Bioscience, Biotechnology, and Biochemistry
Online ISSN : 1347-6947
Print ISSN : 0916-8451
Volume 72 , Issue 5
Showing 1-39 articles out of 39 articles from the selected issue
Award Reviews
  • Takahiro ISHIKAWA, Shigeru SHIGEOKA
    2008 Volume 72 Issue 5 Pages 1143-1154
    Published: May 23, 2008
    Released: May 23, 2008
    [Advance publication] Released: May 07, 2008
    JOURNALS FREE ACCESS
    Ascorbate (AsA), the most abundant water-soluble redox compound in plants and eukaryotic algae, has multiple functions. There is compelling genetic evidence that the biosynthesis of AsA proceeds via a D-mannose/L-galactose pathway and is the most significant source of AsA in plants. AsA plays important roles in antioxidative defense, particularly via the AsA/glutathione cycle. AsA peroxidase (APX) plays a central role in the cycle and is emerging as a key enzyme in cellular H2O2 metabolism. Plants possess diverse APX isoenzymes in cellular compartments, including the chloroplast, cytosol, and microbody. In algae, however, the number and distribution of APX proteins are quite limited. Recent progress in molecular biological analysis of APX isoenzymes has revealed elaborate mechanisms for the tissue-dependent regulation of two chloroplastic APX isoenzymes by alternative splicing, and for redox regulation of cytosolic APX gene expression in response to light stress. Furthermore, transgenic plants overexpressing a chloroplastic APX isoenzyme enable us to evaluate the behavior of the enzyme under conditions of photo-oxidative stress. Molecular physiological analysis has revealed that cytosolic APX is part of the system modulating the cellular H2O2 level in redox signaling.
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  • Shou TAKASHIMA
    2008 Volume 72 Issue 5 Pages 1155-1167
    Published: May 23, 2008
    Released: May 23, 2008
    [Advance publication] Released: May 07, 2008
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    Sialic acids are negatively charged acidic sugars, and sialylglycoconjugates often play important roles in various biological phenomena. Sialyltransferases are involved in the synthesis of sialylglycoconjugates, and 20 members of the mammalian sialyltransferase family have been identified to date. These sialyltransferases are grouped into four families according to the carbohydrate linkages they synthesize: β-galactoside α2,3-sialyltransferases (ST3Gal I-VI), β-galactoside α2,6-sialyltransferases (ST6Gal I and II), GalNAc α2,6-sialyltransferases (ST6GalNAc I-VI), and α2,8-sialyltransferases (ST8Sia I-VI). Analysis of the amino acid sequence similarities, substrate specificities, and gene structures of mouse sialyltransferases has revealed that they can be further divided into seven subfamilies. The genomic structural resemblance of members of the same subfamily suggests that they arose from a common ancestral gene through gene duplication events. These multiple sialyltransferase genes are needed for fine control of the expression of sialylglycoconjugates, resulting in a variety of developmental stage- and tissue-specific glycosylation patterns.
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  • Tomonobu TOYOMASU
    2008 Volume 72 Issue 5 Pages 1168-1175
    Published: May 23, 2008
    Released: May 23, 2008
    [Advance publication] Released: May 07, 2008
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    Cyclic diterpenoids are commonly biosynthesized from geranylgeranyl diphosphate (GGDP) through the formation of carbon skeletons by specific cyclases and subsequent chemical modifications, such as oxidation, reduction, methylation, and glucosidation. A variety of diterpenoids are produced in higher plants and fungi. Rice produces four classes of diterpene phytoalexins, phytocassanes A to E, oryzalexins A to F, oryzalexin S, and momilactones A and B. The six diterpene cyclase genes involved in the biosynthesis of these phytoalexins were identified and characterized. Fusicoccin A was produced by the phytopathogenic Phomopsis amygdali and served as a plant H+-ATPase activator. A PaFS, encoding a fungal diterpene synthase responsible for fusicoccin biosynthesis, was isolated. The PaFS is an unusual chimeric diterpene synthase that possesses not only terpene cyclase activity (the formation of fusicoccadiene, a biosynthetic precursor of fusicoccin A), but also prenyltransferase activity (the formation of GGDP). Thus, we identified a unique multifunctional diterpene synthase family in fungi.
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Analytical Chemistry Note
Biochemistry & Molecular Biology Regular Papers
  • Fabiana de Souza GOUVEIA, Fabrícia GIMENES, Adriana FIORINI, Ma ...
