Electrolyzed anode water (EAW) markedly extended the vase life of cut carnation flowers. Therefore, a flower-senescence hormone involving ethylene decomposition by EAW with potassium chloride as an electrolyte was investigated. Ethylene was added externally to EAW, and the reaction between ethylene and the available chlorine in EAW was examined. EAW had a low pH value (2.5), a high concentration of dissolved oxygen, and extremely high redox potential (19.2 mg/l and 1323 mV, respectively) when available chlorine was at a concentration of about 620 μM. The addition of ethylene to EAW led to ethylene decomposition, and an equimolar amount of ethylene chlorohydrine with available chlorine was produced. The ethylene chlorohydrine production was greatly affected by the pH value (pH 2.5, 5.0 and 10.0 were tested), and was faster in an acidic solution. Ethylene chlorohydrine was not produced after ethylene had been added to EAW at pH 2.6 when available chlorine was absent, but was produced after potassium hypochlorite had been added to such EAW. The effect of the pH value of EAW on the vase life of cut carnations was compatible with the decomposition rate of ethylene in EAW of the same pH value.
  These results suggest that the effect of EAW on the vase life of cut carnations was due to the decomposition of ethylene to ethylene chlorohydrine by chlorine from chlorine compounds.

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  The mechanical properties of pullulan films prepared at various temperatures were investigated. The films prepared at high temperatures (40°C and 60°C; H-films) did not show any clear plastic deformation in tensile test, indicating that they were brittle. In contrast, those prepared at low temperatures (4°C, 13°C, and 25°C; L-films) showed such deformation. The latter films had higher values for both tensile strength and elastic modulus than the former, indicating that the L-films were stiffer and more flexible than the H-films. Stretching the L-films clearly showed a shear deformation band inclined at 45° to the stretching direction, indicating that they were amorphous.

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  Hexadecadien-1-ol and the derivatives (acetate and aldehyde) with a conjugated diene system have recently been identified from a pheromone gland extract of the persimmon fruit moth (Stathmopoda masinissa), a pest insect of persimmon fruits distributed in East Asia. The alcohol and acetate showed their base peaks at m/z 79 in a GC-MS analysis by electron impact ionization, but the aldehyde produced a unique base peak at m/z 84, suggesting a 4,6-diene structure. To confirm this inference, four geometrical isomers of each 4,6-hexadecadienyl compound were synthesized by two different routes in which one of two double bonds was furnished in a highly stereoselective manner. Separation of the two isomers synthesized together by each route was facilely accomplished by preparative HPLC. Their mass spectra coincided well with those of natural components, indicating that they were available for use as authentic standards for determining the configuration of the natural pheromone. Furthermore, other hexadecadienyl compounds, including the conjugated diene system between the 3- and 10-positions, were synthesized to accumulate the spectral data of pheromone candidates. 5,7-Hexadecadienal interestingly showed the base peak at m/z 80; meanwhile, the base peaks of its alcohol and acetate were detected at m/z 79 like the corresponding 4,6-dienes. The base peaks of all 6,8-, 7,9-, and 8,10-dienes universally appeared at m/z 67 like 9,11-, 10,12-, and 13,15-dienes, the spectra of which have already been published. Although 3,5-hexadecadienal was not prepared, base peaks at m/z 67 and 79 were recorded for the alcohol and acetate, respectively.

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   The 4-oxofurofuran lignan, (+)-aptosimon (1), was synthesized from γ-butyrolactone (9). To construct the two benzylic chiral center of (+)-aptosimon (1), highly erythro selective aldol condensation and stereoconvergent SN1 intramolecular cyclization were used as the key reactions.

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  Neocurdione, isoprocurcumenol, and a new sesquiterpene, 9-oxo-neoprocurcumenol, were isolated from fresh rhizomes of Curcuma aromatica and Curcuma zedoaria (Zingiberaceae) as attachment inhibitors against the blue mussel, Mytilus edulis galloprovincialis. The structures of these compounds were elucidated on the basis of spectroscopic evidence.

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  Chitinase cDNAs from Leucaena leucocephala seedlings were cloned by PCR amplification with degenerate primers based on conserved class I chitinase sequences and cDNA library screening. Two closely related chitinase cDNAs were sequenced and inferred to encode precursor proteins of 323 (KB1) and 326 (KB2) amino acids. Expression of the KB2 chitinase from a pET32a plasmid in Origami (DE3) Escherichia coli produced high chitinase activity in the cell lysate. The recombinant thioredoxin fusion protein was purified and cleaved to yield a 32-kDa chitinase. The recombinant chitinase hydrolyzed colloidal chitin with endochitinase-type activity. It also inhibited growth of 13 of the 14 fungal strains tested.

