Dissimilatory nitrate reductase (DNR) in Escherichia coli cells grown under anaerobic conditions with KNO₃ was activated upon incubation of the cells with NaCl and inactivated with cysteine or glutathione (GSH). Cysteine was shown to exert its effect, at least in part, after conversion to GSH. NaCl-induced activation of DNR was prevented by concomitant addition of the extracts of many kinds of vegetable, although none of the extracts inactivated it when added in the absence of NaCl. Further, the vegetable extracts, in contrast to cysteine or GSH, were found to inhibit DNR activity itself when added to the reaction mixture for the enzyme assay.

View full abstract

The mechanism for the ameliorating effect of dietary fiber on rat intestines was investigated by using Tween 20 and amaranth as toxic chemicals. Ingestion of the diet containing 10% Tween 20 or 5% amaranth induced marked decreases in the body weight gain, food consumption, and segmental sucrase activity in the small intestine. It also induced severe diarrhea in the rats. These adverse effects of the chemicals were prevented by the concurrent ingestion of Gobo dietary fiber (GDF). We found that the amount of concanavalin A-binding glycoprotein (CBGP) in the feces of rats which had ingested the chemicals with GDF was much larger than in that of the rats fed only on the basal diet. In addition, the toxic chemicals or GDF alone in the diet resulted in a definite change in the electrophoretic profile of fecal CBGP, while the concurrent ingestion of GDF and the chemicals produced no change in the electrophoretic profile. These results suggest that renewal of the epithelium caused by the addition of dietary fiber overcome the adverse effects from solubilization

View full abstract

TTUR 2-2, an alkalophilic Bacillus strain isolated from soil, grew well in media containing cholic acid (CA) at 5% or higher and efficiently converted 7α- and 12α-hydroxyl groups of CA to keto groups, with the conversion rate for both hydroxyl groups reaching 100% by 72 hours of cultivation. The strain also converted a 3α-hydroxyl group to a keto group, but the conversion rate was about 5% at 72 hours. The strain neither affected any other part of the CA molecule, nor oxidized 7β- or 12β-hydroxyl groups. By NTG mutagenesis, the following mutants were acquired ; (1) converting only the 7α- and 12α-hydroxyl groups, (2) converting only the 12α-hydroxyl group, and (3) converting only the 7α-hydroxyl group. These mutants selectively produce 12-ketochenodeoxycholic acid (12KCDCA), 7-ketodeoxycholic acid (7KDOCA), and 7, 12-diketolithocholic acid (7, I2DKLCA), from CA ; and 7-ketolithochohc acid (7KLCA) from chenodeoxycholic acid (CDCA), respectively, at high yields, close to 100%.

View full abstract

The purification and some properties of two types of lipase (Lipase I and Lipase II) from Rhizopus niveus are described. The enzymes were purified to homogeneity by column chromatographies on DEAE-Toyopearl (1 pass) and CM-Toyopearl (2 passes). Lipase I consists of two polypeptide chains [a small peptide with sugar moiety (A-chain) and a large peptide of molecular weight 34, 000 (B-chain)]. Lipase II has a molecular weight of 30, 000 consisting of a single polypeptide chain. Lipase I appeared to be converted to Lipase II by limited proteolysis by a specific protease a small amount of which is in the culture supernatant from Rh. niveus, because one of the peptides formed has the same N-terminal sequence and C-terminal amino acid as Lipase II, as well as the molecular mass estimated by SDS-PAGE. Lipase I had a pH optimum of 6. 0-6. 5 and a temperature optimum of 35°C, while, for Lipase II these values were pH 6. 0 and 40°C. Both enzymes were obtained in the crystalline state using the hanging drop method of vapor diffusion and PEG as the precipitating agents.

View full abstract

A photoactivatable heterobifunctional fluorescent reagent, 1-azido-5-naphthalene sulfonyl (ANS) hydrazide, was synthesized and characterized. ANS-hydrazide reacted with lactose to form a photoactivatable hydrazone. The derivative (ANS-lactose) had the same binding affinity for a conger eel lectin as lactose judging from the hemagglutinating-inhibition assay with rabbit erythrocytes. ANS-lactose was used for photoaffinity labeling of a conger eel lectin. The photolabeled lectin was digested with chymotrypsin to isolate photolabeled peptides by reversed-phase HPLC by monitoring fluorescence. A major labeled peptide was located at positions 31-45 in the lectin by amino acid analysis and N-terminal sequencing. The identified segment was close to the highly conserved region throughout animalβ-galactoside-binding lectins.

