified. By DNA microarray, we investigated changes in gene expression in hairless mouse skin when a damage-inducing AD diet and a glucosylceramide diet (GluCer) were imposed. GluCer administration potentially suppressed the upregulation of six genes and the downregulation of four genes in the AD group. Examination of the epidermal and/or dermal expression of Npr3, Cyp17a1, Col1a1, S100a9, Sprr2f, Apol7a, Tppp, and Scd3 revealed responses of various parts of the skin to the diets. In normal hairless mice, GluCer administration induced an increase in the dermal expression of Cyp17a1 and the epidermal expression of Tppp, and a decrease in the epidermal expression of S100a9. Our results provide information on gene expression not only in whole skin but also in the epidermis and dermis that should prove useful in the search for the mechanisms underlying the effects of GluCer on damaged and normal skin.

抄録全体を表示

The effects of 16 lectins isolated from foodstuff on the transport system across human intestinal Caco-2 cell monolayers were investigated by using four fluorescent markers: lucifer yellow (LY) for the paracellular pathway, fluorescein (FL) for the monocarboxylic acid transporter-mediated pathway, rhodamine 123 for the P-glycoprotein-mediated efflux pathway, and calcein for the multidrug resistance associated protein-related efflux pathway. The transepithelial electrical resistance (TER) values for the monolayers were also measured. WGA from wheat germ, ABA from white mushroom, AOL from Aspergillus oryzae, and CSL3 from chum salmon eggs (each at 100 µg/mL) decreased the TER value by 20–40% which resulted in increased LY transport. These lectins, as well as such other lectins as SBA from soybean, RBA from rice bran, and Con A from jack bean, affected other transport pathways too. These results indicate that the lectins modulated the transepithelial transport system in different ways, probably because of their specific binding characteristics toward Caco-2 cell monolayers.

抄録全体を表示

Sparassis crispa (SC) is an edible mushroom with various medicinal properties. In this study, we investigated to determine whether SC would affect skin conditions in rats and humans. Oral administration of SC increased both turnover of the stratum corneum and dermal soluble collagen content in collagen synthetic activity-reduced model rats. To investigate the effects of oral intake of SC in humans, we performed a randomized, double-blind, placebo-controlled study. We found that cheek transepidermal water loss was significantly lower in the experimental group than in the control group at 4 weeks of ingestion. This study suggests that SC is effective and safe for the improvement of skin conditions.

抄録全体を表示

Tea catechins have strong bitterness and influence the taste of tea. Among the 25 human bitter-taste receptors (TAS2Rs), we found that hTAS2R14 responded to catechins, in addition to hTAS2R39, a known catechin receptor. Although hTAS2R14 responded to (−)-epicatechin gallate and (−)-epigallocatechin gallate, it did not respond to (−)-epicatechin and (−)-epigallocatechin.

抄録全体を表示

Lactococcus lactis IO-1 (JCM7638) produces L-lactic acid predominantly when grown at high xylose concentrations, and its utilization is highly desired in the green plastics industry. Therefore it is worthwhile studying its genomic traits. In this study, we focused on (i) genes of possible horizontal transfer derivation (prophages, the nisin-sucrose transposon, and several restriction-modification systems), and (ii) genes for the synthetic pathways of amino acids and vitamins in the IO-1 genome. In view of the results of this analysis, we consider their meanings in strain IO-1.

抄録全体を表示

Glutaminase, an enzyme that catalyzes the conversion of L-glutamine to L-glutamate, enhances the umami taste in soy sauce. The Aspergillus sojae genome contains 10 glutaminase genes. In this study, we estimated that approximately 60% of the glutamate in soy sauce is produced through the glutaminase reaction. To determine which glutaminase is involved in soy sauce glutamate production, we prepared soy sauces using single and multiple glutaminase gene disruptants of A. sojae. The glutamate concentration in soy sauce prepared using the ΔgahA-ΔgahB-ΔggtA-Δgls disruptant was approximately 60% lower than that in the control strain, whereas it was decreased by approximately 20–30% in the ΔgahA-ΔgahB disruptant. However, the glutamate concentration was unchanged in the soy sauces prepared using the ΔgahA-ΔggtA-Δgls and ΔgahB-ΔggtA-Δgls disruptants. These results indicate that four glutaminases are involved in glutamate production in soy sauce, and that the peptidoglutaminase activities of GahA and GahB increase the glutamate concentration in soy sauce.

