Erbstatin, a tyrosine kinase inhibitor, has antitumor activity on L1210 mouse leukemia. We tried to find the mechanism of this cytotoxic effect of erbstatin. Erbstatin increased trypan blue exclusion in cultured L1210 cells. The cytotoxic action of erbstatin was accompanied with shrinking of the cells and internucleosomal DNA fragmentation, characteristics of apoptosis. The DNA fragmentation was induced with other tyrosine kinase inhibitors as well, methyl 2, 5-dihydroxycinnamate and herbimycin A. In addition, intracellular tyrosine phosphorylation was inhibited by erbstatin before cell death. Erbstatin-induced DNA fragmentation was prevented by zinc ions, but not by cycloheximide or 12-O-tetradecanoylphorbol 13-acetate (TPA). Thus, erbstatin induced apoptotic cell death in L1210 cells by inhibiting tyrosine kinase

View full abstract

Butterfat was interesterified in phosphatidylcholine reverse micellar systems containing hexane at different ratios of water per mol surfactant (W₀), with a commercial lipase (Lipase N, Amano). The amount and composition of free fatty acids were found by gas-liquid chromatography. Triacylglycerols were separated from interestified butterfat by column chromatography on a Supelclean LC-SI column. The purified triacylgylcerols were separated analytically by high-pressure liquid chromatography. The positional distribution of fatty acids was identified by enzymatic deacylation with commercial pancreatic lipase. The addition of up to 2.2% water (W₀ = 17.4) to the reaction mixture had little effect on the interesterification yield, but shifted the reaction equilibrium in favor of hydrolysis by 4-fold. Further additions of water up to 4.4% (W₀ = 34.8) resulted in maximum yield, with a decreased degree of hydrolysis, from 2.2 to 1.8mmol of free fatty acids per gram of butter. The interesterified butterfat triacylglycerols had an increased amount of palmitic acid (C16 : 0) at the sn-2 position with a decrease in the proportions of small chain fatty acids (C4-C10).

View full abstract

Alkyl β-D-glucosides and n-alcohols were separated by high-performance liquid chromatography on a porou gel of a trimethylolpropane trimethacrylate homopolymer, using a mixture of water with methanol or acetonitrile as the eluent. The effect of the methanol and acetonitriIe concentrations on the separability was examined, and the optimum concentration of each to separate alkyl glucosides and alcohols with alkyl chain lengths of six to twelve was determined. A model for analyzing the elution characteristics of the solutes for various levels of methanol or acetonitrile is proposed, based upon the chemical potential of the solutes and eluent at the interface between the gel and eluent phases. Conventional column chromatographic separation of an alkyl glucoside and an alcohol was also performed by using gel with a larger particle diameter.

View full abstract

An extracellular, novel alkaline lipase produced by Pseudomonas fluorescens AK102 was purified by ultrafiltration, ammonium sulfate precipitation, and DEAE-Toyopearl 650M and Phenyl-Toyopearl 650M column chromatographies. The purified enzyme was homogeneous on SDS-PAGE. The molecular weight was estimated to be about 33, 000 by SDS-PAGE. The isoelectric point was pH 4.0 by isoelectric focusing. The pH stability was 4 to 10 and the optimum pH was 8 to 10. The optimum temperature was 55°C and the enzyme was stable below 50°C. The enzyme unspecifically liberated short chain to long chain fatty acids from p-nitrophenyl esters, methyl esters, and triglycerides. In the presence of an anionic surfactant, the enzyme was characteristically stable. These results suggested that the enzyme can be used as a home laundry product ingredient.

View full abstract

The gene encoding L-lactate dehydrogenase (LDH) was cloned from an industrial dairy strain of Streptococcus thermophilus M-192 using a synthetic oligonucleotide probe based on the N-terminal amino acid sequence of the purified enzyme, and its nucleotide sequence was determined. The enzyme was deduced to have 328 amino acid residues with a molecular weight of 35, 428 and found to have high sequence similarity to LDHs from other lactic acid bacteria (89.0% to Streptococcus mutans, 76.3% to Lactococcus lactis subsp. lactis, 67% to Lactobacillus casei, and 60% to Lactobacillus plantarum). The gene contained a promoter-like sequence similar to the Escherichia coli promoter consensus, and expression of the S. thermophilus LDH gene was observed in E. coli cells.