View full abstract

Bacillus sphaericus DR-1-189[pBHB5022] was found to have an electron transport chain including at least three cytochromes with absorption maxima at 552, 563, and 602 nm, which were reduced by succinic acid, NADH, or NADPH. Electron transport inhibitors such as antimycin A, 3, 3', 5'-triiodo-L-thyronine and 1-(2'-thenoyl)-3, 3, 3-trifluoroacetone (TTFA), blocked this electron transport, with resultant inhibition of cell growth and of biotin synthesis by the cells. When DR-1-189[pBHB5022] was mutagenized to yield mutants resistant to TTFA, the amounts of cytochrome 552 and those of biotin secreted increased in proportion to the extent of resistance of cells to TTFA. A mutant resistant to 250 μg/ml of TTFA produced 48mg biotin per liter in a medium containing pimelic acid. These results suggest that biotin production by B. sphaericus DR-1-189[pBHB5022] is closely related to its electron transport chain.

View full abstract

The function of the gene, act, which encodes lipase activator and that is downstream from the gene that encodes lipase, lip, in Pseudomonas sp. strain KWI-56, was studied with Escherichia coli as the host organism. E. coli carrying both the lip and act genes in either cis or trans produced an active lipase, but E. coli carrying only the lip gene produced an equal amount of inactive lipase protein. The active and inactive lipases had the same molecular weight and the same N-terminal amino acid sequence, so their primary structures were the same. The inactive lipase was not activated spontaneously, but it was activated in vitro by the addition of a crude cell extract of E. coli carrying an expression plasmid of the act gene. Lipase from Pseudomonas sp. strain KWI-56 was completely without enzymatic activity when denatured, but regained activity in vitro upon addition of the crude cell extract. These results indicate that the act gene affects the activation involved in the conformational change of the lipase protein to its active form.

View full abstract

Several purified or commercially available alcohol oxidoreductases of different kinds were tested for their ability to convert the racemic, glycerol-based C₃-synthons glycidol (2, 3-epoxy-1-propanol) and solketal (2, 2-dimethyl-4-(hydroxymethyl)-1, 3-dioxolane), with adequate activity and enantioselectivity. Quinohaemoprotein alcohol dehydrogenases (enzymes containing haem c as well as pyrroloquinoline quinone (PQQ) as cofactors) appeared to be excellently suited for such use. The bacteria from which the enzymes were purified had the same enantiomer preference and had an efficient respiratory chain for reoxidation of these dehydrogenases. In some cases, however, whole cells gave a lower enantiomer ratio (E) than the pure enzyme. NAD-dependent alcohol dehydrogenases also are present in these bacteria, but their presence may not explain the lower ratio because they oxidized the C₃-synthons little if at all. It seems, therefore, that different kinetic mechanisms are responsible for the discrepancy between the effects of whole cells and purified enzymes.

View full abstract

Age-associated changes in several antioxidative factors were studied in blood and liver from mice prone to accelerated senescence aged 0. 7 to 1. 4 years and mice resistant to accelerated senescence aged 1. 0 to 2. 1 years. In both strains, activities of hepatic glutathione peroxidase (GSH-Px) and glutathione-S-transferase (GST) decreased significantly with age, and the activity of superoxide dismutase (SOD), and the concentration of glutathione (GSH) in liver and blood cells tended to decrease with age. The GSH concentration in the plasma and blood cells, and the hepatic GST and SOD activities were significantly lower in the mice prone to senescence than other strain, but the hepatic GSH concentration and GSH-Px activity were similar in the two strains. These results suggest that GSH and hepatic antioxidative enzymes are associated to some extent with senescence in mice with accelerated senescence.

View full abstract

To discover the specificity of Streptomyces β-xylanase toward xylan having glucose stubs, glucoxylan was prepared by the hydrogenation of cotton-seed cake glucuronoxylan. The glucoxylan was hydrolyzed by the β-xylanase of Streptomyces olivaceoviridis E-86, and three kinds of glucoxylo-oligosaccharides were isolated from the hydrolysate by chromatographies on a charcoal column, preparative paper partition, and a Toyopearl HW-40F column. The isolated oligosaccharides had the structures of 2³-α-glucopyranosylxylotriose, 2³-α-(4-O-methyl-glucopyranosyl)xylotriose and 2⁴-α-glucopyranosylxylotetraose. From the structure of the above oligosaccharides and the results of our previous studies, we suggest that the specificity of Streptomyces β-xylanase toward glucose stubs is the same as that toward glucuronic acid stubs, but differs considerably from that toward arabinose stubs.

