Bioscience, Biotechnology, and Biochemistry
Online ISSN : 1347-6947
Print ISSN : 0916-8451
Volume 65 , Issue 8
Showing 1-42 articles out of 42 articles from the selected issue
Organic Chemistry
Biochemistry & Molecular Biology
  • Kazuya ISHIGE, Tomoki HAMAMOTO, Toshikazu SHIBA, Toshitada NOGUCHI
    2001 Volume 65 Issue 8 Pages 1736-1740
    Published: 2001
    Released: July 31, 2002
    JOURNALS FREE ACCESS
    A novel method for synthesizing CMP-NeuAc was established. We first confirmed that the putative neuA gene of Haemophilus influenzae, identified by its whole genome sequence project, indeed encodes CMP-NeuAc synthetase (EC2.7.7.43). The enzyme requires CTP as a cytidylyl donor for cytidylylation of NeuAc. The enzyme was coupled with an enzymatic CTP-generating system from CMP and inorganic polyphosphate as a sole phospho-donor driven by the combination of poly-phosphate kinase and CMP kinase, where phosphorylation of CMP is done by the combined activity expressed by both enzymes, and subsequent phosphorylation of CDP by polyphosphate kinase itself occurred efficiently. When CMP-NeuAc synthetase of H. influenzae, poly-phosphate kinase, and CMP kinase were added to the reaction mixture containing equimolar concentrations (15 mM) of CMP and NeuAc, and polyphosphate (150 mM in terms of phosphate), CMP-NeuAc was synthesized up to 10 mM in 67% yield.
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  • Masahiro NAGASE, M.Murshedul ALAM, Akiko TSUSHIMA, Takumi YOSHIZAWA, N ...
    2001 Volume 65 Issue 8 Pages 1741-1747
    Published: 2001
    Released: July 31, 2002
    JOURNALS FREE ACCESS
    The molecules participating in apoptosis induced by T-2 toxin in human leukemia HL-60 cells were investigated. The rank order of the potency of trichothecene mycotoxins to induce internucleosomal DNA fragmentation was found to be T-2, satratoxin G, roridin A >> diacetoxyscirpenol > baccharin B-5 >> nivalenol, deoxynivalenol, 3-acetyldeoxynivalenol, fusarenon-X, baccharin B-4=vehicle control. Western blot analysis of caspase-3 in T-2-treated cells clearly indicated the appearance of its catalytically active fragment of 17-kDa. Increased caspase-3 activity was also detected by using a fluorogenic substrate, DEVD-AMC. Next, cells exposed to T-2 led to cleavage of PARP from its native 116-kDa form to the 85-kDa product. Moreover, DFF-45/ICAD were cleaved to give a 12.5-kDa fragment via T-2 treatment. T-2 caused the release of cytochrome c from mitochondria into the cytosol. Increased enzymic activity of caspase-9 on LEHD-AMC was shown. These data indicate that T-2-induced apoptosis involves activation of caspase-3 and DFF-40/CAD through cytosolic accumulation of cytochrome c along with caspase-9 activation.
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  • Tamo FUKAMIZO, Tsugihisa YAMAGUCHI, Tomohiro ARAKI, Takao TORIKATA, Ar ...
    2001 Volume 65 Issue 8 Pages 1766-1773
    Published: 2001
    Released: July 31, 2002
    JOURNALS FREE ACCESS
    Binding of a highly de-N-acetylated chitosan to Japanese pheasant lysozyme (JPL), which differs from hen egg white lysozyme (HEWL) by nine amino acid substitutions (including Arg114→His), was investigated by 1H-NMR spectroscopy. The profile of the one-dimensional spectrum of JPL is essentially identical to that of HEWL. Using two-dimensional spectral of JPL and HEWL, several aromatic and aliphatic proton resonances of JPL were assigned by comparison. When a highly de-N-acetylated chitosan (number-average degree of polymerization, about 18; degree of acetylation, 0.04), where the N-acetylated units are predominantly surrounded by de-N-acetylated units (a monoacetylated chitosan), was added to the JPL solution, the NMR signals were clearly affected in Trp28 C5H and Ile98 γCH, as in the case of binding to HEWL. The dissociation constant of the monoacetylated chitosan evaluated from the NMR signal responses was calculated to be 0.23±0.05 mM (−31.5 kJ/mol), which is similar to that of HEWL (0.11±0.02 mM, −33.3 kJ/mol). Thus, the Arg→His substitution of the 114th amino acid, which participates in sugar residue binding at the right-sided subsite F, did not significantly affect the chitosan binding. In addition, the C2H signal of His114 of JPL was not affected by the chitosan binding. These results suggest that the monoacetylated chitosan binds to subsites E and F through the left-sided binding mode.