抄録全体を表示

Torulaspora delbrueckii metabolism was assessed in a synthetic culture medium similar to grape must under various conditions: no aeration and three different oxygen feeds, in order to determine the effect of oxygen on metabolism. Carbon and nitrogen mass balances were calculated to quantify metabolic fluxes. The effect of oxygen was to decrease the flux of carbon going into the fermentation pathway in favor of growth. In the absence of aeration, higher amounts of glycerol were produced, probably to maintain the redox balance. The oxygen requirement of this strain was high, since even for the highest air supply oxygen became limiting after 24 h. Nevertheless, this strain developed well in the absence of oxygen and consumed 220 g/L of sugars (glucose/fructose) in 166 h at 20 °C, giving a good ethanol yield (0.50 g/g).

抄録全体を表示

Nucleoside diphosphates linked to moiety X (Nudix) hydrolase functions were investigated in hypoxic Aspergillus nidulans cells. Among three nudix hydrolase isozymes, NdxA transcription was up-regulated under oxygen (O₂)-limited conditions. A gene disruptant of the NdxA-encoding gene (NdxAΔ) accumulated more NADH and ADP-ribose than the wild type (WT) under the same conditions. These results indicate that NdxA hydrolyzes these nucleotides in hypoxic fungal cells, which accords with the thesis that NdxA hydrolyzes NADH and ADP-ribose. Under O₂-limited conditions, NdxAΔ decreased glucose consumption, the production of ethanol and lactate, cellular ATP levels, and growth as compared with WT. WT cultured under hypoxia converted exogenously added fructose 1,6-bisphophate, a glycolytic intermediate, to glyceraldehyde 3-phosphate (GAP). The hypoxic NdxAΔ cells accumulated 3.0- to 4.2-fold more GAP than WT under the same conditions, indicating that NdxA increased GAP oxidation by a glycolytic mechanism. Steady-state kinetics indicated that NADH and ADP-ribose competitively inhibited fungal GAP dehydrogenase (GAPDH) with K_i values of 34- and 55-µM, respectively. These results indicate that NdxA hydrolyzes cellular NADH- and ADP-ribose, derepresses GAPDH activity, and hence up-regulates glycolysis in hypoxic A. nidulans cells. That NdxAΔ consumed less pyruvate and tricarboxylate cycle intermediates than WT suggests that NdxA-dependent hydrolysis of nucleotides controls the catabolism of these carbon sources under O₂-limited conditions.

抄録全体を表示

Trichothecene 3-O-acetyltransferase (TRI101) is an indispensable enzyme for the biosynthesis of trichothecenes, a group of mycotoxins produced by Fusarium graminearum. In this study, an inhibitor of TRI101 was identified by chemical array analysis using compounds from the RIKEN Natural Products Depository (NPDepo) library. Although the addition of the identified enzyme inhibitor to the fungal culture did not inhibit trichothecene production, it can serve as a candidate lead compound in the development of a mycotoxin inhibitor that inactivates fungal defense mechanisms.

抄録全体を表示

We examined in vitro the adhesion of Enterococcus faecium NHRD IHARA (NHRD IHARA) to porcine small intestinal mucin (PSIM) and inhibition of the adherence of enteropathogenic bacteria due to pre-incubation of PSIM with NHRD IHARA. NHRD IHARA exhibited an effective barrier function in porcine small intestinal mucus layer.

抄録全体を表示