The induction of thermotolerance by split-dose hyperthermia was analyzed in the radioresistant bacterium Deinococcus radiodurans wild type strain R₁. Immediately after a 30 min exposure to 52°C, the cells were maintained at 30°C for various intervals up to 6hr in a medium and then exposed again to 52°C for various periods, that is, "split-dose hyperthermia." Induced thermotolerance was maximum at the 4 hr interval. However, when chloramphenicol was added to the medium or the cells were incubated in a phosphate buffer, the thermotolerance induction decreased to the level of the survival curve without initial 52°C treatment. Also, when the cells previously heated were maintained at 42°C for 6 hr instead of 30°C, this tolerance induction was gently caused.
Therefore, these results suggest that proteins synthesized de novo during the interval incubation were involved either in the appearance of thermotolerance or in the recovery from the injury induced by 52°C heating for 30 min.

View full abstract

A biofertilizer, showing antagonistic activity against potato common scab in pot tests, was produced from swine feces with a newly isolated strain, CH-33, identified as Streptomyces albidoflavus. This strain characteristically grew on fresh swine feces at 20-35°C without sterilization or any additives, and produced an antibiotic substance against Streptomyces scabies, the common scab-pathogen, during composting. The addition of the biofertilizer at from 0.1 g to 1.6g total nitrogen (N) per 600 g humic volcanic ash soil in a pot did not inhibit the growth of Brassica rapa var. perviridis but increased it, even at the highest nitrogen content tested. Common scab was completely inhibited when the biofertilizer was added at 0.1 g to 1.6g as nitrogen (N) per 4kg of scab-infected soil in a pot. Thus a biofertilizer suppressing plant pathogenic microorganisms was developed.

View full abstract

The effect of melanoidin, prepared with a D-glucose and glycine system, on cholesterol metabolism in rats was examined. Male rats of the Wistar strain weighing ca. 140 g were fed a highcholesterol diet supplemented with nondialyzable melanoidin for three weeks. The dietary melanoidin suppressed elevation of the cholesterol level in both plasma and liver, while the cholesterol levels in feces and the contents of the caecum decreased unexpectedly. Neutral steroids other than cholesterol also decreased. However, chromatograhy of fecal steroids indicated that melanoidin markedly affected the intestinal metabolism of neutral steroids, including cholesterol. On the other hand, the fecal recovery of radioactivity from orally ingested 26 [27]-¹⁴C cholesterol was promoted by the added melanoidin, although the radioactive species were not identified. This suggested that the cholesterol level decreased due to the interference of melanoidin in the intestinal absorption of cholesterol.

View full abstract

A Pseudomonas strain capable of utilizing 1, 6- and 2, 6-naphthalenedisulfonic acid as sole sources of carbon was isolated from soil. Sulfonate was released from 1, 6- and 2, 6-naphthalenedisulfonic acid into the culture medium. When the cells were incubated with 1, 6- or 2, 6-naphthalenedisulfonic acid in the incubation medium containing m-sulfobenzoic acid as a metabolic inhibitor, 5-sulfosalicylic acid was accumulated in the medium. The bacterium degraded 5-sulfosalicylic acid and gentisic acid. These results suggest that the bacterium assimilates 1, 6- and 2, 6-naphthalenedisulfonic acid via 5-sulfosalicylic acid.

View full abstract

A novel enzymatic method for the production of L-carnitine (L-4-trimethylamino-3-hydroxybutyric acid) has been developed.
Microorganisms that asymmetrically produce L-carnitine from trans-crotonbetane(trans-4-trimethylamino-2-butenoic acid), which was easily supplied through the chemical dehydration of DL-carnitine with an acid, were screened from stock cultures in our laboratory. Of the 686 strains of microorganisms examined, i.e., 416 bacteria, 37 actinomycetes and 233 yeasts, strains belonging to the genera, Proteus and Escherichia, were found to be efficient producers of L-carnitine from trans-crotonbetaine. Through the screening, Proteus mirabilis AJ-2772 and AJ-12137, with potent abilities, were selected as representative strains. The trans-crotonbetaine-hydrating enzyme was inducibly and intracellularly produced on either the addition of trans-crotonbetainQ or L-carnitine to the medium under partially anaerobic conditions.
With intact cells of P. mirabilis AJ-2772, 40 g/l of L-carnitine chloride was produced from 62.5g/l of trans-crotonbetaine.

View full abstract

UV-induced structural chromosome aberrations were suppressed by post-treatment with tannic acid (m-galoyl gallic acid) in cultured Chinese hamster ovary cells. A significant decrease in the frequencies of chromosome aberrations in the presence of tannic acid was observed in cells treated with mitomycin C or methylmethanesulfonate but not with X-rays or bleomycin. The effects of tannic acid were dependent on the DNA pre-replicational G1 phase of the cell cycle. Furthermore, tannic acid enhanced the liquid-holding effect on UV-irradiated cells at G1 phase. On the other hand, the number of surviving UV-irradiated cells increased in the presence of tannic acid. These results suggest the possibility of promotion by tannic acid of an excision repair system of mammalian cells.