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  • Ken-ichi HONJOH, Hideyuki SHIMIZU, Noriko NAGAISHI, Hiroko MATSUMOTO, ...
    2001 Volume 65 Issue 8 Pages 1796-1804
    Published: 2001
    Released: July 31, 2002
    JOURNALS FREE ACCESS
    A cryoprotective protein, HIC6, was expressed transgenically in tobacco, a cold-sensitive plant, and the localization of the protein within the cell as well as freezing tolerance of the transgenic tobacco was investigated. For constitutive expression of HIC6 in tobacco, its corresponding gene was subcloned into pBI121. Through the transformation with pBI121/hiC6, fifteen transgenic tobacco lines were acquired, out of which twelve lines expressed the HIC6 protein. None of the transgenic tobacco lines, however, showed significant differences in freezing tolerance from the control plants (wild-type and transformed with pBI121) at −1, −3, and −4°C, with the exception that their freezing temperature was −2°C. In order to increase the accumulation level of HIC6, pBE2113 with a stronger promoter was used. Eight lines expressed the protein out of thirteen lines transformed with pBE2113/hiC6. The accumulation levels of the protein were clearly higher in the tobacco plants transformed with pBE2113/hiC6 than in those with pBI121/hiC6. The HIC6 protein seemed to be localized in mitochondria of the transgenic tobacco plants. Freezing-tolerance test at −1 - −4°C showed that the degree of electrolyte leakage was significantly lower in the plants with pBE2113/hiC6 than in the control plants. A leaf browning observation also showed that high accumulation of HIC6 significantly suppressed injury caused by freezing to the transgenic tobacco at −3°C.
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  • Machiko NAKAMURA, Natsuko KUROIWA, Yoshiki KONO, Akira TAKATSUKI
    2001 Volume 65 Issue 8 Pages 1812-1823
    Published: 2001
    Released: July 31, 2002
    JOURNALS FREE ACCESS
    Clofibrate-induced retrograde Golgi membrane movement was blocked or retarded when NRK cells were treated with sodium azide/2-deoxyglucose, nocodazole, taxol, and destruxin B, indicating that it depends on energy, and the dynamic state of microtubules, and being acidic or vacuolar-type ATPase function. PDMP and phospholipase A2 inhibitors also blocked it. These characteristics are similar to those of brefeldin A (BFA) and nordihydroguaiaretic acid (NDGA), inducers of retrograde Golgi membrane movement. However, clofibrate was distinguished from BFA in that BFA action was insensitive to phospholipase A2 inhibitors and from NDGA in that NDGA stabilized microtubules against nocodazole and its action was almost insensitive to taxol. The trans Golgi network (TGN) was resistant to clofibrate, while BFA and NDGA dispersed it. To our knowledge, clofibrate is the first drug to show such different effects on the Golgi and TGN and, therefore, is expected to be a useful tool to distinguish their architecture and/or membrane dynamics.
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  • Hiroshi TSUJIBO, Chiaki TAKADA, Akihiko TSUJI, Mitsuo KOSAKA, Katsushi ...
    2001 Volume 65 Issue 8 Pages 1824-1831
    Published: 2001
    Released: July 31, 2002
    JOURNALS FREE ACCESS
    The intracellular β-xylosidase was induced when Streptomyces thermoviolaceus OPC-520 was grown at 50°C in a minimal medium containing xylan or xylooligosaccharides. The 82-kDa protein with β-xylosidase activity was partially purified and its N-terminal amino acid sequence was analyzed. The gene encoding the enzyme was cloned, sequenced, and expressed in Escherichia coli. The bxlA gene consists of a 2,100-bp open reading frame encoding 770 amino acids. The deduced amino acid sequence of the bxlA gene product had significant similarity with β-xylosidases classified into family 3 of glycosyl hydrolases. The bxlA gene was expressed in E. coli, and the recombinant protein was purified to homogeneity. The enzyme was a monomer with a molecular mass of 82 kDa. The purified enzyme showed hydrolytic activity towards only p-nitrophenyl-β-D-xylopyranoside among the synthetic glycosides tested. Thin-layer chromatography analysis showed that the enzyme is an exo-type enzyme that hydrolyze xylooligosaccharides, but had no activity toward xylan. High activity against pNPX occurred in the pH range 6.0-7.0 and temperature range 40-50°C.