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  It has been reported that Pseudomonas putida 9816 is able to grow on methanol, but it does not have methanol dehydrogenase or oxidase activity. To utilize methanol it requires yeast extract. The utilization of methanol can be accelerated by adding formate, which obviously helps oxidize methanol and win biologically useful energy. This pseudo-oxidation is catalyzed by a reverse formaldehyde dismutase. Thus, methanol can be both assimilated and dissimilated. Formate alone cannot replace yeast extract. The strain is auxotrophic.

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  A ribonuclease inhibitory activity was detected in the fruits of common apple, Malus x domestica, cv. Fuji, and purified by affinity chromatography on ribonuclease A-Sepharose. It inhibited hydrolysis of cyclic-2′:3′-CMP by bovine pancreatic ribonuclease A with an apparent inhibition constant of about 5×10^-8 M. Matrix-assisted laser desorption/ionization time-of-flight mass spectrometry of the purified protein gave two peaks corresponding to the mass numbers of 55,658 and 62,839, while three bands of 43-, 34-, and 21-kDa were detected by SDS-PAGE. These results suggested that the inhibitor preparation was a mixture of two proteins comprised of 43- and 21-kDa subunits or of 34- and 21-kDa subunits. Attempts to separate these two proteins were unsuccessful. Amino acid composition and N-terminal amino acid sequence of these subunits were also identified and N-terminal sequences showed some similarity to that of cottonseed storage globulin. The significance of the presence of ribonuclease inhibitors in apple fruits is not clear, but it might allow some speculation about their possible involvement in the control of the self-incompatibility ribonuclease of Rosaceae plants.

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  We demonstrated that Mortierella alpina 1S-4 has two Δ6-desaturases, which are involved in the desaturation of linoleic acid to γ-linolenic acid. For one of the two Δ6-desaturases, designated as Δ6I, gene cloning and its heterologous expression in a fungus, Aspergillus oryzae, has previously been reported. In addition, we indicated in this paper that there is an isozyme of the two Δ6-desaturases, designated as Δ6II, in M. alpina 1S-4. The predicted amino acid sequences of the Mortierella Δ6-desaturases were similar to those of ones from other organisms, i.e. borage and Caenorhabditis elegans, and had a cytochrome b₅-like domain at the N-terminus, being different from the yeast Δ9-desaturase, which has the corresponding domain at the C-terminus. The full-length Δ6II cDNA was expressed in A. oryzae, resulting in the accumulation of γ-linolenic acid (which was not detected in the control Aspergillus) up to 37% of the total fatty acids. The analysis of real-time quantitative PCR (RTQ-PCR) showed that the quantity of Δ6I RNA was 2.4-, 9-, and 17-fold higher than that of Δ6II RNA on 2, 3, and 4 days in M. alpina 1S-4, respectively. M. alpina 1S-4 is the first fungus to be confirmed to have two functional Δ6-desaturase genes.

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  Ancylobacter aquaticus strain KNK607M, which had high NAD-dependent formate dehydrogenase (FDH) activity, was newly isolated. The enzyme, purified to homogeneity, was a dimer composed of identical subunits with a molecular mass of 44 kDa. The specific activity was 9.5 u/mg, and the enzyme was optimum at pH 6.3 and 50°C, most stable at pH 7.0, and stable at 50°C or lower. The apparent K_m values for formate and NAD⁺ were 2.4 and 0.057 mM, respectively. The enzyme was specific to formate and was inhibited by SH reagents and heavy metal ions. The cloned gene of FDH contained one open reading frame (ORF) of 1206 base pairs, predicted to encode a polypeptide of 401 amino acids, with a calculated molecular weight of 43,895; this gene was highly expressed in E. coli cells. The FDH had high identity to other FDHs, i.e., those of Pseudomonas, Mycobacterium, Moraxella, and Paracoccus, which were 91.3%, 90.8%, 84.2%, and 82.3%, respectively.