View full abstract

Factors affecting heterotrophic nitrification by Alcaligenes faecalis OKK17, which was isolated from sewage sludge, were examined. Specific nitrifying activity increased as the pH increased up to 8.5. Most of the nitrogenous compounds (88%) in the culture supernatant were converted to hydroxylamine or nitrite at pH 9 but 87% of them remained as ammonium at pH 7. These results imply that the substrate for heterotrophic nitrification is ammonium and that the organism oxidizes ammonium to lower its toxic effect. Although the addition of acetate to a defined medium increased growth of the bacterium up to C/N = ca. 6, the accumulation of nitrification products almost paralleled the growth and the specitic nitrifying activity decreased. Pyruvate and oxaloacetate increased the specific nitrifying activity six- to eightfold compared with the other organic acids examined, but the key enzyme activities in the glyoxylate cycle were not increased. Acetate, glyoxylate, and malonate did not increase the specitic nitrifying activity, but they increased the enzyme activities. These results imply that the involvement of acetate metabolism in the heterotrophic nitrification is unlikely.

View full abstract

FR901228, a novel antitumor antibiotic, reversed the transformed morphology of the Ha-ras transformants, Ras-1 cells, and inhibited their growth. The reduction of c-myc expression was observed in FR901228-treated Ras-1 cells by RNA dot-blot hybridization. This reduction of c-myc expression and morphological reversion of the transformed cells to normal were correlated with growth inhibition (G₀/G₁ arrest in cell cycle).

View full abstract

The effect of an agent that lowers the taurine concentration of various tissues, guanidinoethane sulfonate (GES), on the activities of enzymes involved in sulfur amino acid metabolism in the tissues of rats fed a taurine-free puritied diet was evaluated. Rats given 1% GES in drinking water profoundly decreased taurine concentration in various tissues. GES slightly but significantly increased the concentration of glutathione in liver, kidneys, and heart. The drug caused decreases in the hepatic activities of cysteine dioxygenase and cysteinesultinic acid decarboxylase which are involved in a metabolic pathway for the conversion of cysteine to taurine. However, the renal and brain cysteinesulfinic decarboxylase activities were not affected by the drug. This drug did not change the activities of cystathionine synthase and γ-glutamyIcysteine synthetase in the liver and kidneys. GES treatment slightly increased the renal but not the hepatic aspartate aminotransferase activity. The depression of taurine synthesis in the liver may be one factor accounting for the potency of this drug to reduce taurine concentration in tissues.

View full abstract

Colony groups that form large amounts of slime externally were obtained from activated sludge among phthalate-assimilating microbes. That slime, which R-3 mixed microbes produced externally, had a high level of flocculation activity. R-3 mixed strains, one of such colony groups, efficiently produced a bioflocculant (APR-3) in liquid cultures of production medium, especially that containing starch and glucose (1 : 1) as carbon sources. Identitication of this group of bioflocculant-producing microbes (R-3 mixed strains) showed that it was comprised of four strains belonging to the genera Oerskovia, Acinetobacter, Agrobacterium, and Enterobacter. The bioflocculant (APR-3) was purified electrophoretically to homogeneity by ethanol and CPC precipitation. Its molecular mass is at least 2×10⁶ Da. APR-3 is an acidic polysaccharide made of glucose, galactose, succinic acid, and pyruvic acid (molar ratio : 5.6 : 1 : 0.6 : 2.5).

View full abstract

When beer was stored for 0, 1, 2, 3, and 4 weeks at 37°C, a stale flavor was generated with the increasing storage period. The 132 peaks detected by a GC analysis of the medium-and high-boiling-point volatiIes in beer were used as variables for a multivariate analysis. Fifteen compounds, including furfural and 2-furfurylethyl ether, which had rotated factor loadings of more than 0.5 on a second factor axis, were extracted as components that were highly correlated with the formation of the stale flavor. Furfuryl acetate, 5-methylfurfural, and an unknown compound were extracted by a stepwise discriminant analysis as important indices for the discrimination of beer according to storage period.

View full abstract

The bacterial isolates WS, WL, and YL, which were identified as Pseudomonas, grew in 2-chloroacrylate (2-CAA) and 2-chloropropionate (2-CPA) media to produce 2-halo acid dehalogenases. Pseudomonas sp. YL showed the highest 2-CPA dehalogenase activity among the three strains, and inducibly produced two kinds of 2-halo acid dehalogenases in a medium containing either 2-CAA or 2-CPA. The 2-CAA-induced enzyme catalyzed dehalogenation of D- and L-2-CPA to produce L- and D-lactate, respectively. Its pH optimum of the dehalogenation and activity staining indicated that a single enzyme catalyzes the dehalogenation of both enantiomers of 2-CPA. The 2-CPA-induced enzyme catalyzed production of D-lactate from L-2-CPA, but did not act on D-2-CPA. The two dehalogenases are different from each other in mobility upon polyacrylamide gel electrophoresis and sensitivity towards thiol reagents.