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  • Hidekazu KATAYAMA, Tsuyoshi OHIRA, Koji NAGATA, Hiromichi NAGASAWA
    2001 Volume 65 Issue 8 Pages 1832-1839
    Published: 2001
    Released: July 31, 2002
    JOURNALS FREE ACCESS
    In crustaceans, molt-inhibiting hormone (MIH) is presumed to regulate molting through suppressing synthesis and/or secretion of ecdysteroids by the Y-organ. Recently, a recombinant MIH of the kuruma prawn Penaeus japonicus was produced in E. coli. To approximate the secondary structure of native and recombinant MIH of P. japonicus containing six cysteine residues, the arrangements of disulfide bridges in both MIHs were determined by characterizing their enzymatic digests, and their circular dichroism spectra were measured. The arrangements of disulfide bonds in both MIHs were determined to be identical, and they were linked between Cys7 and Cys44,Cys24 and Cys40, and Cys27 and Cys53. The circular dichroism spectra of both MIHs were very close, and demonstrated that they were rich in α-helix. α-Helix contents in native and recombinant MIHs were calculated to be 49.3% and 46.0%, respectively. All these results strongly suggested that the recombinant MIH was folded in the same manner as the native MIH.
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  • Hirotaka INOUE, Noriaki OZAKI, Hiromichi NAGASAWA
    2001 Volume 65 Issue 8 Pages 1840-1848
    Published: 2001
    Released: July 31, 2002
    JOURNALS FREE ACCESS
    Organic matrices in calcified hard tissues have been considered to control calcification. A matrix peptide, designated CAP-1, was extracted and purified by anionexchange and reverse-phase high performance liquid chromatographies from the exoskeleton of the crayfish, Procambarus clarkii. The amino acid sequence of CAP-1 was determined by mass spectral and sequence analyses of the intact peptide and its enzymatically digested peptides. CAP-1 consisted of 78 amino acid residues, including a phosphoserine residue, and was rich in acidic amino acid residues. CAP-1 had a RebersRiddiford consensus sequence, which is conserved in cuticle proteins from many arthropods. CAP-1 inhibited precipitation of calcium carbonate in an in vitro anticalcification assay dose-dependently, and completelyinhibited it at 3×10-7 M. CAP-1 also showed chitinbinding ability, indicating that this molecule was bifunctional and played an important role in formation of the exoskeleton.
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  • Arisa HIGA, Tomoe HIDAKA, Yuji MINAI, Yoshitaka MATSUOKA, Minoru HAGA
    2001 Volume 65 Issue 8 Pages 1852-1855
    Published: 2001
    Released: July 31, 2002
    JOURNALS FREE ACCESS
    The relationship between active oxygen radicals and peroxidase induction on disease resistance in rice blades was investigated. Nitric oxide was produced in the whole blade stimulated by blast fungus elicitor. The induction of peroxidase activity was detected in active oxygen radical-treated rice blades 1 hour after treatment and thereafter. These results suggest that active oxygen radicals produced by stimulation with the elicitor could trigger peroxidase induction.
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  • Susumu MASUDA, Hiroshi KAMADA, Shinobu SATOH
    2001 Volume 65 Issue 8 Pages 1883-1885
    Published: 2001
    Released: July 31, 2002
    JOURNALS FREE ACCESS
    A chitinase activity was detected in fractions of xylem sap collected from the cut surface of cucumber stems. A 28-kDa acidic protein was purified from the active fractions and its N-terminal amino acids sequence was found to be identical to that of a chitinase gene. Cucumber roots produce and secrete an acidic chitinase, one of the PR proteins, into xylem sap and deliver it to aboveground organs.
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  • Tadao OIKAWA, Yasuyuki TSUKAGAWA, Masashi CHINO, Kenji SODA
    2001 Volume 65 Issue 8 Pages 1889-1892
    Published: 2001
    Released: July 31, 2002
    JOURNALS FREE ACCESS
    The transglycosylation of p-nitrophenyl-β-D-cellotrioside to cellotetraose catalyzed by endo-1,4-β-glucanase (cellulase, EC 3.2.1.4) from a psychrotrophic yeast, Rhodotorula glutinis KUJ 2731, was increased by addition of a miscible organic solvent in the reaction mixture. Among various organic solvents tested, acetone was most effective. The transglycosylation activity increased with an increase in acetone concentrations, while hydrolysis activity was suppressed. The transglycosylation preferably occurred at acidic pH with the optimum pH at 2 in 10 mM Gly-HCl buffer. The optimum temperature of transglycosylation was found to be 50°C in the presence of 40% acetone.
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  • Lenka WEIGNEROVÁ, Jaroslav SPÍZEK, Lucie NAJMANOVÁ ...