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  We purified and characterized an aminopeptidase from Streptococcus thermophilus YRC001 to obtain an enzyme for the application of reducing bitter-defect in cheese manufacturing. The purified enzyme was a monomer, and its molecular mass was estimated to be 90-100 kDa. It had a broad substrate specificity, and mostly hydrolyzed lysyl and leucyl peptides. The optimal temperature and pH for the enzyme were 35°C and pH 6.5, respectively. EDTA, o-phenanthroline, and p-chloromercuribenzoate inhibited its activity, therefore it was considered to be a metallopeptidase. The purified enzyme efficiently reduced the bitterness of a trypsin digest of reconstituted skim milk. Therefore, we cloned a gene for the enzyme from YRC001. The nucleotide sequence of a 2,940-bp XbaI fragment containing the gene was analyzed. The gene encoded 849 amino acids, and the calculated molecular mass for the mature enzyme (initial methionine is removed) was 96,434. The deduced amino acid sequence showed high homology with the known bacterial lysyl aminopeptidase (aminopeptidase N).

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  We have previously reported that transforming growth factor β (TGF-β) triggers two independent senescence programs, 1) replicative senescence dependent upon telomere shortening and 2) premature senescence independent of telomere shortening, in the cell line of A549 human lung adenocarcinoma. In this study, we examined the possibility that cancer cell tumor phenotypes could be suppressed by forced senescence. We used A549 cells treated with TGF-β for a long time (over 50 days), where senescence was induced in a telomere-shortening-dependent or an independent way. Fully senescent A549 cells were elongated, acquired contact inhibition capabilities when reaching confluence, and secreted the senescence-associated cytokine IL-6. Furthermore, senescent A549 cells had no tumorigenicity in nude mice. These results indicate that the forced induction of senescence in cancer cells may be a novel and potentially powerful method for advancing anti-cancer therapy.

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  Chitinase C (ChiC) is the first bacterial family 19 chitinase discovered in Streptomyces griseus HUT6037. In vitro, ChiC clearly inhibited hyphal extension of Trichoderma reesei but a rice family 19 chitinase did not. In order to investigate the effects of ChiC as an increaser of plant resistance to fungal diseases, the chiC gene was introduced into rice plants under the control of the increased CaMV 35S promoter and a signal sequence from the rice chitinase gene. Transgenic plants were morphologically normal. Resistance to leaf blast disease caused by Magnaporthe grisea was evaluated in R₁ and R₂ generations using a spray method. Ninety percent of transgenic rice plants expressing ChiC had higher resistance than non-transgenic plants. Disease resistance of sibling plants within the same line was correlated with the ChiC expression levels. ChiC produced in rice plants accumulated intercellularly and had the hydrolyzing activity against glycol chitin.

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  Hairy roots of goldenrod (Solidago altissima L.) were induced by infecting axenic plants with Agrobacterium rhizogenes strain A4. Growth and allelopathic polyacetylene (cis-dehydromatricaria ester, cis-DME) production of two independent hairy root clones were examined in several culture media and light regimes. cis-DME contents in hairy roots were at the same level as those in normal roots. cis-DME production in root cultures was several-fold lower than that of native plants and greatly repressed by light.

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  A histidine-tagged G protein of bacteriophage φX174 (HisG) bound strongly with lipopolysaccharide (LPS) of Escherichia coli C, one of a φX174-sensitive Ra strain. The dissociation constant, K_d, was measured to be 0.16±0.04 μM by fluorometric titration. HisG showed slightly less affinity to LPSs of the insensitive Rc and Rd₂ strains having shorter R-core polysaccharide sequences than that of the sensitive Ra strains. The difference between the two types of LPS was demonstrated by CD spectra; LPSs of the sensitive strains increased the signal intensity for β-sheet, while the insensitive strains decreased it. The chemically degraded LPS derivatives lacking a hydrophobic lipid region showed much less affinity to HisG, indicating the importance of the lipid region of LPS for strong binding with HisG. On the other hand, since the degraded derivatives increased the intensity of CD spectra, the polysaccharide region is thought to contribute to the conformation change of the protein.

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  A fibrinolytic protease was purified from an Oriental medicinal herb, Spirodela polyrhiza (Choi, H. S., et al., Biosci. Biotechnol. Biochem., 65, 781-786 (2001)). The protease hydrolyzed not only fibrin but also fibrinogen. The enzyme had an anticoagulant activity measured with activated partial thromboplastin time, thrombin time, and prothrombin time in rat plasma. It doubled all three at 69, 29, and 221 nM, respectively. The protein had anticoagulant activity when given intravenously and orally. The maximum delay in the activated partial thromboplastin time was at the dose of 0.52 and 4.2 mg/kg for intravenous and oral administration, respectively. This protein may be useful in clinical applications for anticoagulation.