View full abstract

A bacterial arginase was purified to homogeneity from a strain of Bacillus brevis. The native enzyme, with an estimated MW of 143, 000, migrated on SDS-PAGE as a single polypeptide of estimated MW of 33, 000. The enzyme, highly specific to L-arginine, showed the maximum activity at pH 11. 0 in the presence of Mn²⁺ ions and the p^I was 4. 8 by isoelectric focusing. The enzyme activity was increased significantly by the addition of Mn²⁺, Ni²⁺, or Co²⁺ ions, and inhibited potently by chemicals such as HgCl₂, N-bromosuccinimide, or glutathione. The K_ms for L-arginine and L-canavanine were 0. 69 and 22. 2 mM, respectively. The enzyme was inhibited competitively by γ-guanidinobutyric acid, and non-competitively by L-lysine, L-ornithine, creatine, blasticidin S, and edeine B₁. Analysis of the N-terminal amino acid sequence of the purified bacterial enzyme found 33-36% homologies with the Agrobacterium, yeast, rat, and human enzymes.

View full abstract

Addition of small amounts of calcium ion markedly accelerated the transesterification of N-acetyl-L-tyrosine methyl ester to its ethyl ester by the catalysis of α-chymotrypsin in organic solvents. Maximum increase of the reaction rate was about 12-fold in the presence of 25 μM of calcium ion in ethanol. The rate increase was strongly dependent on calcium ion concentration and nature of organic solvents. Esterification of N-acetyl-L-tyrosine and hydrolysis of N-acetyl-L-tyrosine ethyl ester by α-chymotrypsin in organic solvents were also accelerated by calcium ion. The reactions obeyed Michaelis-Menten kinetics, and the acceleration of the reactions was due to the increase in k_cat.

View full abstract

The transformation of schizandrin (I) into gomisin A (II) was accomplished by use of a combination of biotransformation and chemical reactions. The biotransformation, microbial O-demethylation of I by Cunninghamella echinulata var. elegans (ATCC 9245) produced two novel metabolites [3-norschizandrin (IV) and 2-norschizandrin (VI)] and two known metabolites [gomisin T (III) and 13-norschizandrin (V)]. Among those metabolites, compound III was derived to II by the O-demethylation with a Lewis acid in the presence of an acid scavenger, followed by methylenation.

View full abstract

Chemical modification of Purified D-glucosaminate dehydratase (GADH) apoenzyme by N-ethylmaleimide (NEM) and by 7-chloro-4-aminobenzo-2-oxa-1, 3-diazole (NBDCl) resulted in the time- and concentration- dependent inactivation of the enzyme in each case. The inactivation followed pseudo-first-order kinetics and a double-logarithmic plot of the observed pseudo-first-order rate constant against reagent concentration proved evidence for an approximately first-order reaction, suggesting that the modification of a single cysteine residue per mole of enzyme resulted in inactivation. Amino acid analysis of the NEM-inactivated enzyme showed that three moles of cysteine residues among six moles per mole of subunit were modified under these conditions, therefore one of the three cysteine residues modified by NEM may be essential for activity. Pyridoxal 5'-phosphate (PLP) and D-glucosaminate (GlcNA) protected the enzyme against inactivation by NEM and NBDCl. The apoenzyme was inactivated by EDTA and activity of enzyme was restored by incubation with Mn²⁺ in the presence of PLP. Incubation of the EDTA-treated enzyme with NEM inhibited the restoration of activity. These results suggest that one of the cysteine residues of GADH may be chelated to a Mn²⁺ at or near the active site of GADH, contributing to formation of the active enzyme.