    2001 Volume 65 Issue 8 Pages 1897-1899
    Published: 2001
    Released: July 31, 2002
    JOURNALS FREE ACCESS
    Lincomycin (1), a glycosidic antibiotic, active against Gram-positive bacteria, was modified enzymatically with the aim of improving its physico-chemical and biological properties. Compound 1 was glycosylated using jack bean α-mannosidase to produce 7-O-α-D-mannopyranosyl-lincomycin (2).
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  • Wataru KITAGAWA, Asaka SUZUKI, Toshihiro HOAKI, Eiji MASAI, Masao FUKU ...
    2001 Volume 65 Issue 8 Pages 1907-1911
    Published: 2001
    Released: July 31, 2002
    JOURNALS FREE ACCESS
    To address the multiplicity of aromatic ring hydroxylation dioxygenases, we used PCR amplification and denaturing gradient gel electrophoresis (DGGE). The amplified DNA fragments separated into five bands, A to E. Southern hybridization analysis of RHA1 total DNA using the probes for each band showed that band C originated from a couple of homologous genes. The nucleotide sequences of the bands showed that bands A, C, and E would be parts of new dioxygenase genes in RHA1. That of band B agreed with the bphA1 gene, which was characterized previously. That of band D did not correspond to any known gene sequences. The regions including the entire open reading frames (ORFs) were cloned and sequenced. The nucleotide sequences of ORFs suggested that the genes of bands A,C, and E may respectively encode benzoate, biphenyl, and polyhydrocarbon dioxygenases. Northern hybridization indicated the induction of the gene of band A by benzoate and biphenyl, and that of the gene of band C by biphenyl and ethylbenzene, supporting the above notions. The gene of band E was not induced by any of these substrates. Thus the combination of DGGE and Southern hybridization enable us to address the multiplicity of the ring hydroxylation dioxygenase genes and to isolate some of them.
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  • Masao SAKURAI, Hiroyuki ADACHI, Kazuo SUTOH
    2001 Volume 65 Issue 8 Pages 1912-1916
    Published: 2001
    Released: July 31, 2002
    JOURNALS FREE ACCESS
    GAPA is an IQGAP-related protein and is involved in Dictyostelium cytokinesis. Since mammalian IQ-GAPs are effectors for Rac/Cdc42, GAPA is also predicted to bind to small GTPases, which are to be identified. In this study, mutant GAPAs were examined for functions in cytokinesis by genetic complementation of gapA- cells. Positively charged side chains of Arg442 and Lys474 of GAPA, predicted to be present on the surface of interaction with small GTPases, were found to be essential, suggesting an interaction between GAPA and putative small GTPase in cytokinesis. Also, results from truncated GAPAs indicated that almost the entire region of GAPA homologous to IQGAP is required for cytokinesis in Dictyostelium.
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  • Takeshi SAITO, Norio TANAKA, Takao SHINOZAWA
    2001 Volume 65 Issue 8 Pages 1928-1931
    Published: 2001
    Released: July 31, 2002
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    A transformation vector, pLS-hph, was constructed with the promoter and terminator of the glyceraldehyde-3-phosphate dehydrogenase (GPD) gene derived from an ectomycorrhizal basidiomycete, Lyophyllum shimeji, and with the hygromycin B (HmB) phosphotransferase (hph) gene from Escherichia coli. This vector was introduced into protoplasts of L. shimeji and 3.4 transformants per μg plasmid DNA were obtained. In most of the transformants, multiple copies of the vector were integrated into the genomic DNA. The results indicate that pLS-hph is a useful vector for L. shimeji.
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Food & Nutrition Science
  • Hisashi YOSHIOKA, Yasushi SENBA, Kieko SAITO, Takahide KIMURA, Fumiko ...
    2001 Volume 65 Issue 8 Pages 1697-1706
    Published: 2001
    Released: July 31, 2002
    JOURNALS FREE ACCESS
    A spin-trapping method was applied to examine the formation of the hydroxyl (OH) radical from a tea catechin-Cu(II) system to elucidate a previous result that some tea catechin-Cu(II) systems induced DNA scission. Three tea catechins, (−)-epigallocatechin (EGC), (−)-epigallocatechin gallate (EGCg) and (−)-epicatechin (EC), were used. The spin-trapping agent, 5,5'-dimethyl-pyrroline-1-oxide (DMPO), was dissolved in a pH 9 phosphate buffer solution, then a catechin and Cu(II) were added in that order, and the ESR spectral change was monitored for one hour. The order of adding the catechin and Cu(II) was then reversed, and the ESR spectral change was again monitored to examine the coordinating activity of each catechin toward the Cu(II) ion and the effect on OH radical generation. The intensity changes of the spin adducts, DMPO-OH, DMPO-CH3 and DMPO-H, were analyzed, the results suggesting that the OH radical generated in the system decomposed DMPO, resulting in the formation of DMPO-CH3 and DMPO-H. The results show that EGC formed a stable complex with Cu(II) and generated the OH radical. EGCg seemed to have this activity, but the OH radical that was generated was scavenged by the gallate group existing in the complex. EC did not show strong coordinating and OH-generating activities. These characteristics of the three catechins are consistent with the results shown for DNA scission.