View full abstract

Streptococcus mutans induces both H₂O₂-forming and H₂O-forming NADH oxidases in the presence of O₂ [M. Higuchi, J. Gen. Microbiol., 130, 1819-1826 (1984)]. In this paper, a nox-1 gene encoding H₂O₂-forming NADH oxidase (NOX-1) from Streptococcus mutans was cloned, and the nucleotides sequenced. The structural gene of nox-1 consisted of 1530 base pairs, which encode a polypeptide consisting of 510 amino acids with a predicted molecular mass of 55, 196 Da. The deduced N-terminal amino acid sequence was consistent with that previously found for the purified NOX-1 protein. The nox-1 gene was expressed in Escherichia coli using its own promoter. Alignment of the amino acid sequence of NOX-1 with those of NADH oxidases from other microorganisms showed identities of 55.6%, 20.8%, 20.3%, and 7.3% for those of Amphibacillus xylanus Ep01, Streptococcusfaecalis 10C1, Thermoanaerobium brockii Rt8. G4, and Thermus thermophilus HB8, respectively.

View full abstract

This work was designed to examine culture variables influencing the production of uridine by the Bacillus subtilis mutant No. 556. A combination of corn steep liquor (4%) + corn gluten meal (0.7%) + urea (2%) was the most suitable nitrogen source for the production of uridine ; a high level of ammonium ions in the medium brought about abundant cell growth in the early stage of incubation and was detrimental to the fermentation. The addition of calcium salts, such as C_aCO₃ and C_aCl₂, was also essential for obtaining high yields of uridine. Among the carbon sources tested, glucose gave the best result. The optimum temperature and oxygen transfer rate for producing uridine were around 38°C and 20 x 10^-7 molO₂ / min / ml, respectively. Under optimum conditions in a 6000-liter fermentor, strain No. 556 produced 65mg/ml of uridine from 18% glucose.

View full abstract

We investigated the effects of various dietary fibers or their likenesses on the apparent fat digestibility by rats fed on a high-fat diet. Each of 23 diffrent fibers was added at 5% (w/w) to a purified diet containing 20% (w/w) corn oil. The rats were fed these diets for 2 weeks, and the feces were collected from each animal during the last 3 days. When compared with cellulose (control), 10 of the tested fibers significantly increased the fecal lipid excretion. Among these fibers, chitosan markedly increased the fecal lipid excretion and reduced the apparent fat digestibility to about a half relative to the control. The apparent protein digestibility was not greatly affected by chitosan. The fatty acid composition of the fecal lipids closely reflected that of the dietary fat. These results suggest that chitosan has potency for interfering with fat digestion and absorption in the intestinal tract, and for facilitating the excretion of dietary fat into the feces.

View full abstract

Rats were fed for 2 weeks on five different high-fat diets containing cellulose (control) or chitosan with and without organic acids (i. e., ascorbic, lactic and citric acids) as the dietary fiber. The apparent fat digestibility in the chitosan-receiving groups was significantly lower than in the control group. The addition of ascorbic acid to chitosan caused a larger increase in the fecal fat excretion than otherwise, without considerably affecting the apparent protein digestibility. The effect is thought to have occurred because chitosan was so well dissolved and mixed with fat by the action of ascorbic acid that it encapsulated fine fat droplets on its gel after contacting the pancreatic juice in the samll intestine. The intake of a mixture of chitosan and ascorbic acid is suitable for reducing excess fat.

View full abstract

A DNA fragment containing the gene for a thermostable hydantoinase was cloned from a thermophile, Bacillus stearothermophilus NS1122A in Escherichia coli. Nucleotide sequencing showed that the DNA fragment contains one open reading frame, which is predicted to encode a peptide of 471 amino acids, with a calculated molecular weight of 51, 724. When the hydantoinase gene was under the control of both the lpp promoter and the lac promoter-operator, and its expression was induced by isopropyl-1-thio-β-D-galactopyranoside, it was overexpressed in E. coli leading to the formation of an insoluble aggregate. The enzyme was purified to homogeneity from the insoluble aggregate. The molecular mass of the purified active enzyme was approximately 200 kDa by gel filtration. Although the monomer had no activity, the activity was restored by incubation with Mn²⁺ or Co²⁺ at pH 8. 1. These findings suggested that the hydantoinase is a metalloenzyme and the oligomeric structure is required for activity. The oligomeric structure is suggested to contribute to thermostability.