View full abstract

Male Sprague-Dawley rats, 4 weeks (young) or 8 months (adult) of age, were fed one of three purified diets free of or containing either 0. 5% cholesterol or 0. 5% oxidized cholesterol (92% oxidized cholesterol) for 3 weeks. Feeding of oxidized cholesterol caused a significant reduction of food intake, body weight gain, and relative liver weight in rats of both ages. The activity of the HMG-CoA reductase and cholesterol 7α-hydroxylase of liver microsomes, the key enzymes in cholesterol synthesis and catabolism, respectively, was lowered by oxidized cholesterol compared to the diet free of cholesterol in both ages, and the difference was significantly in the adult. On the other hand, the activity of Δ6-desaturase of liver microsomes, a key enzyme in linoleic acid metabolism to arachidonic acid, was significantly increased by oxidized cholesterol in adult rats, leading to the increase in linoleic acid desaturation index [(20 : 3n-6+20 : 4n-6)/18 : 2n-6] in liver phospholipids. Oxidized cholesterol reduced the concentration of cholesterol in serum and liver. Also, the fecal excretion of acidic steroids was lower in rats fed the oxidized cholesterol diet than in those fed the cholesterol-free diet. Thus, oxidized cholesterol significantly influenced cholesterol and fatty acid metabolism in particular in adult rats.

View full abstract

Dry seeds of soybean [Glycine max (L. ) Merr. var. Bay] were irradiated by X-rays (21. 4kR) and the M₂ generations were examined for the oleic acid content in their oil. The genetic variability of oleic acid content in the oil of the Bay variety was significantly increased by the X-ray treatment when compared with the Bay control. A mutant, designated as M23, was selected from 2747 M₂ plants, having an oleic acid content of 46. 1%, twice as much as that in the normal Bay variety. A distinct inverse relationship was observed between the oleic and linoleic acid contents in M23. Mutant M23 isolated from the M₂ generations was confirmed to be always associated with a high oleic acid content under different environmental conditions in the M₃ generations.

View full abstract

The effects of amino acid mixtures simulating to casein and soybean protein on insulin- and glucagondependent linoleic acid desaturation were studied in rat hepatocytes of up to 72 h of culture. The desaturation increased with the insulin molarity in the medium, but decreased with the glucagon molarity. The soybean protein type of amino acid mixture, compared to the casein type of amino acid mixture, potentiated the insulin-dependent desaturation. Thus, dietary proteins could moidify in vivo insulin-dependent intracellular metabolism through postprandial amino acids in blood plasma.

View full abstract

Cells at the apical part of developing stolons of the potato (Solanum tuberosum L. cv. Norin 1) were analyzed for the occurrence of putative precursors to amyloplasts, designated "amyloplast initial." Ultrastructural studies showed that the cells contained the expected novel organelle. It was about 1μm in diameter, devoid of thylakoid membranes, and was stained to a similar extent as the stroma of amyloplasts by uranyl acetate and lead citrate. Formation of thylakoid membranes and starch granules takes place at an early stage of development of these initials when they are just a few μm in diameter. At this stage, proliferation of the initials takes places by division at random sites.

View full abstract

To express the cloned β-amylase cDNA in Escherichia coli under control of the tac promoter, a plasmid pBETA92 was constructed. The plasmid consisted of 6312 bp. An extract of E. coli JM109 harboring pBETA92 had β-amylase activity that produced β-maltose from soluble starch. The enzyme production started in the logarithmic phase, increased linearly, and reached a maximum after 12 h. The recombinant barley β-amylase gave two major (pI 5. 43 and 5. 63) and four minor (pI 5. 20, 5. 36, 5. 80, and 6. 13) activity bands on isoelectric focusing, and their pIs didn't change throughout the incubation. But Western blot analysis found that one β-amylase having a molecular weight of about 56, 000 was synthesized. The recombinant β-amylase was purified from the cells by consecutive column chromatography. The purified enzyme gave a single band of protein on SDS-PAGE but showed heterogeneity on isoelectric focusing. The N-terminal amino acid sequence showed that the recombinant β-amylase lacked four amino acids at positions 2-5 (Glu-Val-Asn-Val) when compared with the presumed amino acid sequence of barley β-amylase. Therefore, the recombiant β-amylase consisted of 531 amino acids, and its molecular weight was calculated to be 59, 169. The N-terminal amino acid sequence of the recombinant β-amylase and the nucleotide sequence of the junction position in plasmid pBETA92 indicated that GTG (Val-5 in the case of barley β-amylase)at positions 27-29 from the SD sequence (AGGA) was the translation initiation codon. The properties of the recombinant β-amylase were almost the same as those of barley β-amylase except for the pI and the K_m values for maltohexaose and maltoheptaose. The pI of recombiant barley β-amylase calculated by Genetyx Version 9 based on the presumed amino acid sequence was 5. 60, but the real pIs were 5. 20-6. 13. Therefore, some post-translational reaction(s) might happen after protein synthesis in E. coli cells, and this modification might cause the differences in the pI and the K_m values for maltohexaose and maltoheptaose between the barley and the recombinant β-amylases.