    2008 Volume 72 Issue 5 Pages 1190-1198
    Published: May 23, 2008
    Released: May 23, 2008
    [Advance publication] Released: May 07, 2008
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    The Rhynchosciara americana C3-22 gene is located in an amplified domain and is developmentally expressed. The aim of the present work was to identify intrinsically bent DNA sites in a segment containing the gene promoter and downstream sequence. The results indicated that this gene is flanked by intrinsically bent DNA sites. Three bent DNA sites (b−3, b−2, and b−1) were localized in the promoter, and one was localized downstream of the gene (b+1). These sites had helical parameters that confirmed the curved structure, as well as segments with left-handed superhelical writhe. In silico analysis of the promoters of four other insect genes, which encode secreted polypeptides, showed that they all had curved structures and similar helical parameters. Correlation with other results indicates that the detected intrinsically bent DNA sites that flank the C3-22 gene might be a consensus feature of the gene structure in the amplified domains.
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  • Sayoko SUZUKI, Shin-ichi ISHIKAWA, Keizou ARIHARA, Makoto ITOH
    2008 Volume 72 Issue 5 Pages 1212-1218
    Published: May 23, 2008
    Released: May 23, 2008
    [Advance publication] Released: May 07, 2008
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    It has been demonstrated that the composition of molecular species of adipose tissue triacylglycerols (TGs) from farm animals are not equally synthesized and that some molecular species are preferentially synthesized. The objective of the present study was to determine whether exogenous fatty acids (FAs) would affect the TG composition. To this end, the composition of TG molecular species stored in porcine adipocytes differentiated with several long-chain FAs was analyzed by gas chromatography. The addition of each FA for 6 d increased TG molecular species having two or three added FAs. However, the molecular species compositions at 15 d after the addition of each FA resembled those of cells with no added FAs. Moreover, some common molecular species in all experimental cells increased, as well as cells with no added FAs. It was concluded that the addition of FAs increases the contents of specific molecular species, but does not affect the synthetic processes of individual TG molecular species.
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  • Erick K. CHERUIYOT, Louis M. MUMERA, Wilson K. NG’ETICH, Ahmed H ...
    2008 Volume 72 Issue 5 Pages 1219-1226
    Published: May 23, 2008
    Released: May 23, 2008
    [Advance publication] Released: May 07, 2008
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    An experiment was conducted to determine the association of tea catechins to water stress in tea, with the objective of determining their suitability as indicators for predicting drought tolerance in tea (Camellia sinensis). The study consisted of six tea clones (BBK 35, TRFK 6/8, TRFK 76/1, TRFK 395/2, TRFK 31/30, and TRFK 311/287) and four levels of soil water content (38, 30, 22, and 14% v/v), which were arranged in a complete randomized design and replicated 3 times. The treatments were maintained for a period of 12 weeks. Tea shoots were sampled for catechin analysis during the 6th week of water treatment, in which fresh shoots with two leaves and a bud were plucked and steamed for 2 min, and dried at 70 °C to constant weight. Subsequently, the samples were ground and analyzed for catechins using an HPLC system. The total catechins showed significant correlation with shoot growth (r=0.65, P=0.006), soil water content (r=0.54, P=0.0066), and water stress index (r=0.67, P=0.0004). The epicatechin (EC) correlated with shoot growth (r=0.58, P=0.0032), soil water content (r=0.62, P=0.0014), and water stress index (r=0.63, P=0.0010). Similarly, epigallocatechin (EGC) correlated with shoot growth (r=0.65, P=0.0006), soil water content (r=0.50, P=0.0133), and water stress index (r=0.60, P=0.0021). However, epigallocatechin gallate (EGCg) and epicatechin gallate (ECG) showed no significant response to changes in soil water content. The shoot contents of EC and EGC in the six clones showed varied responses, with a distinct pattern in the water-stress tolerant clones (TRFK 6/8 and TRFK 31/30). The results suggest a potential use for EC and EGC as indicators in predicting drought tolerance in tea.
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  • Tomoyuki MIYABAYASHI, Masashi YAMAMOTO, Akiko SATO, Seiji SAKANO, Yasu ...
    2008 Volume 72 Issue 5 Pages 1242-1248
    Published: May 23, 2008
    Released: May 23, 2008
    [Advance publication] Released: May 07, 2008
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    Embryonic stem cells (ESCs), which have characteristics such as self-renewal, indefinite proliferation, and pluripotency, are thought to hold great promise for regenerative medicine. ESCs are generally cultured on mouse embryonic fibroblast (MEF) or MEF conditioned medium (MEF-CM). However, for therapeutic applications, it is preferable for ESCs to be cultured under chemically defined conditions.