View full abstract

To clarify the absolute configurations of antafumicins A and B, which are antifungal substances from Aspergillus niger NH-401, the total synthesis of (2S)-antafumicins was accomplished by starting from (S)-malic acid in 12 steps. Based upon the physicochemical data of the synthetic samples, the absolute configurations of naturally occurring antafumicins A and B were determined as (2R, 4S)- and (2R, 4R)-4-(3-acetyl-2, 6-dihydroxyphenyl)-2-methoxy-4-butanolide, respectively.

View full abstract

A simple measuring system for the frequency of recombinant clones generated by R-protein mediated recombination was constructed. By applying the system, the frequencies of recombinant clones due to site-specific recombination between two loci on one or two different nonhomologous chromosome(s) were monitored during cultivation. Subsequently, it was found that the frequencies increase nearly in proportion to the culture period and are different among the individual two selected loci. Thus, the system is thought to be a candidate for analyzing physicochemical situations, such as folding and localization, of chromosomes in the nucleus.

View full abstract

Panax ginseng extract contains nerve growth factor-like immunoreactive substance (Panax ginseng NGF) that is cross-reactive with anti-mouse NGF IgG. Panax ginseng NGF-like substance and mouse NGF are almost equivalent with respect to their neurite outgrowth-stimulating activity and are immunologically indistinguishable. The molecular weight of Panax ginseng NGF-like substance estimated by the gel filtration method is identical with that of mouse NGF. The isoelectric point of Panax ginseng NGF is about 9.1, like that of mouse NGF. These results suggest that the root of Panax ginseng contains a biologically active NGF-like immunoreactive substance.

View full abstract

The substrate specificity of β-fructofuranosidase from Bifidobacterium adolescentis G1 for fructooligosaccharides [1^F (1-β-D-fructofuranosyl)_n-1 sucrose, GF_n, (n=3-8)] andinulooligosaccharides [1^F (1-β-D-fructofuranosyl)_n-1 fructose, F_n, (n=2-7)] were investigated. The K_m (mM) and k₀ (s^-1) values were : GF₃, 1.1 and 155 ; GF₄, 1.6 and 154 ; GF₅, 3.2 and 252 ; GF₆, 4.2 and 186 ; GF₇, 7.1 and 260 ; GF₈, 7.0 and 180 ; F₂, 4.9 and 213 ; F₃, 1.5 and 423 ; F₄, 2.4 and 311 ; F₅, 1.9 and 458 ; F₆, 8.7 and 369 ; F₇, 8.1 and 323, respectively. The enzyme preferred oligosaccharides to inulin and sucrose as the substrate. The enzyme hardly catalyzed transfructosylation from GF₂. The dependence of rate parameters on the degree of polymerization differed from that of Penicillium trzebinskii exo-inulinase. The subsite atfinities in the active site were 0.93, 4.33, 1.15, -0.48, 0.38, -1.07, and -0.04 kcal/mol for subsites 1, 2, 3, 4, 5, 6, and 7, respectively.

View full abstract

The effects on serum cholesterol level were examined in rats fed on various xenobiotics. The hypercholesterolemia induced by polychlorinated biphenyls (PCB) was characterized in rats, from which Iipoproteins were isolated by ultracentrifugation. A dietary addition of 0.03% PCB, 0. 3% chloretone, 0.1% aminopyrine, or 0.2% 2, 6-di-tert-butyl-β-cresol (BHT) resulted in a significant increase in serum cholesterol, although the chemical structure of each of these xenobiotics was different. The serum cholesterol level was markedly increased by one month of PCB feeding, the effect of PCB on the serum phospholipid level beingsimilar. The serum triglyceride level transiently increased within 7days of feeding with PCB diet. PCB feeding resulted in the elevation of all lipoproteins, including VLDL, LDL, HDL₁, and HDL₂, a marked increase being observed in HDL₁. Both HDL₁ and HDL₂ isolated from PCB-treated rats contained more apolipoprotein A-I (apo A-I) and less apo E than normal. VLDL isolated from PCB-treated rats had more cholesterol and apo E, but less apo C than that of the control animals. These data demonstrate that PCB feeding resulted in increased VLDL rich in cholesterol and apo E, and increased HDL rich in apo A-I. This experimentally induced hypercholesterolemia resulting in apo A-I-rich HDL would be a useful model for investigating the metabolism of apo-A-I and HDL.