View full abstract

Bruised and substandard persimmons were processed to produce persimmon extracts. Optimal fluorometric conditions (wavelength of excitation, pH range, and concentration) for measurement of the content of phenolic acids were developed in this study. The main component found in persimmon extract was p-coumaric acid.

View full abstract

The digestibility of triacylglycerols (TAGs) comprising branched-chain fatty acids (BCFAs) and the effects on plasma and hepatic lipid levels in rats were investigated by feeding male Sprague-Dawley rats diets containing 5% (w/w) experimental fats for two weeks. Safflower oil (SO), tripalmitoylglycerol (C16TAG), tri-2-methylpalmitoylglycerol (MeC16TAG), and tri-5, 9, 13-trimethylmyristoylglycerol (Me₃C14TAG) were used as experimental fats. The apparent digestibility followed the order of SO > C16TAG > Me₃C14TAG > MeC16TAG. Plasma total and free cholesterol levels were significantly lower in the MeC16TAG-fed rats, while plasma HDL-cholesterol level was lower in the Me₃C14TAG-fed rats. The plasma TAG level was also very low in the Me₃C14TAG group. However, liver lipid level was not influenced by the dietary fat type. 2-Methylpalmitic acid (MeC16) was not detected in liver lipids of the MeC16TAG group although a low level of MeC16 was present in plasma lipids. A low level of 3, 7, 11-trimethyllauric acid (Me₃C12) and high level of 5, 9, 13-trimethylmyristic acid (Me₃C14) existed in plasma and liver lipids of the Me₃C14TAG-fed rats. These results demonstrated that BCFA-containing TAGs affect the lipid metabolism in rats.

View full abstract

The mechanism by which the Escherichia coli pro V promoter is activated more than 100-fold in response to the medium osmolarity, without the help of any known trans-acting activators, is not yet fully understood. In this context, it has recently begun to be realized that structural features, not the primary sequences, of cis-acting DNA elements may be important for transcriptional regulation in prokaryotes. From this point of view, in this study the pro V promoter was characterized by constructing a series of spacer-insertion mutants in a pro V-lacZ fusion on the chromosome. Here it was found that the upstream cis-acting sequence must be positioned stereospecifically with respect to the principal - 35 and - 10 regions for the pro V promoter to be fully activated. In this regard, it was suggested that an overall DNA structure, particularly DNA curvature, is an important cis-acting parameter for activation of the pro V promoter.

View full abstract

The formation of βCD inclusion complex for aliphatic ethyl ester, with ethyl hexanoate as a guest molecule was investigated. Proton T₁ measurements by the IR method and 1D difference nuclear Overhauser enhancement experiments showed that ethyl hexanoate exists with the groups (H-2 and H-3) around the ester bond nearer to the narrower βCD cavity and the other protons are proximal to the H(CD)-3 and H(CD)-5 protons within the cavity with a folded structure. The complexation of βCD was greatly affected by the addition of ethanol. With increasing ethanol concentration, a marked increase in volatility of ethyl hexanoate from the βCD solution was observed ; the volatility in 30% ethanol was about 3 times larger than in 0% one. On the basis of the observed NOE and T₁ values, the lowering of the ability of βCD complexation in ethanol solution is assumed to be caused by the situation shift of guest molecule toward the narrower side or outside of the cavity.

View full abstract

The effects of bile acids on the proliferation and IgM production of human lymphoblastoid Namalwa cells and on the Ig production of rat spleen lymphocytes were examined. Among the free bile acids examined, two dihydroxy bile acids, CDCA and DCA, inhibited the proliferation of Namalwa cells and Ig production by rat spleen lymphocytes at concentrations above 20 μg/ml, while the inhibitory effect of a trihydroxy bile acid, CA, was much weaker. The inhibitory effects of their conjugated bile acids were weaker than those of the free ones, and the DCA derivatives were more toxic than the CA ones. These results suggest that dihydroxy bile acids were more toxic to Ig production by spleen lymphocytes than trihydroxy ones. The effect of bile acids on Ig production by the lymphocytes was examined in the presence of such mitogens as LPS, PHA, ConA, and PWM. As a result, TDCA inhibited their IgG and IgM production at 200μg/ml independently of the mitogen addition, while TCA was almost ineffective. It thus seems likely that the bile acid inhibits the Ig production by spleen lymphocytes through non-specific inhibition of the both T and B cell functions.