    Here, we report synthetic compounds that allow expansion of undifferentiated mouse ESCs in the absence of MEF, Leukemia Inhibitory Factor (LIF), and Fetal Bovine Serum (FBS). ESCs cultured for more than 30 d in a serum-free medium supplemented with indole derivertives retained their characteristic morphology and expressed markers such as SSEA-1, OCT3/4, Rex-1, Sox2, and Nanog. They consistently differentiated into many types of cells, including neurons, muscle cells, and hepatocytes.
    These results indicate that our compounds provide a more efficient and safer large-scale culture system for pluripotent ESCs, and hence might contribute to the use of ESCs in therapeutic applications.
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  • Xiaomin HU, Jia LI, Bjarne Munk HANSEN, Zhiming YUAN
    2008 Volume 72 Issue 5 Pages 1257-1263
    Published: May 23, 2008
    Released: May 23, 2008
    [Advance publication] Released: May 07, 2008
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    The surface layer protein encoding genes from five mosquito-pathogenic Bacillus sphaericus isolates were amplified and sequenced. Negative staining of the S-layer protein extracted from the cell wall of wild-type B. sphaericus C3-41 was prepared. It showed a flat-sheet crystal lattice structure. Two genes encoding the entire and N-terminally truncated S-layer protein (slpC and ΔslpC respectively), were ligated into plasmid pET28a and expressed in Escherichia coli. SDS–PAGE revealed that about 130 KD and 110 KD proteins could be expressed in the cytoplasm of recombinant E. coli BL21(pET28a/slpC) and E. coli BL21(pET28a/ΔslpC) respectively. Furthermore, an intracellular sheet-like or fingerprint-shape structure was investigated in two recombinant strains, which expressed SlpC and ΔSlpC protein respectively, by ultrathin microscopy study, but bioassay results suggested that the S-layer protein of wild B. sphaericus C3-41 and recombinant E. coli BL21 (pET28a/slpC) have no direct toxicity against mosquito larvae. These results should provide information for further understanding of the function of S-layer protein of pathogenic B. sphaericus.
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  • Nan-Yin CHEN, Hsi-Huai LAI, Tai-Hao HSU, Fang-Yi LIN, Jian-Zhi CHEN, H ...
    2008 Volume 72 Issue 5 Pages 1283-1289
    Published: May 23, 2008
    Released: May 23, 2008
    [Advance publication] Released: May 07, 2008
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    Tremella mesenterica (TM) is a common food and folk medicine widely used in several Asian countries as a tonic for the lungs. In the present study, we compared the effects of extracellular polysaccharides (EPS), intracellular polysaccharides (IPS), and ethanol extract (EE) of Tremella mesenterica on the induction of apoptosis into human lung carcinoma A549 epithelial cells. The EE, but not the EPS or the IPS, almost completely inhibited the growth of A549 cells. The results of Annexin V-FITC/PI staining and flow cytometric analysis indicated that the percentage of Annexin V+/PI cells in EE-treated cells increased to 32.8%. The results of further investigation showed a disruption of mitochondrial transmembrane potential (ΔΨm), the production of reactive oxygen species (ROS), and the activation of caspase-3 protein in EE-treated cells. These findings suggest that EE can decrease cell viability and induce apoptosis in A549 cell lines by activating a mitochondrial pathway.
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  • Kazumi HIRAGA, Yoshie UENO, Kohei ODA
    2008 Volume 72 Issue 5 Pages 1299-1306
    Published: May 23, 2008
    Released: May 23, 2008
    [Advance publication] Released: May 07, 2008
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    In this study, the glutamate decarboxylase (GAD) gene from Lactobacillus brevis IFO12005 (Biosci. Biotechnol. Biochem., 61, 1168–1171 (1997)), was cloned and expressed. The deduced amino acid sequence showed 99.6% and 53.1% identity with GAD of L. brevis ATCC367 and L. lactis respectively. The His-tagged recombinant GAD showed an optimum pH of 4.5–5.0, and 54 kDa on SDS–PAGE. The GAD activity and stability was significantly dependent on the ammonium sulfate concentration, as observed in authentic GAD. Gel filtration showed that the inactive form of the GAD was a dimer. In contrast, the ammonium sulfate-activated form was a tetramer. CD spectral analyses at pH 5.5 revealed that the structures of the tetramer and the dimer were similar. Treatment of the GAD with high concentrations of ammonium sulfate and subsequent dilution with sodium glutamate was essential for tetramer formation and its activation. Thus the biochemical properties of the GAD from L. brevis IFO12005 were significantly different from those from other sources.