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  • Yoko NISHIKAWA, Yuka TOYOSHIMA, Tadao KURATA
    2001 Volume 65 Issue 8 Pages 1707-1712
    Published: 2001
    Released: July 31, 2002
    JOURNALS FREE ACCESS
    Dehydro-L-ascorbic acid (DAA), an oxidation product of L-ascorbic acid (vitamin C), is unstable in the neutral and basic pH regions. When DAA was incubated in a phosphate buffer with deuterium oxide (pH 7.4), it was degraded to form the main degradation compound, which was identified as 3,4-dihydroxy-2-oxo-butanal (L-threosone). This compound was also formed from diketo-L-gulonic acid (DKG) in a phosphate buffer with deuterium oxide. L-threosone had reducing activity, probably due to its enolization, and is likely to have been involved in the formation of the reducing activity that was observed in aqueous DAA and DKG solutions. As a reactive dicarbonyl compound, L-threosone might also take some role in the cross-linking of tissue proteins that are formed in vivo in the Maillard reaction.
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  • Simon POON, Adrienne CLARKE, Graeme CURRIE, Carolyn SCHULTZ
    2001 Volume 65 Issue 8 Pages 1713-1723
    Published: 2001
    Released: July 31, 2002
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    A peptide derived from apomyoglobin by cyanogen bromide cleavage was found to be an active emulsifier. This molecule, peptide 1-55, has two potential amphipathic α-helices and a hydrophilic C-terminal domain. The importance of this molecule was investigated by testing the products of gene constructs based on the sequence of peptide 1-55, but lacking one of the three domains. The emulsifying activity of the peptides lacking either of the α-helices was correlated with the hydrophobic moments of their respective helices. The hydrophobic moment is a measure of the amphipathicity of α-helices; a hydrophobic moment analysis of other emulsifying peptides supports the hypothesis that a high hydrophobic moment contributes to good emulsifying properties in a molecule which contains α-helices.
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  • Jun WATANABE, Soichi TANABE, Michiko WATANABE, Takanori KASAI, Kei SON ...
    2001 Volume 65 Issue 8 Pages 1729-1735
    Published: 2001
    Released: July 31, 2002
    JOURNALS FREE ACCESS
    Brown Norway rats were immunized with gluten, and then fed a diet containing hypoallergenic fluor or an amino acid mixture. The rats were then made to inhale a solubilized gluten to induce gluten-specific bronchial asthma. The antibody levels in the serum of rats were measured by ELISA, and cell counts were done on cytospin preparations of bronchoalveolar lavage fluid. Body weight was decreased after allergen challenge in rats fed the amino acid mixture but not in rats fed the hypoallergenic flour. Antibody levels in the serum were significantly lower in rats fed hypoallergenic flour than in those fed the amino acid mixture. Differential cell counts in the bronchoalveolar lavage fluid showed that the numbers of eosinophils, lymphocytes, and neutrophils were significantly lower in rats fed the hypoallergenic flour than in those fed the amino acid mixture. These results suggest that hypoallergenic flour actively suppresses the allergic reactions, probably by inducing oral tolerance.
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  • Hisanao TAKEUCHI, Takeshi NAKAMOTO, Yoshiaki MORI, Masako KAWAKAMI, Hi ...