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  Serine (Ser) decarboxylase (SDC) catalyzes the conversion of Ser to ethanolamine (EA) in plants, while the physiological implications of the enzyme activity remain elusive. Here, we report that SDC gene expression in Arabidopsis was greatly induced by treatments with Ni²⁺ (24-fold) and Mn²⁺ (4-fold), and discuss possible genetic engineering strategies using the SDC gene for environmental stress management.

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  In addition to momilactone, phytocassanes A through E (diterpene phytoalexins) were detected in rice leaves in fields suffering from rice blast. Furthermore, phytocassane accumulation was most abundant at the edges of necrotic lesions, indicating that the phytoalexins prevent subsequent spread of the fungus from the infected site. In pot experiments the pattern of phytocassane accumulation in rice leaves in an incompatible interaction (infection with an avirulent race of Magnaporthe grisea) was more rapidly induced than in a compatible interaction (infection with a virulent race of M. grisea).

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  The antimutagenic activity of four isoflavones isolated from soybean miso toward three kinds of mutagens, AF-2, MNNG, and Trp-P-1, was evaluated by the Ames test. 8-Hydroxyisoflavones had greater suppressive potency than that of daidzein, and 6-hydroxydaidzein had almost the same activity as daidzein. These results indicated the number of hydroxy and methoxy groups and the position of these functional groups were important for antimutagenic activity.

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  Trichothecene 3-O-acetyltransferase (encoded by Tri101) inactivates the virulence factor of the cereal pathogen Fusarium graminearum. Zearalenone hydrolase (encoded by zhd101) detoxifies the oestrogenic mycotoxin produced by the same pathogen. These genes were introduced into a model monocotyledon rice plant to evaluate their usefulness for decontamination of mycotoxins. The strong and constitutive rice Act1 promoter did not cause accumulation of TRI101 protein in transgenic rice plants. In contrast, the same promoter was suitable for transgenic production of ZHD101 protein; so far, five promising T0 plants have been generated. Low transgenic expression of Tri101 was suggested to be increased by addition of an Ω enhancer sequence upstream of the start codon.

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  We found that a carbonic anhydrase, YadF, is essential for cell growth in the absence of another carbonic anhydrase, CynT, in Escherichia coli. However, mutant strains lacking both of them grew at high CO₂ concentrations (5%), where non-enzymatic mechanisms generate HCO₃^-. This suggests that these carbonic anhydrases are essential because they maintain HCO₃^- levels at ambient CO₂ concentrations.

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  Voltage-dependent K⁺ channels consist of a voltage-sensing region and a pore-forming region. Here we have identified the negative residues of the second transmembrane segment in the plant voltage-dependent K⁺ channel, KAT1, which involves the function of voltage sensing. Point mutations at D95 and D105 but not D89 and D116 failed to complement the K⁺ uptake deficient properties of the mutant yeast. In vitro translation and translocation experiments showed that the membrane integration of the third and fourth segments involving voltage sensor were impaired by the replacement of D95 or D105 by serine. These data show that both the residues play a crucial role in the membrane topogenesis of the voltage sensor in KAT1.

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  The Schizosaccharomyces pombe Och1p is required for the initiation of outer chain elongation of N-linked oligosaccharides. In this report, we investigated the transcriptional control of the S. pombe och1⁺ gene and found that the expression of the och1⁺ gene was not regulated during the cell cycle, but was induced by NaCl and KCl through a transcription factor, Atf1p.

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  Pyruvate was catabolized anaerobically by resting cells of Fusarium oxysporum to form formate and acetate. Addition of nitrate decreased the accumulation of formate in the medium with concomitant formation of nitrite and N₂O. The results suggested a unique metabolic pathway that occurs in fungi mediated by pyruvate-formate lyase to supply electrons via formate to fungal denitrification.

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  It has been found that commercial water-soluble soybean polysaccharides (SSPS) can make cooked rice and noodles non-sticky and prevent rice grains and noodles from adhering to each other. We studied in detail the phenomenon of rice cooked with SSPS. We assumed that the phenomenon resulted from the interaction between SSPS and starch during cooking, and studied the effects of SSPS on the gelatinizing behavior of rice starch by using a Rapid-Visco-Analyzer. The addition of SSPS reduced the viscosity of the gelatinized starch. This lower final viscosity of the rice starch was more distinct from than that of potato starch. These results imply that the properties of SSPS in forming a non-sticky condition might result from a decrease in the viscosity of the gelatinized starch.