View full abstract

The gene for monoamine oxidase (MAO) was cloned from an Escherichia coli genomic library and MAO was overproduced in the periplasmic space. The enzyme was purified to homogeneity by preparation of a periplasmic fraction, followed by ammonium sulfate fractionation and DEAE-cellulose column chromatography. Crystals were obtained by the hanging drop method using sodium citrate as a precipitant. The enzyme was found to be a dimer of identical subunits with a molecular weight of 80, 000, and showed the highest activity at pH 7.5 and 45°C. The enzyme was inhibited by a MAO specific inhibitor, hydroxylamine, hydrazine, phenelzine, isoniazid, and tranycypromine. The enzyme oxidized tyramine, phenethylamine, and tryptamine at higher rates, but not oxidized diamine and polyamines such as putrecine and spermine. The antibody against E. coli MAO cross-reacted with purified MAO A from Klebsiella aerogenes.

View full abstract

Dextrin dextranase (EC 2.4. 1.2 ; DDase), which is produced by Acetobacter capsulatus ATCC 11894, acted on a mixture of stevioside and starch hydrolysate with isoamylase, so that the enzyme was found to convert stevioside to predominantly mono-glucosyl-stevioside (SG1) and di-glucosyl-stevioside (SG2), and little of the stevioside initially added remained. SG1 was separated into two compounds (SG1a and SG1b) by reversed-phase high-pressure liquid chromatography. The structures of SG1a, SG1b, and SG2 were analyzed and concluded to be 13-O-(6-α-glucosyl-2-β-glucosyl-β-glucosyl)-19-O-β-glucosyl-steviol, 13-O- [(6-α-glucosyl)(2-β-glucosyl)-β-glucosyl]-19-O-P-glucosyl-steviol, and 13-O-[(6-α-glucosyl)(6-α-glucosyl-2-P-glucosyl)-β-glucosyl]-19-O-β-glucosyl-steviol, respectively. During the reaction for production of glycosylsteviosides, DDase catalyzes transglucosylations from glucosyl donor compounds to stevioside to be SG1a and SG1b, and to SG1b to be SG2 rapidly forming α-1, 6 glucosidic linkages. However transglucosylation to SG1a to be SG2 rarely occurred, and the conversions among stevioside and these glycosyl-steviosides were catalyzed by the action of DDase to transfer α-1, 6 linked glucosyl residues, forming α-1, 6 linkages

View full abstract

The inhibition by oligo- and poly-L-glutamic acids of calcium phosphate insolubilization was investigated with special attention to their molecular weights. The inhibitory activity was evaluated by the induction time from base titration experiments. The inhibitory activity of oligo-L-glutamic acids increased as their molecular weights (residue numbers) increased, and that of oligo-L-glutamic acid (20 mer) was almost the same as that of poly-L-glutamic acids (70 mer or 340 mer). Calcium binding properties of oligo-L-glutamic acids were measured by using a calcium-ion-selective electrode. The apparent amount of calcium bound to oligo-L-glutamic acids tended to increase as their molecular weights increased. The calcium-binding isotherm of oligo-L-glutamic acid (20 mer) was almost identical with that of poly-L-glutamic acid (70 mer). Moreover, the effect of poly-L-glutamic acid on the crystalline forms of insolubilized calcium phosphates was investigated by X-ray diffraction analysis, with the result that CaHPO₄·2H₂O (DCPD), a precursor of hydroxyapatite, was not observed.

View full abstract

The scope and limitation of a bottom-fermentation yeast (Saccharomyces cerevisiae IFO 0565) toward the reduction of 3-oxoalkanoic esters were examined. The substrate specificity of this microorganism for various kinds of 3-oxoalkanoic esters was studied. This microorganism was distinct from converntional bakers' yeast in terms of its selectivity in the reduction and its high expression of a hydrolytic enzyme. 3-Oxoalkanoic ester with an aromatic substituent, a halogen substituted 3-oxoalkanoic ester, an aliphatic longer-chain 3-oxoalkanoic ester and its α, α-difluoro analog were also accepted by this microorganism. The products are useful intermediates in the synthesis of physiologically active compounds.