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  • Masakazu KAWAMURA, Shogo ITO, Norihito NAKAMICHI, Takafumi YAMASHINO, ...
    2008 Volume 72 Issue 5 Pages 1307-1316
    Published: May 23, 2008
    Released: May 23, 2008
    [Advance publication] Released: May 07, 2008
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    In the model higher plant Arabidopsis thaliana, the CCA1 (CIRCADIAN CLOCK-ASSOCIATED 1) gene plays important circadian clock-associated roles. The CCA1 protein is a member of a small subfamily of single MYB-related transcription factors. This family consists of several homologous CCA1-like transcription factors, including the closest homolog LHY (LATE ELONGATED HYPOCOTYL). To gain insight into the molecular function of CCA1 and its homologs, here we took a unique genetic approach that was recently developed for Arabidopsis thaliana. Through this strategy, referred to as CRES-T (Chimeric REpressor Silencing Technology), a transgenic plant was constructed to produce a dominant negative transcriptional repressor (designated CCA1-SRDX). By employing the resulting transgenic lines, together with previously established cca1 lhy double mutant and CCA1-ox (over-expressing) plants, their circadian clock-associated phenotypes were examined and compared with each other. The observed clock-associated phenotypes of the CCA1-SRDX plants were very similar to those of CCA1-ox, but not to those of cca1 lhy, suggesting that CCA1 acts predominantly as a transcriptional repressor in nature. However, the developmental morphology (or architecture) of adult CCA1-SRDX plants were quite different from that of CCA1-ox, suggesting that CCA1 might also be implicated, directly or indirectly, in an as yet unknown circadian-associated output pathway at a late developmental stage.
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  • Hajime SHIBUYA, Taisei KIKUCHI
    2008 Volume 72 Issue 5 Pages 1325-1332
    Published: May 23, 2008
    Released: May 23, 2008
    [Advance publication] Released: May 07, 2008
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    A family of endoglucanases belonging to glycoside hydrolase family (GHF) 45 have been isolated from the pine wood nematode Bursaphelenchus xylophilus. Here we describe the purification and characterization of the recombinant enzymes, named Bx-ENG-1, 2, and 3, expressed in Pichia pastoris. The respective molecular masses of purified Bx-ENG-1, 2, and 3 were estimated to be 18, 33–39, and 100–140 kDa by SDS–PAGE, and 18, 67, and 252 kDa by gel filtration, suggesting that Bx-ENG-1 existed in an unglycosylated monomeric form and Bx-ENG-2 and Bx-ENG-3 in a glycosylated dimeric form. The enzymatic properties of the recombinant enzymes were similar to each other: optimal activity at 60 °C at about pH 6.0, like other endoglucanases of GHF45. The recombinant enzymes displayed the highest activity toward lichenan, and lower activities were observed on carboxymethyl cellulose and amorphous cellulose. Nematode enzymes also hydrolyzed glucomannan, the most abundant hemicellulose in the cell walls of softwood. These substrate specificities suggest that B. xylophilus endoglucanases acted on the cellulose-hemicellulose complex in the cell walls, resulting in a weakening of the mechanical strength of the cell walls to facilitate the nematode’s feeding on plant cells.
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  • Takayuki INOUYE, Masaki ODAHARA, Tomomichi FUJITA, Mitsuyasu HASEBE, Y ...
    2008 Volume 72 Issue 5 Pages 1340-1347
    Published: May 23, 2008
    Released: May 23, 2008
    [Advance publication] Released: May 07, 2008
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    RecA protein is widespread in bacteria, and it plays a crucial role in homologous recombination. We have identified two bacterial-type recA gene homologs (PprecA1, PprecA2) in the cDNA library of the moss Physcomitrella patens. N-terminal fusion of the putative organellar targeting sequence of PpRecA2 to the green fluorescent protein (GFP) caused a targeting of PpRecA2 to the chloroplasts. Mutational analysis showed that the first AUG codon acts as initiation codon. Fusion of the full-length PpRecA2 to GFP caused the formation of foci that were colocalized with chloroplast nucleoids. The amounts of PprecA2 mRNA and protein in the cells were increased by treatment with DNA damaging agents. PprecA2 partially complemented the recA mutation in Escherichia coli. These results suggest the involvement of PpRecA2 in the repair of chloroplast DNA.