View full abstract

We have engineered a conventional yeast, Saccharomyces cerevisiae, to confer a novel biosynthetic pathway for the production of β-carotene and lycopene by introducing the bacterial carotenoid biosynthesis genes, which are individually surrounded by the promoters and terminators derived from S. cerevisiae. β-Carotene and lycopene accumulated in the cells of this yeast, which was considered to be a result of the carbon flow for the ergosterol biosynthetic pathway being partially directed to the pathway for the carotenoid production.

View full abstract

Synthesis of platelet-activating factor (PAF) was stimulated by the treatment of cells with the calcium ionophore A23187 in Saccharomyces cerevisiae. The amount of PAF production increased A23187-concentration- and time-dependently. The maximum PAF synthesis was observed when the yeast cells were treated with 2 μM of A23187 for 5 min, and the PAF amount decreased at the higher concentrations of A23187 and/or for the longer incubation time. Treatment of the cells with 1 mM of PMSF (phenylmethylsulfonyl fluoride), an inhibitor of PAF catabolism, resulted in transient accumulation of PAF ; however, at more than 2mM of PMSF, PAF production was reduced. Synthesized PAF was not released into the extracellular fraction and was found to be cell-associated. In the absence of A23187 or CaCl₂, and in the presence of EGTA, a Ca²⁺ -chelator, no new synthesis of PAF was observed. These results suggested A23187-induced PAF production was Ca²⁺ dependent, and proceed via PAF remodeling pathway. PAF synthesis in response to A23187 was not detectable in the yeast cells in the logarithmic phase, but increased and reached maximum in the stationary phase of the cell growth. These tindings suggest that PAF is related to yeast cell growth.

View full abstract

(2S, 4R, 5S)-2, 4, 6-Trimethyl-5-heptanolide (1), a sex pheromone component for Macrocentrus grandii, was synthesized by starting from methyl (R)-citronellate (2) and employing bromolactonization (10→11)as the key reaction.

View full abstract

Achatin-I (Gly-D-Phe-Ala-Asp) is a tetrapeptide isolated from the ganglia of an African giant snail (Achatina fulica Firussac). The compound and its derivatives were synthesized, and their effects on the action potential of the crayfish giant axon elicited by electrical stimulation were examined by an intracellular recording method. The membrane potential after the action potential had fallen was elevated by some compounds at an acidic but not neutral pH level. An analysis of the structure-activity relationship for the activity to elevate the afterpotential at an acidic pH level, suggests that the factors favorable for activity were the β-turn conformation, a β- or γ-carboxyl group in the C-terminal residue, appropriate hydrophobicity and/or bulkiness in the protecting groups for the α-amino group at the N-terminus, and the number of negative charges in the entire molecule.

View full abstract

tRNA (adenine-1-)-methyltransferase was purified to homogeneity from an extreme thermophile, Thermus thermophilus HB27, by several steps of column chromatographies. The molecular weight of this enzyme was about 60, 000 as analyzed by SDS polyacrylamide gel electrophoresis. K_m for E. coli tRNA^Glu₂ was 100 nM and that for the methyl group donor, S-adenosyl-L-methionine, was 7. 8 μM. The substrate specificity of the enzyme was investigated by using T7 RNA polymerase transcripts and tRNA fragments obtained by partial digestion with RNases. The enzyme was able to transfer the methyl group to the 3'-half fragment of E. coli initiator tRNA, however, the extent of methylation was elevated by more than five times when the 5'-half fragment was added and annealed to the 3'-half. This indicates that the main recognition site of the enzyme is within the 3'-half region of tRNA molecule, while the tertiary interaction between the T-loop and the D-loop is very effective for the adequate methylation reaction.