    2001 Volume 65 Issue 8 Pages 1748-1754
    Published: 2001
    Released: July 31, 2002
    JOURNALS FREE ACCESS
    The effects of different types of dietary fat on the activities of hepatic enzymes related to fatty acid synthesis {glucose-6-phosphate dehydrogenase (G6PDH) and acetyl-CoA carboxylase ACC)}, oxidation {acyl-CoA synthetase (AST), carnitine palmitoyl transferase (CPT), and peroxisomal β-oxidation (P βOX)}, and lipogenesis {phosphatidate phosphohydrolase (PAP), diacylglycerol acyltransferase (DGAT), and phosphocholine diacylglycerol transferase (PCDGT)}, and plasma and liver lipid levels were investigated in male Wistar rats. The animals were 6 weeks old and about 120 g of body weight, and were fed on test diets containing 20% of a mixture of tripalmitin, tristearin and corn oil (SFA), olive oil (OLI), sunflower oil (SUN), linseed oil (LIS), and sardine oil (SAR) for 2 weeks. The concentrations of plasma total cholesterol (T-CHOL), high-density lipoprotein-cholesterol (HDL-CHOL), triacylglycerol (TG) and phospholipid (PL) were generally higher in the rats fed on SEA and OLI than in those given SUN, LIS and SAR. The rats fed on OLI had a higher level of liver T-CHOL than those fed on the other fats. The liver TG content was nearly higher from the intake of SFA and OLI than from SUN, LIS and SAR, although the liver PL level was not affected by the type of dietary fat. The SFA and OLI groups had the highest activities of hepatic G6PDH and ACC, and the SAR group, the lowest activities. The activities of AST and CPT, and peroxisomal P βOX in the liver were higher in the rats fed on the LIS and SAR diets than in those given the other diets. The hepatic PAP activity was higher from the intake of OLI and SUN, and tended to be higher from SFA than from LIS and SAR. The activity of liver DGAT was higher from SFA and inclined to be higher from OLI, SUN, and LIS than from SAR, while the PCDGT activity in the liver was not effected by the type of dietary fat. The concentrations of plasma and liver TG were generally positively correlated with the activities of liver enzymes related to the synthesis of fatty acids and lipids, and negatively with those involved in fatty acid oxidation. Based on these results, it is suggested that the levels of plasma and liver TG were controlled by different types of dietary fat through changes in the hepatic enzyme activities related to fatty acid synthesis, lipogenesis, and fatty acid oxidation.
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  • Nobuhiro NAKANO, Norifumi SHIRASAKA, Kazuki MASUOKA, Tetsuo MURAKAMI, ...
    2001 Volume 65 Issue 8 Pages 1859-1863
    Published: 2001
    Released: July 31, 2002
    JOURNALS FREE ACCESS
    The inhibitory effects of such vanillylamides as capsaicin and nine capsaicinoids on fatty acid desaturation in liver cells were investigated by using the cultured rat liver cell line, BRL-3A. When capsaicin was added to the medium, it had a relatively strong inhibitory effect on Δ6 desaturation and clear inhibitory effects on Δ5 and C24Δ6 desaturation (Δ6 desaturation of C24-polyunsaturated fatty acids). Capsaicinoids with side carbon chain lengths of C10:0 and C12:0 expressed the maximum inhibitory effects of the nine capsaicinoids on fatty acid desaturation in the BRL-3A cells. The inhibitory effects of the capsaicinoids were not correlated with their pungency.
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  • Masaki TERAHARA, Saori KURAMA, Naoki TAKEMOTO
    2001 Volume 65 Issue 8 Pages 1864-1868
    Published: 2001
    Released: July 31, 2002
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    Ether extracts of lactic acid bacteria were analyzed for prevention of the oxidation of erythrocyte membrane and human low-density lipoprotein in vivo. Streptococcus thermophilus 1131 and Lactobacillus delbrueckii subsp. bulgaricus 2038, yogurt starters, were chosen as test-strains, and ether extracts of these cultures were used as samples. Both strain 1131 and strain 2038 produced radical scavengers and inhibited oxidation of erythrocyte membranes and low-density lipoproteins. The antioxidative activity of strain 2038 was higher than that of strain 1131.
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  • Osamu CHONAN, Rie TAKAHASHI, Masaaki WATANUKI
    2001 Volume 65 Issue 8 Pages 1872-1875
    Published: 2001
    Released: July 31, 2002
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    Rats fed a diet containing β1-4 linked galactooligosaccharides (GOS) (5 g/100 g of diet) absorbed calcium and magnesium more efficiently than those fed the control diet. However, the increment obtained through GOS-feeding was reduced by neomycin sulfate 0.67 g/100 g of diet). Since the decrease in cecal pH in rats fed GOS was suppressed by neomycin-feeding, bacterial action in the digestive tract was considered to be reduced by neomycin-feeding. Our findings suggest that the action of intestinal bacteria is necessary for the effects of GOS.
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  • Yasuhisa WAKABAYASHI
    2001 Volume 65 Issue 8 Pages 1893-1896
    Published: 2001
    Released: July 31, 2002
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    Germanium-132 (Ge-132) was given at 200 mg/kg of body weight to 8-week-old Kurosawa and Kusanagi hypercholesterolemic (KHC) rabbits. Thirty-six weeks later, the susceptibility of plasma low-density lipoprotein to oxidation and the morphology of atherosclerosis in the aorta and coronary artery were investigated. Treatment with Ge-132 resulted in decreases in the oxidation rate and in the formation rate of thiobarbituric acid-reactive substances following copper-induced oxidation of LDL. Ge-132 is suggeseted to possess antioxidative properties, but this did not lead to any attenuation of atherosclerotic progression in the KHC rabbits.