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Biochemistry & Molecular Biology Notes
Food & Nutrition Science Regular Papers
  • Seung-Hae BAEK, Min PARK, Jae-Hee SUH, Hye-Seon CHOI
    2008 Volume 72 Issue 5 Pages 1176-1182
    Published: May 23, 2008
    Released: May 23, 2008
    [Advance publication] Released: May 07, 2008
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    The protective effects of an extract of young radish (Raphanus sativus L) cultivated with sulfur (sulfur-radish extract) and of sulforaphane, an isothiocyanate, on carbon tetrachloride (CCl4)-induced liver injury were observed in mice. CCl4 produced a marked increase in the serum level of alanine aminotransferase (ALT), primed lipid peroxidation, and resulted in intense necrosis due to oxidative stress. Oral administration of the sulfur-radish extract and of sulforaphane after CCl4-induced liver injury both decreased the serum level of ALT, reduced the necrotic zones, inhibited lipid peroxidation, and induced phase 2 enzymes without affecting cytochrome P450-2E1 (CYP2E1). These results suggest that the administration of the sulfur-radish extract and of sulforaphane may partially prevent CCl4-induced hepatotoxicity, possibly by indirectly acting as an antioxidant by improving the detoxification system.
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  • Toshio NORIKURA, Akiko KOJIMA-YUASA, Miki SHIMIZU, Xuedan HUANG, Sheng ...
    2008 Volume 72 Issue 5 Pages 1183-1189
    Published: May 23, 2008
    Released: May 23, 2008
    [Advance publication] Released: May 07, 2008
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    Blumea balsamifera is known to improve physiological disorders such as rheumatism and hypertension, but its anticancer activity has not been well elucidated. In this study, we found that Blumea balsamifera MeOH extract (BME) induced growth-inhibitory activity in rat and human hepatocellular carcinoma cells without cytotoxicity in rat hepatocytes which were used as a normal cell model. BME induced cell cycle arrest at the G1 phase via decreases in the expression of cyclin-E and phosphorylation of retinoblastoma protein. Furthermore, BME reduced the level of a proliferation-inducing ligand, that stimulates tumor cell growth. These findings suggest that BME has possible therapeutic potential in hepatoma cancer patients and that depletion of cellular APRIL is an important mechanism in the growth-inhibitory effect of BME.
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  • Makoto TAKAHASHI, Dai KITAMOTO, Tomohiro IMURA, Hirosuke OKU, Kensaku ...
    2008 Volume 72 Issue 5 Pages 1199-1205
    Published: May 23, 2008
    Released: May 23, 2008
    [Advance publication] Released: May 07, 2008
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    In order to use liposomes as an efficient carrier of functional food materials, liposomes encapsulating a ukon extract (LUE) were prepared by the mechanochemical method under different conditions, and were physico-chemically and biochemically characterized.
    After a homogenization treatment, the size of LUE decreased with decreasing concentration of the extract from 10 to 2.5 wt %, but did not decrease below 570 nm. LUE were thus subjected to microfluidization. The LUE solutions obtained from less than 5 wt % of the extract remained well dispersed for at least 14 d, whereas those from 10 wt % showed phase separation. With 5 wt % of the extract, the size of LUE obtained at an inlet pressure of 100 MPa was smaller than that obtained at 20 MPa, and reached below 180 nm. Under optimal conditions, resulting LUE was confirmed to be small unilamellar vesicles (SUV) with a diameter of approximately 100 nm by freeze-fracture electron microscopy (FFEM).
    When used for treating simulated gastric and intestinal fluids, LUE obtained by microfluidization showed a 2-fold higher residual rate of curcumin than the uncapsuled extract itself. The bioactivity of LUE was further examined for its suppressive effect on carbon tetrachloride (CCl4)-induced liver injury by using mice. Orally administrated LUE at a dose of 10 mg/kg as the extract had a much higher suppressive effect on the serum aspartate aminotransferase (AST) and alanine aminotransferase (ALT) levels, compared to the uncapsuled extract at a dose of 33 mg/kg.
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  • Tomomi IKEUCHI, Tomomi AOKI, Tadashi YOSHIDA, Koji TAKAHASHI, Makoto H ...