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  To characterize the factors involved in the production of volatile aldehydes during mashing, a model mashing experiment was done. After we inactivated the endogenous lipoxygenase (LOX) activity in the mash by mashing at 70°C for 30 min, further incubation with recombinant barley LOX-1 stimulated the accumulation of 2(E)-nonenal; however, this effect was significantly reduced by boiling the mash sample. The result suggests that both LOX-1 and a heat-stable enzymatic factor are involved in the production of 2(E)-nonenal during mashing. Malt contained fatty acid hydroperoxide lyase-like activity (HPL-like activity) that transformed 9-hydroperoxy-10(E),12(Z)-octadecadienoic and 13-hydroperoxy-9(Z),11(E)-octadecadienoic acid into 2(E)-nonenal and hexanal, respectively. Proteinase K sensitivity tests showed that they are distinct factors. 9-HPL-like activity survived through the mashing at 70°C for 30 min but was inactivated by boiling, suggesting it will be the heat-stable enzymatic factor found in the model mashing experiment.

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  We localized the T cell epitope regions of chicken ovomucoid (OVM), a potent egg allergen, with the overlapping pin-peptides covering the entire sequence of OVM and three strains of mice with different haplotypes. In C3H/He (H-2^k) mice, the T cells recognized relatively broad regions on OVM; the dominant regions were 49-93 and 97-114 residues, and the subdominant regions were 7-21, 37-48, 94-96, 115-123 and 145-177 residues. In contrast, a more limited number of T cell epitope regions were localized in BALB/c (H-2^d) and C57BL/6 (H-2^b) mice. The T cells from BALB/c mice recognized 100-114 and 157-171 residues, and the T cells from C57BL/6 mice recognized only 157-180 residues. These results were confirmed by using peptides separately synthesized and purified on the putative epitope regions. The roles of the carbohydrate moieties and cysteine residues involved in the disulfide bridges of OVM were also examined, and we found that they were not important in recognition by the T cell/antigen presenting cell.

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  The effect on immunoglobulin production of a commercially available casein phosphopeptide preparation (CPP-III) consisting mainly of bovine αs2-casein (1-32) and β-casein (1-28) in mice that had orally ingested lipopolysaccharide (LPS) from Salmonella typhimurium was investigated. No significant difference in body weight gain was observed between the mice fed on the CPP-III-added diet and those fed on the control diet. The mice fed on the CPP-III-added diet exhibited similar serum and intestinal IgG, IgM, and IgE responses towards LPS to those fed on the control diet. In contrast, fecal and intestinal anti-LPS IgA and total IgA in mice fed on the CPP-III-added diet were significantly higher than in those fed on the control diet. Spleen cells from mice fed on the CPP-III-added diet produced larger amounts of IgA, IL-5, and IL-6 than cells from mice fed on the control diet. These results suggest that dietary casein phosphopeptide may protect a host from invasion of the intestinal mucosa by food-born pathogenic microorganisms.

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  The status of hyaluronan, the major glycosaminoglycan in the skin, is regulated by many factors such as cytokines and glucocorticoids. To examine whether and how protein malnutrition affects the status of skin hyaluronan, the hyaluronan content and mRNA levels of hyaluronan synthases (has) were analyzed in the skin of rats fed on a protein-free diet or on a 12% gluten diet. When these malnourishing diets had been given for 1 week, the hyaluronan content was significantly reduced as compared with that in rats fed on a 12% casein diet. Substantial falls in the mRNA levels of rhas2 and rhas3 were also observed. The reduction of mRNAs was already evident on the second day of treatment with the malnourishing diets. These results suggest that protein malnutrition has a primary impact on the gene expression of rhass, which leads to the reduction of hyaluronan content and to disfunction of the skin.

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  The effects of dietary casein level (5-40%) on the liver microsomal phospholipid profile, Δ6-desaturase activity and related variables were investigated in rats to examine whether the dietary protein level affected the Δ6-desaturase activity through an alteration of the liver microsomal phospholipid profile. The effects of supplementing a 10% casein diet with certain amino acids were also investigated. The concentration of hepatic S-adenosylmethionine (SAM), the ratio of phosphatidylcholine (PC) to phosphatidylethanolamine (PE) and the Δ6-desaturase activity in liver microsomes, and the ratio of arachidonate to linoleate of microsomal PC increased with increasing dietary casein level. There were significant correlations between the dietary methionine content and hepatic SAM concentration, hepatic SAM concentration and microsomal PE concentration, and microsomal PE concentration and Δ6-desaturase activity. Supplementation of the 10% casein diet with methionine significantly increased the hepatic SAM concentration, PC/PE ratio, Δ6-desaturase activity, and arachidonate/linoleate ratio, whereas cystine supplementation had no or little effect on these variables. These increases induced by methionine were significantly suppressed by additional glycine. The results obtained here, together with those in our previous report, suggest that quantity and type of dietary protein might affect the Δ6-desaturase activity through an alteration of the liver microsomal profile of phospholipids, especially PE, and that the alteration of phospholipid profile might be mediated by a hepatic SAM concentration that reflects the dietary methionine level.