View full abstract

(2S, 2'R, 3R, 4E, 8E)-N-2'-Hydroxyoctadecanoyl-1-O-(β-D-glucopyranosyl)-9-methyl-4, 8-sphingadienine (Pen III), a cerebroside isolated from Penicillium funiculosum A-1 as the fruiting inducer against Basidiomycete Schizophyllum commune, was synthesized by starting from D-glucose, L-serine, homoprenyl acetate and stearic acid.

View full abstract

A spectrophotometric assay for aminopeptidase in biological fluids has been developed using a thermostable alanine dehydrogenase (AlaDH, EC 1.4.1.1) from Bacillus stearothermophilus. L-Alanine produced by the aminopeptidase with L-alanine amide or L-leucyl-L-alanine as the substrate, is oxidatively deaminated to pyruvate in the presence of NAD⁺ by the action of AlaDH. Aminopeptidase activity was continuously monitored by measuring the absorbance at 340nm corresponding to NADH production. The measured aminopeptidase activity was found to be linear up to 700-800 units / liter at 37°C. The reagents were stable in solution for at least 4 weeks at 10°C. This method was applicable to the assay of serum aminopeptidase. The assays had a high degree of precision even at low aminopeptidase activity and correlated well with the conventional assay methods.

View full abstract

he absolute configuration of dehydrodiconiferyl alcohol was elucidated by chemical degradation. (+)-Dehydrodiconiferyl alcohol was prepared by optically resolving the recemate with HPLC and degraded to methylsuccinic acid. This methylsuccinic acid was the (R)-(+)-enantiomer, the optical purity of which was confirmed by HPLC after suitable conversion. This result shows that the absolute configuration of C-2 of (+)-dehydrodiconiferyl alcohol was S. H-2 and H-3 of (+)-dehydrodiconiferyl alcohol are trans, so the absolute configuration of C-3 must be R. Thus, (+)-dehydrodiconiferyl alcohol was 2S and 3R, and the (-)-enantiomer was 2R and 3S. The absolute configuration of natural glucosides of dehydrodiconiferyl alcohol was also elucidated.

View full abstract

Two chitin synthase genes (chs1 and chs2) were isolated from Rhizopus oligosporus by plaque hybridization probed with the chitin synthase 2 gene of Saccharomyces cerevisiae. From their deduced amino acid sequences, they were both class II chitin synthases according to the classification proposed by Bowen et al. ¹⁾ The expression of these genes was controlled differently in each stage of differentiation. It was suggested that the gene products of chs1 and chs2 function mainly in the hyphae growing stage but not in the late stage of spore formation. When each of these genes was expressed in S. cerevisiae, elevation of chitin synthase activity was observed in both cases.

View full abstract

An E. coli expression clone coding for human proinsulin, which was fused to NH₂-terminal β-galactosidase, was engineered for the separation from host proteins by introducing peptide devices, and for the sequential removal of the fused polypeptide by cyanogen bromide in front of the NH₂ terminalresidue (methionine) of the human proinsulin gene. Short synthetic genes encoding oligopeptide residues including (Glu)_n, (His)_n, (Trp)_n, and (Ser)_n(n=10 or 11), which have certain characteristic physical properties such as metal-affinity, polarity, hydrophobicity, and hydrophilicity, respectively, were inserted at the junction region of the gene fusion. Interestingly, it was found that among the oligopeptides, the oligohistidine residue as an affinity-tag has greatly facilitated the procedures for FPI purification, particularly in the manner of selective metal-affinity precipitation. The chelating peptide covering the NH₂-terminal beta-galactosidase portion could then be removed simply after purification to generate a protein with the natural amino acid sequence of proinsulin by cyanogen bromide.

View full abstract

Erbstatin, a tyrosine kinase inhibitor, inhibited p59^fyn- and p56^1ck-kinases in vitro with IC₅₀s of 0.21 and 0.18μg/ml, respectively. Inositol phosphates formation, enhanced by anti-CD3, wasalso inhibited by erbstatin. Moreover, erbstatin treatment prevented anti-CD3-induced interleukin 2 (IL-2) production, but phorbol ester-induced IL-2 production was not affected by erbstatin.

View full abstract

The cytotoxicity of hydrogen peroxide (H₂O₂) toward Chinese hamster V79 cells was assessed with a colony-formation assay. Decylubiquinone, a coenzyme Q homolog, suppressed H₂O₂-induced cytotoxicity. Almost all the incorporated decylubiquinone was detected as decylubiquinol after incubating the cells for 4h. These results suggest that decylubiquinol reduced in the cells acted as an antioxidant against H₂O₂-induced oxidative cell injury.