View full abstract

Two kinds of miso, one with added precultured yeast and the other without, were compared with respect to the changes in the concentration of HEMF formed and the number of yeast cells in the process of aging. In miso without added yeast, the HEMF concentration increased with the increasing number of existing yeast cells. In miso without yeast aged for 21 days after the miso mash, 0. 06ppm HEMF was detected when the cell number was 2. 2 x 10₃ cell/g. In yeast-added miso aged for 7 days after the miso mash, no HEMF was detected, although the number of yeast cells was 1. 6 x 10₆ cell/g. In yeast-added miso aged for 14 days after the miso mash, HEMF was first detected. The pH levels of miso without yeast and with added yeast when HEMF was first detected were 5. 59 and 5. 57, respectively. It is suggested that the formation of HEMF in miso containing a high concentration of reducing sugar and salt was related to the growth of yeast and started when the pH level fell to less than 5. 6.

View full abstract

A strain of Bacillus circulans, MCI-2554, which produced high levels of cycloinulo-oligosaccharide fructanotransferase (CFTase), was isolated from soil. The high enzyme activity results in the production of large amounts of cycloinulo-oligosaccharides (cyclofructan) composed of cycloinulohexaose and cycloinuloheptaose. Optimal culture conditions for production of CFTase were identified, and CFTase activity of 1. 89 U/ml was obtained in the culture supernatant.

View full abstract

The effect of the oral administration of large amounts of arginine or lysine on tumor growth and hypercholesterolemia was studied in rats into which hepatoma AH109A cells had been transplanted. Arginine reduced both hepatoma growth and hypercholesterolemia, and lysine tended to have these effects. The oral administration of large amounts of arginine might be useful to decrease cachexia arising from cancer and to inhibit the tumor growth.

View full abstract

We determined capsaicinoids and their free phenolic inermediates in the placenta of two cultivars of Capsicum annuum during the fruit-developing period. One cultivar was highly pungent, while the other had no pungency. Although substantial amounts of free phenolics were detected in both cultivars, they were not correlated with the production of capsaicinoids in either cultivar.

View full abstract

Four phytotoxic compounds (1-4) were obtained from a culture of Bipolaris bicolor El-1, a fungal pathogen of gramineous plants. Compounds 1-3 were respectively identified as cochlioquinone A, cochlioquinone B, and stemphone by spectrometric analyses. Compound 4, named isocochlioquinone A, was found to have an identical molecular formula to that of 1 and was determined to be an intramolecular redox isomer of 1. Compounds 1-4 inhibited the root growth of such plant seedlings as finger millet and rice.

View full abstract

The seeds of Annona squamosa (Annonaceae) furnished squamostanal-A (1), a novel annonaceous acetogenin. The structure of squamostanal-A was established by spectral methods to be (5S)-3-(12-formyldodecyl)-5-methyl-2, 5-dihydrofuran-2-one.

View full abstract

Large-scale preparation of (R)-(-)-1, 3-butanediol (R-BDO), an important chiral synthon, from the racemate by Candida parapsilosis IFO 1396 was investigated. We found that ethanol accumulated during culture enhanced the secondary alcohol oxido-reduction activity of cells. Large-scale preparation of R-BDO was done using a fermentor. 3092g of R-BDO was obtained from the racemate by the use of this strain with 94. 0°C enantiomeric excess.

View full abstract

Methyl and ethyl 3-alkylthiazolidine-4(R)-carboxylates (1-6 and 7-12), which were newly synthesized, inhibited growth of the roots of Brassica campestris L. subsp. rapa Hook fil. et ANDERS. of the methyl esters, methyl 3-methylthiazolidine-4(R)-carboxylate (2) was the strongest inhibitor. Of the ethyl esters, ethyl 3-methylthiazolidine-4(R)-carboxylate (8) was the strongest inhibitor. The chlorophyll concentration in the cotyledons of plants treated with any of the compounds, except for was decreased.

View full abstract

The putative precursors to plastids, "plastid initials, " were present in callus cells of apple. The ultrastructure resembled mitochondria in size but differed from them in that they contained no lamellar structures. They showed amoeboid changes, and small vesicular structures were abundant. Formation of thylakoid membranes and starch granules appeared to start in the developing plastid initials.