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  The phytate content in soymilk is known to affect tofu curdling. A rapid measurement of phytate from a water extract of soybean (raw soymilk) in an early stage of tofu processing was investigated using mid-infrared spectroscopy (IR) with an ATR accessory. IR absorption of phytate was observed from 1200 cm^-1 to 900 cm^-1, and saccharide and protein in the extract also had IR absorption in the same region. In order to separate phytate from other components, the phytate was precipitated completely by the addition of calcium under alkaline condition (pH 11.5). The precipitate was dissolved in citrate buffer (pH 6.0) and then used for IR measurement. The absorbance at 1070 cm^-1 correlated well with the phytate content of the soymilk. The measurement of phytate in raw soymilk can be done rapidly by FT-IR measurement with an ATR accessory and gives reproducible values, which can be used for the measurement of phytate content in various soybeans for tofu making.

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  Probiotic supplements induce immunological responses in the host, and dietary fructooligosaccharides (FOS) stimulate the growth of selected intestinal microflora. In this study we investigated the immunological influences of orally administrated FOS. BALB/c mice were oral administered 0-7.5% FOS for 6 weeks, and the intestinal mucosal immune responses were measured. In the 2.5%-FOS group, fecal IgA was significantly increased. IgA secretion by Peyer's patch (PP) cells was upregulated in a dose-dependent way in response to FOS and CD4⁺ T cells from PP showed a dose-dependent increase in production of interferon-γ and interleukin (IL) 10, and a high response in production of IL-5 and IL-6. In contrast, FOS suppressed serum IgG1. Our findings suggest that FOS supplementation changes the intestinal environment of microflora, and leads to upregulation of IgA secretion in CD4⁺ PP cells in intestinal mucosa, and to suppression of the systemic immune response to type 2 helper T (Th2) dominant.

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  Ovalbumin, a member of the serpin superfamily, is transformed via an intermediate state into a non-cleaved, thermostabilized form (S-ovalbumin) during either the storage of unfertilized eggs or development of fertilized eggs; essentially the same thermostabilization also occurs upon in vitro incubation of isolated ovalbumin under alkaline conditions. To investigate the implications of a partial insertion of the α-helical serpin loop into β-sheet A that has been proposed as a conformational mechanism for S-ovalbumin production, we examined the thermostabilization process of ovalbumin with different loop structures. When the thermostabilization processes were compared for the intact, P1-P1′-cleaved and P1-P1′/P8-P7-cleaved forms of egg white ovalbumin, both the rates for the conversion from the native to intermediate and from the intermediate to S-ovalbumin were almost indistinguishable among the three protein forms. Furthermore, the fully loop-inserted form of recombinant ovalbumin mutant R339T that had been thermostabilized by P1-P1′ cleavage with Tm values from 72 to 88°C was further thermostabilized by an alkaline treatment, yielding a final product (loop inserted S-ovalbumin) with a Tm value of 93°C. No significant difference was found between native ovalbumin and S-ovalbumin in respect of the rate of proteolytic cleavage of the loop by elastase and subtilisin. These data strongly suggest that S-ovalbumin is produced by a mechanism other than that of the partial loop insertion model.

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  Our previous study (Biosci. Biotechnol. Biochem., 66, 2449-2457 (2002)), suggested that ferulic acid was transported via a monocarboxylic acid transporter (MCT). Transepithelial transport of ferulic acid was examined in this study by directly measuring the rate of its transport across Caco-2 cell monolayers. Ferulic acid transport was dependent on pH, and in a vectorical way in the apical-basolateral direction. The permeation of ferulic acid was concentration-dependent and saturable; the Michaelis constant was 16.2 mM and the maximum velocity was 220.4 nmol min^-1 (mg protein)^-1. Various substrates for MCTs, such as benzoic acid and acetic acid, strongly inhibited the permeation of ferulic acid, demonstrating that ferulic acid is obviously transported by MCT. Antioxidative phenolic acid compounds from dietary sources like ferulic acid would be recognized and transported by MCT by intestinal absorption.