View full abstract

Basic 7S globulin in soybean seeds is a cysteine-rich glycoprotein with insulin-binding activity, and has 27- and 16-kDa subunits linked by disulfide bonds. After N-glycanase treatment, the 27-kDa subunit did not react with insulin, but the 16-kDa subunit still did. Soybean seeds release much basic 7S globulin when immersed in hot water. Protein kinase activity was detected in the proteins released. The protein kinase activity of the globulin was measured by the phosphorylation method in a gel. The activity was conferred by the 16-kDa subunit. The 16-kDa subunit and the sugar chain of the 27-kDa subunit could bind with insulin, whereas protein kinase activity seemed to be associated with the 16-kDa subunit.

View full abstract

Xyloglucan which was isolated from the walls of suspension-cultured poplar cells showed an average mol wt of 45, 000 ; the extracellular xyloglucan is 25, 000 mol wt. Compositional analysis of the oligosaccharide units with Streptomyces endo-1, 4-β-glucanase indicated that the polysaccharides were mainly constructed of four kinds of oligosaccharide repeating units, XLFG, XXFG, XXLG, and XXXG. Poplar xyloglucans contain these four units distributed at random, but primarily at the same ratio among their molecules.

View full abstract

The cytokinin activity of isoamyl, benzyl and phenyl derivatives of a series of 4-aminopyridopyrimidines and 4-aminoquinazoline was tested on tobacco callus. Among the derivatives, 2-methyl-4-anilinopyrido [3, 4-d] pyrimidine exhibited moderate activity almost comparable to that of kinetin. The position of the nitrogen atom in the pyridine moiety of the pyridopyrimidine ring was critical for this activity.

View full abstract

The antioxidative activity of egg yolk lipoproteins and apoproteins toward linoleic acid (LA) in an emulsion was studied. High-density lipoprotein (HDL) was more effective than low-density lipoprotein (LDL), and apo-HDL was more effective than apo-LDL when each was used after being dissolved in NaOH. The lipid moiety of HDL also had an antioxidative effect on LA directly or on LA in an emulsion, and might have enhanced the antioxidative activity of apo-HDL.

View full abstract

We found that interferon-α produced by human leukocytes contained four subtypes that were glycosylated by N-linkages according to the results of lectin blot analysis. These glycosylated subtypes were type H or type ω and thei sugar moieties had no relation to their biological activities.

View full abstract

Phospholipase D (EC 3.1.4.4) from Streptomyces sp. Catalyzed the transfer reaction of the dipalmitoylphosphatidyl residue from 1, 2-dipalmitoyl-3-sn-phosphatidylcholine (DPPC) to both arbutin and kojic acid in a biphasic system, to afford 1, 2-dipalmitoyl-3-sn-phosphatidylarbutin (DPP-arbutin) and 1, 2-dipalmitoyl-3-sn-phosphatidylkojic acid (DPP-kojic acid), respectively. The transfer reaction of DPPC was accompanied by hydrolysis to phosphatidic acid, and the ratio was significantly affected with organic solvents used in the biphasic system. DPP-arbutin and DPP-kojic acid showed almost the same inhibitory activity to tyrosinase (EC 1.14.18.1) from mushroom as arbutin and kojic acid, respectively.

View full abstract

Irradiation of lycopene in an atmosphere of oxygen and in the presence of a sensitizer (methylene blue) led to the formation of 2-methyl-2-hepten-6-one and apo-6'-lycopenal. A reaction pathway via a dioxetane intermediate is proposed.

View full abstract

The use of N-acetylneuraminic acid, sialyl-lactose, and glycomacropeptide by bifidobacteria and lactobacilli, and their growthpromoting effects on B. longum, B. breve, B. bifidum, and B. infantis were investigated. The data presented here suggest that fortification with N-acetylneuraminic acid-containing substances of infant formula may provide formula-fed infants with a function that human milk possesses.

View full abstract

Tryptamine was identified as a stress compound in UV-irradiated barley leaves. Its induction was also observed to occur upon the inoculation of plant pathogenic fungi. In the interaction between barley and powdery mildew fungus, the induction was related to the degree of resistance and spore germination was inhibited by tryptamine in vitro, suggesting that it acts as a defense substance.

View full abstract

A novel method for screening the melanin biosynthesis inhibitor, using the larval haemolymph of the silkworm, Bombyx mori, was developed, and this screening method applied to cultured broths of fungi. Trichoviridin was found to inhibit not only the melanin formation in larval haemolymph, but also mushroom tyrosinase.