    2008 Volume 72 Issue 5 Pages 1227-1234
    Published: May 23, 2008
    Released: May 23, 2008
    [Advance publication] Released: May 07, 2008
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    Bovine β-lactoglobulin (BLG) was conjugated with cationic saccharides to improve its functions. We used a polylysine-dextran conjugate (PL-Dex) as the cationic saccharide which had been prepared by the Maillard reaction. The molar ratio of PL:Dex was 1:1. The emulsifying property of PL in the acidic pH range was improved by conjugating with Dex. BLG and PL-Dex were conjugated by using microbial transglutaminase (MTGase), the effective conjugation being confirmed by SDS–PAGE. The molar ratio of BLG:PL-Dex was 1:1. Structural analyses by a fluorescence study, ELISA with monoclonal antibodies and measurement of the retinol-binding activity indicated that the conjugates had almost retained the native structure of BLG. The emulsifying property of BLG in the acidic pH range and in the presence of NaCl was improved by conjugating with PL-Dex. The immunogenicity of BLG was reduced by this conjugation, while the antigenicity of the BLG-PL-Dex conjugate was similar to that of BLG in BALB/c mice.
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  • Hiromi IZAWA, Machiko KOHARA, Koichi AIZAWA, Hiroyuki SUGANUMA, Takahi ...
    2008 Volume 72 Issue 5 Pages 1235-1241
    Published: May 23, 2008
    Released: May 23, 2008
    [Advance publication] Released: May 07, 2008
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    Diesel exhaust particles (DEPs) are particulate matter from diesel exhaust that contain many toxic compounds, such as polyaromatic hydrocarbons (PAHs). Some toxicities of PAH are thought to be expressed via aryl hydrocarbon receptors (AhRs). The male reproductive toxicity of DEPs might depend on AhR activation induced by PAHs. We hypothesized that AhR antagonists protect against the male reproductive toxicity of DEPs. Quercetin is a flavonoid and a well-known AhR antagonist, while onion contains many flavonoids, including quercetin. Hence, we examined whether quercetin and onion have alleviative effects against the male reproductive toxicity induced by DEPs. BALB/c male mice were fed quercetin- or onion-containing diets and received 10 injections of DEP suspension or vehicle into the dorsal subcutaneous layer over 5 weeks. The mice were euthanized at 2 weeks, after the last treatment, and their organs were collected. Daily sperm production and total incidence of sperm abnormalities were significantly affected in the DEP groups as compared with the vehicle group, but the total incidence of sperm abnormalities in the quercetin + DEP-treated mice was significantly reduced as compared with the DEP-treated mice. The numbers of Sertoli cells were significantly decreased in DEP-treated mice as compared with the vehicle-treated mice, but, the numbers of Sertoli cells were significantly increased in the quercetin and the onion + DEP-treated mice as compared with the DEP-treated mice. These results clearly indicate alleviative effects of quercetin and onion against the male reproductive toxicity induced by DEP.
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  • Kazumi OSADA, Maryanne CURRAN, Michio KURACHI, Kenji TSUNODA, Kunio YA ...
    2008 Volume 72 Issue 5 Pages 1249-1256
    Published: May 23, 2008
    Released: May 23, 2008
    [Advance publication] Released: May 07, 2008
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    We had previously found that male mice could be trained to discriminate between the urine odor of aged and young adult (adult) mice. We hypothesized that these odors that characterized the older animals might be inhibited by a mixture of extracts (AAM) of mugwort and mushroom, because previous studies have indicated that these extracts could be used to reduce the intensity of unpleasant body odors. The findings of this chemical study strongly suggest that the AAM function helped to modify the aged mouse urine odor so that it more closely resembled the smell of urine from younger mice. Based on the results of the chemical studies, a set of behavioral experiments were therefore conducted. The results of three sets of generalization trials also strongly supported the results of the chemical studies. Togethers, these results suggest that ingested AAM decreased the intensity of odors associated with aging in mice.
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  • Sang-Myeon LEE, Mi-Young YOON, Hae-Ryong PARK
    2008 Volume 72 Issue 5 Pages 1272-1277
    Published: May 23, 2008
    Released: May 23, 2008
    [Advance publication] Released: May 07, 2008
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    This study was conducted to examine the antioxidative and neuroprotective effects of Paeonia lactiflora pall (PLE). Total phenolic content of PLE was 89.65 mg of gallic acid equivalent per gram of PLE. IC50 values for reducing power, hydrogen peroxide scavenging activity, and 1,1-diphenyl-2-picrylhydrazyl (DPPH) radical scavenging activity were 297.57, 3.33, and 32.74 μg, respectively. The protective effect of PLE against H2O2-induced oxidative damage to PC12 cells was investigated by an 3-(4,5-dimethyl-thiazol-2-yl)-2,5-diphenyl-tetrazolium bromide (MTT) reduction assay and lactate dehydrogenase (LDH) release assay. After 2 h of cell exposure to 0.5 mM H2O2, a marked reduction in cell survival was observed. However, this reduction was significantly prevented by 10–100 μg/ml of PLE. H2O2 also induced severe apoptosis of the PC12 cells, which was indicated by a flow cytometric analysis. Interestingly, the H2O2-stressed PC12 cells that had been incubated with PLE had greatly suppressed apoptosis. The results suggest that PLE could be a candidate for a new antioxidant against neuronal diseases.