View full abstract

Invertase was purified from Torulaspora pretoriensis YK-1 by acid treatment and column chromatography on DEAE-Toyopearl650M and phenyl-Toyopearl 650M to homogeneity. The molecular weight of the purified enzyme was estimated to be 130, 000 by SDS-polyacrylamide gel electrophoresis and 530, 000 by gel filtration. The enzyme contained 50% molecular weight as carbohydrate. Properties of the invertase from T. pretoriensis was similar to external invertase from Saccharomyces cerevisiae.

View full abstract

Four geometric isomers of 4, 6-hexadecadienol and the corresponding -dienal and -dienyl acetate isomers were prepared in order to evaluate their pheromone activity toward male eri-silk moths. We used the electroantennography (EAG)-GLC method to determine EAG activity, with results showing that only 4E, 6Z- and 4Z, 6E-hexadecadienal were active, while the other compounds were found to be totally inactive.

View full abstract

Equilibrium controlled enzymatic peptide syntheses in aqueous-organic media were done with chymotrypsin and trypsin modified with varied fatty acid. Palmitoyl-modified chymotrypsin and trypsin synthesized the peptide at 1. 7 and 2. 4 times higher yield than those unmodified. Since an alteration of the substrate specificity was not observed, the chemical modification did not affect the active site of the enzymes.

View full abstract

We have cloned and determined the nucleotide sequences of full-length cDNA clones for chitinase, CH6 and CH16, whose sizes were about 1. 1 kb with signal peptides, from rice cell suspension cultures. Northern blot analysis with CH16 as a probe showed that the chitinase transcript reached maximum at 8h after elicitor-treatment and their sizes were about 1. 1 kb, demonstrating that these clones were induced by elicitor.

View full abstract

"Terasi" starter was composed of Bacillus brevis, Bacillus pumilus, Bacillus megaterium, Bacillus coagulans, Bacillus subtilis, and Micrococcus kristinae in the proportion of 39. 1%, 26. 1%, 8. 7%, 8. 7%, 8. 7%, and 8. 7%, respectively. Most of the isolates showed properties of high esterase (C4) and esterase lipase activities (C8).

View full abstract

We constructed firedly luciferase mutants from Luciola lateralis in which Ala at position 217 was replaced by each of three hydrophobic amino acid residues (Ile, Leu, and Val). These mutants were superior to the wild-type in thermostability. Especially, the purified Ala217Leu mutant still maintained over 70% of the initial activity after 60min at 50°C. This mutant is the most thermostable firefly luciferase obtained.

View full abstract

To test whether calmodulin is present in the red alga Susabi laver (Porphyra yezoensis), materials was concentrated from this alga, using hydrophobic chromatography. In this case, 0. 25M (NH₄)₂SO₄ should be added to bind the protein to the column in the presence of 5 mM CaCl₂. Calmodulin is present in the marine red alga, although at a very low concentration.

View full abstract

Fungitoxic sesquiterpenoid chokols A, B, C, F, and K, chokolic acid B, and chokolal A were synthesized from chokol G by modifying the side chain.

View full abstract

This study was designed to compare the influences of dietary addition of 2% L-cystine and 2% L-cysteine on plasma ceruloplasmin and cholesterol in rats. The result showed that cystine addition depressed plasma ceruloplasmin activity and increased plasma cholesterol, while cysteine addition did not. This study provides the first evidence indicating the differencial metabolic responses to dietary addition of 2% cystine and cysteine.

View full abstract

We have isolated a novel type of mutant strain of Azotobacter vinelandii designated strain WN101 that does not fix nitrogen in the presence of molybdenum but in the presence of tungsten. WN101 produced about twice as much hydrogen as the wild-type strain OP but its production of ethylene from acetylene was less than ten percent of that by OP.

View full abstract

cis-Jasmone 1, an important olfactive ingredient of jasmine flower oil, was synthesized by starting from methyl 3-formylpropionate 2 in a 24% overall yield in 7 steps.

View full abstract

A new brassinosteroid, (22R, 23R, 24S)-22, 23-dihydroxy-24-methyl-5α-cholestan-3, 6-dione (6) which is termed 3-dehydroteasterone, as well as castasterone (2), typhasterol (3), teasterone (4), and 6-deoxocastasterone (5) were identified from wheat (Triticum aestivum L. ) grains by GC-MS. In addition, the presence of a castasterone epimer and a pair of 6-deoxocastasterone epimers was indicated, although their structures remain undetermined. 3-Dehydroteasterone is considered to be an intermediate in the conversion of teasterone to typhasterol.

View full abstract