View full abstract

Whey protein and egg white protein were conjugated with glucose 6-phosphate (G6P) through the Maillard reaction by incubation at 50°C and 65% relative humidity for 1 day. The brown color of whey protein-G6P conjugate developed faster than that of egg white protein-G6P conjugate. No precipitate of calcium phosphate was formd in the presence of 2% protein-G6P conjugate in the solution containing 30 mM calcium, 22 mM phosphate, and 10 mM citrate at pH 6.7.

View full abstract

It is known that the abnormal metabolism of tryptophan is caused by feeding a diet containing 6-aminonicotinamide. However, when rats were fed with a diet containing a large amount of nicotinic acid or nicotinamide, no aggravation of tryptophan metabolism was observed. We found that nicotinamide and nicotinic acid completely repaired the disturbance of tryptophan metabolism caused by 6-aminonicotinamide.

View full abstract

Chromosaponin I (CSI), the naturally occurring form of soya-saponin I, inhibited the oxidation of soybean phosphatidylcholine liposomal membranes induced by a water-soluble radical initiator, 2, 2'-azobis(2-amidinopropane) dihydrochloride. The antioxidative activity of CSI was similar to that of urate. Soyasaponin I, a degraded product of CSI, which has previously been thought to be an antioxidant, exerted no antioxidative activity.

View full abstract

A form II ribulose-1, 5-bisphosphate carboxylase / oxygenase (RubisCO) gene was isolated and sequenced from a marine hydrogen-oxidizing bacterium, Hydrogenovibrio marinus strain MH-110. To our knowledge, this is the first report for the sequence of a form II (L₂-form) RubisCO gene derived from a chemolithoautotrophic bacterium. The form II RubisCO gene coded for 463 amino acid residues (1389 bp, M_r = 50, 655). The deduced amino acid sequence had high homology with those of the L₂-form RubisCO of Rhodospirillum rubrum (63%) and the L₄-form RubisCO from Rhodobacter sphaeroides (62%), but low similarity to other L₈S₈ form RubisCOs (under 40%)

View full abstract

The activity of biotin synthase, responsible for biotin synthesis from dethiobiotin, was demonstrated in a completely defined reaction mixture with cell-free extracts of a Bacillus sphaericus bioB transformant. Among the sulfur compounds tested, only S-adenosyl-L-methionine was active, while L-methionine and L-cysteine had no significant effect. Protein concentrations higher than 15mg/ml in the reaction mixture were needed to detect biotin synthase activity. When dialyzed cell-free extracts were used for the reaction, NADH, NADPH, or FAD among the well-known cofactors tested enhanced the activity, and Fe²⁺, Mn²⁺, and Ca²⁺ among the metal ions tested also had some effects.

View full abstract

New glycosidic aroma precursors (1 and 2) of the main volatile constituents, trans- and cis-linalool 3, 6-oxides (linalool oxides I and II), were isolated from oolong tea leaves (Camellia sinensis var. sinensis cv. Maoxie). The isolation was guided by an enzymatic hydrolysis with acetone powder prepared from fresh tea leaves (cv. Yabukita) followed by GC or GC-MS analyses. Chromatographic purification of hot water extracts of the tea leaves on active charcoal, Amberlite XAD-2, and Sephadex LH-20 columns as well as HPLC gave two new glycosides, trans- and cis-linalool 3, 6-oxide 6-O-β-D-xylopyranosyl-β-D-glucopyranosides (1 and 2).

View full abstract

The esterase catalyzes the stereoselective hydrolysis of methyl DL-β-acetylthioisobutyrate (DL-ester) to give D-β-acetylthioisobutyric acid (DAT). To use the enzyme for DAT production, the esterasegene of Pseudomonas putida was cloned and expressed in E. coli. The recombinant E. coli containing the esterase gene produced a large amount of the enzyme in an active form. This strain could be used for the asymmetric synthesis of DAT.

View full abstract

The production of hydroxyproline by microorganisms was studied to discover the chiral source of the side chain moiety of carbapenem. cis-4-Hydroxy-(L)-proline produced by Helicocerus oryzae and A crocylindrium oryzae was purified from culture broths. The ¹NMR spectra and [α]_D of purified samples were in agreement with those of standard samples. Our finding provided the first evidence for fermentative production of cis-4-hydroxy-(L)-proline. The addition of (L)-proline to the culture broth of H. oryzae increased the production of cis-4-hydroxy-(L)-proline.

View full abstract