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  This study examined the hypoglycaemic activity of Cyclocarya paliurus (Batal.) Iljinskaja (C. paliurus) in ICR mice by oral glucose tolerance testing. The blood glucose level was significantly lower in the C. paliurus extract treatment group than in the control group after animals were given sucrose. This difference was not observed following the administration of glucose. We demonstrated that the chronological change in the level of blood glucose in genetically hyperglycemic obese KK-A^y mice is significantly lower when C. paliurus extract is administered daily for three weeks. An in vitro study showed that C. paliurus inhibits α-glucosidase, a disaccharide-degrading enzyme in the small intestinal mucosa, leading to a decrease in the absorption of glucose into the blood and a subsequent lowering of the blood glucose level.

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  The orange-pigmented Thraustochytrium, CHN-1 strain was found to contain astaxanthin as the main carotenoid pigment. Echinenone, canthaxanthin, phoenicoxanthin and β-carotene were also identified by high-performance liquid chromatography (HPLC) and HPLC-mass spectrometry. The total extractable carotenoid level was found to increase with culture age.

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  The effective substances (polyphenols, phenolic and ascorbic acids, flavonoids and dietary fibers) and antioxidative activities, using different radical-scavenging tests, were determined for Jaffa sweeties and grapefruit. The antioxidative activities comprised the contributions from polyphenols, phenolic acids, flavonoids and ascorbate components, and were well-correlated with polyphenols and flavonoids. The correlation coefficient between the polyphenols and antioxidative activity varied from 0.73 to 0.99. All applied methods showed that sweeties had higher antioxidative activity than grapefruit. Experiments on laboratory animals show that diets supplemented with sweeties, and to a lesser extent with grapefruit, increased the plasma antioxidative potential and improved the lipid metabolism, especially in the rats fed with added cholesterol. These findings provide additional characterization of the nutritional value of citrus fruits and their influence on the lipid metabolism in rats.

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  Glyceraldehyde (200 mM) and N^α-acetyllysine (100 mM) were incubated in 0.2 M sodium phosphate buffer (pH 7.4) at 37°C for a week. A major compound, glyceraldehyde-related Maillard reaction product, was purified from the reaction mixture using reverse phase (ODS)-HPLC. It was identified as 1-(5-acetylamino-5-carboxypentyl)-3-hydroxy-5-hydroxymethyl-pyridinium, named as GLAP (Glyceraldehyde derived Pyridinium compound), using NMR and MS analyses. It was suggested that GLAP as a novel advanced glycation end product (AGE) is one of the key compounds in the glyceraldehyde-related Maillard reaction.

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  β-Lactotensin (β-LT: His-Ile-Arg-Leu) is an ileum-contracting peptide derived from residues No. 146-149 of bovine β-lactoglobulin. The ileum-contracting activity of β-LT was blocked by the NT₁ antagonist SR48692. β-LT was selective for the neurotensin NT₂ receptor while neurotensin was selective for the NT₁ receptor. β-LT is the first natural ligand showing selectivity for the NT₂ receptor. β-LT showed hypertensive activity after intravenous administration at a dose of 30 mg/kg in conscious rats, while neurotensin showed hypotensive activity. The hypertensive activity of β-LT was blocked by levocabastine (1 mg/kg, i.v.), an NT₂ antagonist. SR48692, which blocked the hypotensive activity of neurotensin, had no effect on the hypertensive activity of β-LT. These results suggest that the hypertensive activity of β-LT is mediated by the NT₂ receptor. It was concluded that the NT₁ and NT₂ receptors mediate the opposite effect on blood pressure.

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  We isolated mutants of S. cerevisiae in which expression of the JEN1 gene encoding a pyruvate transporter was insensitive to glucose repression. The isolated mutant GDR19 expressed JEN1 and absorbed pyruvate in the presence of glucose. In a DNA microarray analysis, GDR19 highly expressed many more genes, including JEN1, in the presence of glucose compared with the parental strain B29. Some of these genes are under the control of the transcription factor Mig1p and are normally repressed in the presence of glucose. The concentrations of organic acids in sake made with GDR19 were different from those in sake made with B29. Changes in the pyruvate concentration in the sake mash made with GDR19 were not very different from those in sake mash made with B29, and both GDR19 and B29 expressed JEN1 during fermentation. When the ethanol concentration was over 2%, JEN1 expression in B29 was similar in the presence and absence of glucose. The expression of JEN1 in sake mash in spite of the presence of glucose appeared to be caused by the coexistence of ethanol.