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  • Yumi SUGITA, Hitoshi SHIRAKAWA, Ritsuko SUGIMOTO, Yuji FURUKAWA, Michi ...
    2008 Volume 72 Issue 5 Pages 1290-1298
    Published: May 23, 2008
    Released: May 23, 2008
    [Advance publication] Released: May 07, 2008
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    Biotin functions as a coenzyme for four carboxylases involved in energy metabolism in mammals. Besides these classical functions, biotin has novel functions in the cellular processes via the modulation of gene expression. In this study, we examined the alteration of gene expression by biotin administration in the liver of streptozotocin (STZ)-induced diabetic rats. In comparison with the control, the mRNA levels of phosphoenolpyruvate carboxykinase and glucose-6-phosphatase were significantly reduced and glucokinase mRNA was increased 3 h after the administration of biotin or insulin. The expression of hepatocyte nuclear factor 4α, one of the transcription factors responsible for gluconeogenic gene expression, was decreased by biotin at both mRNA and protein levels. In addition, forkhead box O1 and sterol regulatory element-binding protein 1c mRNA expression that was enhanced by the insulin treatment was inversely decreased by biotin. These results indicate that biotin repressed the gluconeogenic genes and their transcription factors via a pathway independent of insulin-signaling and could improve the diabetic condition.
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Food & Nutrition Science Note
Microbiology & Fermentation Technology Regular Papers
  • Sachiko YAMAMOTO, Yosuke MORIHARA, Mamoru WAKAYAMA, Takashi TACHIKI
    2008 Volume 72 Issue 5 Pages 1206-1211
    Published: May 23, 2008
    Released: May 23, 2008
    [Advance publication] Released: May 07, 2008
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    γ-Glutamylmetylamide synthetase (GMAS) of Methylovorus mays No. 9, produced by Eschericia coli AD494 (DE3) harboring pET21aGM, formed theanine from glutamic acid and ethylamine with coupling of the reaction with sugar fermentation of baker’s yeast cells as an ATP-regeneration system. Theanine formation was stimulated by the addition of Mn2+ to the reaction mixture, whereas Mg2+ was less effective. Increases to a certain level in the concentrations of GMAS and the substrates in the mixture were effective in increasing theanine formation, but high concentrations of ethylamine (900 mM or more) inhibited yeast sugar fermentation, and eventually decreased theanine formation. The inhibitory effect of ethylamine was restored by increasing the concentration of potassium phosphate buffer in the mixture. Approximately 600 mM (110 mg/ml) theanine was formed in 48 h in an improved reaction mixture containing 600 mM sodium glutamate, 600 mM ethylamine·HCl, 300 mM glucose, 200 mM potassium phosphate buffer (pH 7.0), 30 mM MgCl2, 5 mM MnCl2, 5 mM AMP, 30 units/ml of GMAS, and 40 mg/ml of yeast cells. The yield of theanine was 100% on the substrates (glutamic acid and ethylamine) and also on the energy source (glucose consumed).
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  • Takashi YAMADA, Yuhki TAKAHAMA, Yasuhiro YAMADA
    2008 Volume 72 Issue 5 Pages 1264-1271
    Published: May 23, 2008
    Released: May 23, 2008
    [Advance publication] Released: May 07, 2008
    JOURNALS FREE ACCESS
    A bacterium that utilizes 2,4,6-tribromophenol (2,4,6-TBP) as sole carbon and energy source was isolated from soil contaminated with brominated pollutants. This bacterium, designated strain TB01, was identified as an Ochrobactrum species. The organism degraded 100 μM of 2,4,6-TBP within 36 h in a growing culture. In addition, it released 3 mol of bromine ions from 1 mol of 2,4,6-TBP during the complete degradation of 2,4,6-TBP in a resting cell assay. Moreover, cells grown on 2,4,6-TBP degraded 2,6-dibromophenol (2,6-DBP), 4-bromophenol (4-BP), 2,4,6-trichlorophenol (2,4,6-TCP) and phenol. Metabolic intermediates were detected in the reaction mixture of an in vitro assay for 2,4,6-TBP, and they were identified as 2,4-DBP and 2-BP. NADH was required for the debromination of 2,4,6-TBP. These results suggest that 2,4,6-TBP is converted to phenol through sequential reductive debromination reactions via 2,4-DBP and 2-BP by this strain.