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  • Puming HE, Yasuhiro NODA, Kimio SUGIYAMA
    2001 Volume 65 Issue 8 Pages 1924-1927
    Published: 2001
    Released: July 31, 2002
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    A coffee extract significantly suppressed lipopolysaccharide (LPS)-induced hepatitis in D-galactosamine-sensitized rats, as assessed by the plasma alanine and aspartate aminotransferase activities, when it was added to the diet (30 g/kg) and fed to rats for 14 days. Its effect was as strong as that of a green tea extract. The coffee extract suppressed LPS-induced hepatitis when singly force-fed (1.2 g/kg) 1.5 h prior to the injection of the drugs, whereas a decaffeinated cofee extract had no significant effect. The hepatoprotective effect of caffeine was stronger than that of theobromine. These results indicate that coffee can protect animals from LPS-induced hepatitis, and that the effect of coffee might be mainly due to caffeine.
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Microbiology & Fermentation Technology
  • Reiji MARUYAMA, Akiko KAWATA, Shin ONO, Mikio NISHIZAWA, Seiji ITO, Ma ...
    2001 Volume 65 Issue 8 Pages 1761-1765
    Published: 2001
    Released: July 31, 2002
    JOURNALS FREE ACCESS
    The soil bacterium Bacillus cereus Tim-r01 efficiently transformed polyaromatic carboxylic acids (PACA) such as 4-biphenylcarboxylic acid (4-BPCA), 4-biphenylacetic acid, and 4-phenoxybenzoic acid into their corresponding amides. The amidation activity was expressed at 37°C (pH 7-8) in the presence of grown cells in nutrients under an aerobic atmosphere. Other strains of B. cereus, IFO 3001 and IAM 1229, also gave the amide from 4-BPCA. In phosphate-buffered saline (PBS), the addition of normal amino acids was essential, while sulfur-containing amino acids such as methionine and cysteine drastically inhibited the amidation. Tracer experiments using N-15-isoleucine and N-15-alanine showed that the nitrogen atom of the amide came from an amino group of amino acids but not from ammonia or alkylamines.
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  • Hyo-Kon CHUN, Yasuo OHNISHI, Norihiko MISAWA, Kazutoshi SHINDO, Miki H ...
    2001 Volume 65 Issue 8 Pages 1774-1781
    Published: 2001
    Released: July 31, 2002
    JOURNALS FREE ACCESS
    The phdABCD gene cluster in a marine bacterium Nocardioides sp. strain KP7 codes for the multicomponent enzyme phenanthrene dioxygenase. phdA encoding an iron-sulfur protein large subunit α, phdB encoding its small subunit β, phdC encoding ferredoxin, and phdD encoding ferredoxin reductase, were replaced in such a way that the termination codons of the preceding open reading frames were overlapped with the initiation codons of the following genes. This manipulated phdABCD gene cluster was positioned downstream of the thiostrepton-inducible promoter PtipA in a high-copy-number vector pIJ6021, and introduced into the gram-positive, soil-inhabiting, filamentous bacterium Streptomyces lividans. The recombinant S. lividans cells converted phenanthrene into a cis-diol form, which was determined to be cis-3,4-dihydroxy-3,4-dihydrophenanthrene by its UV spectral data as well as HPLC property, using the authentic sample for comparison. This biotransformation proceeded very efficiently; 200 μM and 2 mM of phenanthrene were almost completely converted to its cis-diol form in 6 h and 32 h, respectively. In addition, the S. lividans cells carrying the phdABCD gene cluster were found to transform 1-methoxynaphthalene to two products, which were identified to be 8-methoxy-2-naphthol in addition to 8-methoxy-1,2-dihydro-1,2-naphthalenediol by their EI-MS, 1H- and 13C-NMR spectral data.