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  Glycine aminotransferase (EC 2.6.1.4; GlyAT) was presumed to be an enzyme concerning the supply of glycine for the extracellular porphyrin production by Rhodopseudomonas palustris No. 7. GlyAT was purified from strain No. 7 as an electrophoretically homogeneous protein. The enzyme was a monomer protein with the molecular weight of about 42,000. From the absorption spectrum of the enzyme (350 nm, 410 nm), it was indicated that the enzyme had pyridoxal phosphate as a prosthetic group. The enzyme showed high substrate specificity for glutamate as an amino group donor. Apparent K_ms for glutamate and glyoxylate were 6.20 mM and 3.75 mM, respectively. The V_max and K_cat for glutamate were 66.8 μmol/min/mg protein and 46.8 s^-1, respectively. The V_max and K_cat for glyoxylate were 68.8 μmol/min/mg protein and 48.2 s^-1. The optimum temperature and pH were 40~45°C and 7.0~7.5, respectively. The enzyme activity lowered to about 50% in the presence of 15 mM ammonium chloride.

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  Spontaneously developing pocks (S pocks) of Streptomyces azureus ATCC14921 were formed by the both functions of conjugative plasmid pSA1 and lysogenic phage SAt2. The formation was affected by the dose of UV irradiation. The mean pock diameter in cultures treated with UV light at 0, 7.1, 14.2 and 21.3×10² μW·erg/cm, respectively, were 1.3, 0.4, 2.2, and 0.5 mm. The dose affected conjugative plasmid pSA1 related to pock formation. There was UV damage of autonomous pSA1 replicon and UV induction of the chromosomal integrated sequence. Increases and decreases in the amount of autonomous pSA1 replicon corresponded to increases and decreases, respectively, in the diameter of the pocks. Both pSA1 and SAt2 syntheses were developed in the large pocks (1.3 and 2.2 mm), but only SAt2 synthesis was developed in the pinhole pocks (0.4 and 0.5 mm).

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  Respectively, exogenous pamamycin-607 and A-factor restored or stimulated aerial mycelium formation in 30 (67%) and 6 (13%) of 45 Streptomyces strains, and both restored or stimulated it in 5 strains (11%). Pamamycin-607 production was detected in 3 of those strains that responded to pamamycin-607. These findings indicate that pamamycin-607 acts on the common regulatory system for aerial mycelium formation in Streptomyces spp. but is not a universal autoregulator. Increased or decreased antibacterial production occurred in 5 strains in association with aerial mycelium formation by pamamycin-607 or A-factor.

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  The microbial enantioselective reduction of acetylpyridine derivatives was studied. Many microorganisms were found to reduce 5-acetylfuro[2,3-c]pyridine (AFP) to (S)-5-(1-hydroxyethyl)furo[2,3-c]-pyridine (FPH). Candida maris IFO10003 reduced AFP to (R)-FPH with high enantioselectivity. The microbial reduction reaction was optimized. The aeration conditions and glucose concentration affected the yield and stereoselectivity. The cells accumulated 17.5 g/l (107 mM) of (R)-FPH with a 99% yield and 97% enantiomeric excess (e.e.). A cell-free extract of C. maris accumulated 91.5 g/l (559 mM) with over 99% e.e. with enzymatic NADH regeneration. (R)-FPH is an important intermediate for the synthesis of HIV reverse-transcriptase inhibitor, and other optically active 1-(pyridyl)ethanol derivatives are versatile chiral building blocks for asymmetric synthesis.

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  We isolated aromatics-degrading bacteria from the gut of a lower termite, Coptotermes formosanus, using a mineral salt medium containing various aromatic compounds as the sole carbon source. Two species, Burkholderia sp. strain VE22 and Citrobacter sp. strain VA53, were isolated by aerobic enrichment culture with veratraldehyde and vanillin, respectively. Strain VA53 could also grow and metabolize vanillin anaerobically.

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  2-Deoxyribose 5-phosphate was produced from acetaldehyde and dihydroxyacetone phosphate via D-glyceraldehyde 3-phosphate by Klebsiella pneumoniae B-4-4 through deoxyriboaldolase- and triosephosphate isomerase-catalyzing reactions. Under the optimum conditions, 98.7 mM 2-deoxyribose 5-phosphate was produced from 200 mM acetaldehyde and 117 mM dihydroxyacetone phosphate in 2 h with a molar yield of 84%. The 2-deoxyriobse 5-phosphate produced was directly transformed to 2′-deoxyribonucleoside by phosphopentomutase- and nucleoside phosphorylase-catalyzing reactions.

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