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  • Yoko OHNO, Toshihiko NAKAMORI, Haitao ZHENG, Shin-ichiro SUYE
    2008 Volume 72 Issue 5 Pages 1278-1282
    Published: May 23, 2008
    Released: May 23, 2008
    [Advance publication] Released: May 07, 2008
    JOURNALS FREE ACCESS
    Malic enzyme [L-malate: NAD(P)+ oxidoreductase (EC 1.1.1.39)] catalyzes the oxidative decarboxylation of L-malic acid to produce pyruvic acid using the oxidized form of NAD(P) (NAD(P)+). We used a reverse reaction of the malic enzyme of Pseudomonas diminuta IFO 13182 for HCO3 fixation into pyruvic acid to produce L-malic acid with coenzyme (NADH) generation. Glucose-6-phosphate dehydrogenase (EC1.1.1.49) of Leuconostoc mesenteroides was suitable for coenzyme regeneration. Optimum conditions for the carboxylation of pyruvic acid were examined, including pyruvic acid, NAD+, and both malic enzyme and glucose-6-phosphate dehydrogenase concentrations. Under optimal conditions, the ratio of HCO3 and pyruvic acid to malic acid was about 38% after 24 h of incubation at 30 °C, and the concentration of the accumulated L-malic acid in the reaction mixture was 38 mM. The malic enzyme reverse reaction was also carried out by the conjugated redox enzyme reaction with water-soluble polymer-bound NAD+.
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  • Kohei ISHIKAWA, Takayuki ASAHARA, Yoshiya GUNJI, Hisashi YASUEDA, Kozo ...
    2008 Volume 72 Issue 5 Pages 1317-1324
    Published: May 23, 2008
    Released: May 23, 2008
    [Advance publication] Released: May 07, 2008
    JOURNALS FREE ACCESS
    Methionine auxotrophic mutants of Methylophilus methylotrophus AS1 expressing a mutant form of dapA (dapA24) encoding a dihydrodipicolinate synthase desensitized from feedback inhibition by L-lysine, and mutated lysE (lysE24) encoding the L-lysine exporter from Corynebacterium glutamicum 2256, produced higher amounts of L-lysine from methanol as sole carbon source than did other amino acid auxotrophic mutants. Especially, the M. methylotrophus 102 strain, carrying both dapA24 and lysE24, produced L-lysine in more than 1.5 times amounts higher than the parent. A single-base substitution was identified in this auxotroph in codon-329 of the open reading frame of metF, encoding 5,10-methylene-tetra-hydrofolate reductase. We constructed a metF disruptant mutant carrying both dapA24 and lysE24, and confirmed increases in L-lysine production. This is the first report to the effect that metF deficient increased L-lysine production in methylotroph.
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  • Chang-Hua SHANG, Fen ZHU, Na LI, Xiang OU-YANG, Liang SHI, Ming-Wen ZH ...
    2008 Volume 72 Issue 5 Pages 1333-1339
    Published: May 23, 2008
    Released: May 23, 2008
    [Advance publication] Released: May 07, 2008
    JOURNALS FREE ACCESS
    A gene encoding 3-hydroxy-3-methylglutaryl-coenzyme A reductase (HMGR) was isolated from a triterpene-producing fungus, Ganoderma lucidum (Reishi or Lingzhi). This report provides the complete nucleotide sequence of the full-length cDNA encoding HMGR and its genomic DNA sequence. The cDNA of the HMGR (GenBank Accession no., EU263989) was found to contain an open reading frame (ORF) of 3,681 bp encoding a 1,226-amino-acid polypeptide, whereas the HMGR genomic DNA sequence (GenBank Accession no., EU263990) consisted of 4,262 bp and contained seven exons and six introns. The deduced amino acid sequence of G. lucidum HMGR showed significant homology to the known HMGRs from Ustilago maydis and Cryptococcus neoformans, and contained four conserved domains. Gene expression analysis showed that the expression level was relatively low in mycelia incubated for 10, 12, and 14 d, and reached the highest level in the primordia. Functional complementation of Gl-HMGR in a HMGR-deficient mutant yeast strain indicated that the cloned cDNA encoded a HMG-CoA reductase.
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