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  • Jong-Shik SHIN, Byung-Gee KIM
    2001 Volume 65 Issue 8 Pages 1782-1788
    Published: 2001
    Released: July 31, 2002
    JOURNALS FREE ACCESS
    Microorganisms that are capable of (S)-enantioselective transamination of chiral amines were isolated from soil samples by selective enrichment using (S)-α-methyl-benzylamine (S)-α-MBA) as a sole nitrogen source. Among them, Klebsiella pneumoniae JS2F, Bacillus thuringiensis JS64, and Vibrio fluvialis JS17 showed good ω-transaminase (ω-TA) activities and the properties of the ω-TAs were investigated. The induction level of the enzyme was strongly dependent on the nitrogen source for the strains, except for V. fluvialis JS17. All the ω-TAs showed high enantioselectivity (E>50) toward (S)-α-MBA and broad amino donor specificities for arylic and aliphatic chiral amines. Besides pyruvate, aldehydes such as propionaldehyde and butyraldehyde showed good amino acceptor reactivities. All the ω-TAs showed substrate inhibition by (S)-α-MBA above 200 mM. Moreover, substrate inhibition by pyruvate above 10 mM was observed for ω-TA from V. fluvialis JS17. In the case of product inhibition, acetophenone showed much greater inhibitions than L-alanine for all ω-TAs. Comparison of the enzyme properties indicates that ω-transaminase from V. fluvialis JS17 is the best one for both kinetic resolution and asymmetric synthesis to produce enantiomerically pure chiral amines. Kinetic resolution of sec-butylamine (20 mM) was done under reduced pressure (150 Torr) to selectively remove an inhibitory product (2-butanone) using the enzyme from V. fluvialis JS17. Enantiomeric excess of (R)-sec-butylamine reached 94.7% after 12 h of reaction.
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  • Ruamsub CHUMNANTANA, Kumi HIROSE, Hiromichi BABA, Toshiharu YAGI
    2001 Volume 65 Issue 8 Pages 1789-1795
    Published: 2001
    Released: July 31, 2002
    JOURNALS FREE ACCESS
    Conditions for extracellular production of vitamin B6 compounds (B6), especially pyridoxal 5'-phosphate (PLP) by Schizosaccharomyces pombe leu1 strain were examined. The productivity was dependent on concentration of L-leucine in the culture medium: 30 mg/l gave the highest concentrations of total B6 and PLP. The viable cells harvested at different growth phases showed different productivity: middle and late exponential phase cells showed the highest productivity of total B6 and PLP, respectively. D-Glucose (1%, w/v) among other sugars gave the best productivity. Supplementation of air and ammonium sulfate significantly increased extracellular production of PLP. Superoxide anion producers, menadione and plumbagin, and H2O2 increased the productivity of PLP. Cycloheximide inhibited the increase of PLP by the oxidative stress and in contrast, increased pyridoxine.
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  • Yusuke TAKUSAGAWA, Masato OTAGIRI, Sadaharu UI, Takashi OHTSUKI, Akio ...
    2001 Volume 65 Issue 8 Pages 1876-1878
    Published: 2001
    Released: July 31, 2002
    JOURNALS FREE ACCESS
    The L-2,3-butanediol dehydrogenase produced in E. coli JM109/pLBD2-CTC was purified by 5 steps. The molecular mass of this enzyme was estimated at 110 kDa and the subunit was mesured to be 30 kDa. The L-BDH had some differences from the BDHs from other sources in substrate specificity, pI value, pH stability, effects of divalent cations, and organic acids.
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  • Hirokuni TAJIMA, Hisashi KIMOTO, Akira TAKETO
    2001 Volume 65 Issue 8 Pages 1886-1888
    Published: 2001
    Released: July 31, 2002
    JOURNALS FREE ACCESS
    Hydroxy isothiocyanates, especially 2-(4-hydroxy-phenyl)ethyl isothiocyanate (hITC), were examined for antimicrobial synergism with several antibiotics against Escherichia coli and Staphylococcus aureus, using a multiwell plate system. hITC had antibacterial synergism, specifically with aminoglycoside antibiotics. The synergism was observed in synthetic medium (M9 minimal medium) or soybean casein digest broth, but not in nutrient broth. Synergism was seen in the presence of certain sugars such as glucose, fructose, and maltose in the medium.
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  • Yasuhiro KINOSHITA, Yoshikazu YAMAMOTO, Teiko KUROKAWA, Isao YOSHIMURA
    2001 Volume 65 Issue 8 Pages 1900-1902
    Published: 2001
    Released: July 31, 2002
    JOURNALS FREE ACCESS
    Effects of the nitrogen sources in the medium for the production of secondary metabolites in lichens were examined. The usnic acid production by a mycobiont of the lichen Usnea hirta was higher in the liquid medium containing ammonium and nitrate ions than in those containing amino acids.
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  • Shinobu ODA, Hiromichi OHTA
    2001 Volume 65 Issue 8 Pages 1917-1919
    Published: 2001
    Released: July 31, 2002
    JOURNALS FREE ACCESS
    A double coupling system, which couples metabolism of glucose and transacetylation, is a unique procedure for the production of acetic esters. In the novel coupling system described in this article, acetyl coenzyme A (acetyl-CoA) was supplied via metabolism of both glucose and exogenous saturated fatty acids. While short and middle chain fatty acids having C4-8 were very biotoxic, myristic acid (C14) was effectively used as a source of acetyl-CoA.
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