View full abstract

The screening of facultative anaerobes with melanoidin-decolorizing activity (MDA) was performed, and some strains were isolated from stored waste water of an alcohol fermentation involving molasses. One of them, strain W-NS, showed high and stable MDA, and was identical with Lactobacillus hilgardii.
The decolorization yield of this strain under the optimal conditions was only 28%, however, it was improved to 40% by immobilization of cells within Ca-alginate gel. Unlike Basidiomycetes and Ascomycetes, this strain decolorized smaller molecular weight fractions of melanoidin, quickly. Also, the MDA of this strain toward various synthetic melanoidin was quite different from that of Basidiomycetes.

View full abstract

The continuous decolorization of molasses waste water (MWW) by immobilized cells of Lactobacillus hilgardii W-NS was studied. The immobilized cells showed the maximal decolorization yield in the presence of 1% glucose with a medium pH of 5.0 at 45°C. On successive decolorization of MWW, with recycling of the immobilized cells, more than 90% of the maximal decolorization yield was maintained for one month when 0.05% peptone was added to MWW. In contrast, on continuous decolorization in a column type reactor, a sufficient decolorization yield could not be maintained, the decolorization yield dropping to half the maximal level during operation for 5 days. But the feeding of MWW adjusted to the pH of 7.3, compared with the maximal pH value (5.0), slowed down the decrease in the decolorization yield.

View full abstract

A synthesis of L-glutamic acid oligopeptide from L-glutamic acid-α, γ-diethyl ester hydrochloride is demonstrated by the use of papain and α-chymotrypsin. An examination of this polymerization reaction by turbidimetry, a micro-biuret assay and gel chromatography showed that effective synthesis was achieved at pH 7-8.5 in phosphate buffer at 25-35°C for both enzymes. It was revealed by gel chromatography that a higher molecular peptide fraction was accumulated during the reaction as a precipitate, which was determined to be peptides composed of 5 to 9 glutamic acid residues. When 100 mM of the substrate was incubated with 10 μM of papain, more than an 80% overall reaction yield (42% as a precipitate) was attained after 24 hr of the reaction.

View full abstract

Water-soluble phospholipase B was purified to homogeneity from Torulaspora delbrueckii cell washings. The washings were concentrated by ultrafiltration, and then a fraction with phospholipase B activity was precipitated with ammonium sulfate, and purified by sequential column chromatographies on Octyl-Sepharose CL-4B, DEAE-Sephacel, and Sepharose 6B. The molecular weight of the enzyme was estimated to be 170, 000 - 200, 000 by SDS-polyacrylamide gel electrophoresis and by gel filtration with a Sephadex G-200 column. The isoelectric point of the enzyme was 4.0. The purified enzyme had two pH optima at pH 2.5 and pH 7.5. The activity at acidic pH was greatly stimulated by the divalent metal ions tested, but the activity at alkaline pH was stimulated mainly by Ca₂₊ and Fe₂₊. The purified enzyme had both lysophospholipase activity and phospholipase B activity in a ratio of 37:1 at acidic pH and 73:1 at alkaline pH. The amino acid composition of the enzyme was characterized by high contents of Asp, Ser, Leu, and Gly.

View full abstract

Eight geometrical isomers of 10, 12, 14-hexadecatrienyl acetate, female sex pheromone candidates of the mulberry pyralid (Glyphodes pyloalis Walker), were synthesized by six routes. Each route consisted of reactions by which two of three double bonds were introduced stereo-specifically and another rather nonspecifically, giving mainly two geometrical isomers, which were analyzed by 2D-NMR after separating by reversed-phase HPLC or by a reaction with tetracyanoethylene. The signals of the olefinic protons and carbons of each geometrical isomer were assigned by COSY spectra and by C-H COSY spectra, respectively, in addition to the signals of the allylic protons and carbons, and its chemical structure was revealed by the values of their chemical shifts and coupling constants. Furthermore, a new empirical rule concerning the chemical shift changes of these carbons by converting the configuration of the conjugated triene system is suggested.

View full abstract

Accumulated phytoalexins in ultraviloet-irradiated detached rice leaves were detected by gas chromatography-mass spectrometry-selected ion monitoring (GC-MS-SIM) after pretreating with a BOND ELUT C₁₈ cartridge. An accumulation of oryzalexins A, B, C and D, momilactones A and B, and an unknown antifungal substance accompanied by the appearance of brown spots on the leaf surface was observed. Momilactone A was detected in abundance, and among the oryzalexins, oryzalexin D was a major substance. The maximum accumulation of these phytoalexins, except for oryzalxins C and D, was found 3 days after irradiation. However, the maximum accumulation of oryzalexin D was observed 2 days after irradiation and that of oryzalexin C was observed 4 days after it. The content of these diterpenoid phytoalexins in rice leaves was dependent on leaf aging, an accumulation of these phytoalexins in the uppermost leaves being much lower than that in the aged leaves (lower leaves), and brown spots scarcely ever appeared on the surface of the uppermost leaves.

View full abstract

The endo α-1, 4 polygalactosaminidase from Pseudomonas sp. 881 was purified from the culture filtrate by ethanol precipitation and sequential column chromatographies on CM-Sephadex C-25, Sephadex G-50 and Phenyl-Sepharose CL-4B. The purified enzyme was electrophoretically homogeneous and its molecular weight and isoelectric point were 31, 000 and 6.7, respectively. The optimum pH and temperature for hydrolysis of polygalactosamine were 5.0 and 55°C, respectively. The enzyme was stable up to 45°C for 15min and from pH 4.0 to 7.6 at 37°C for 1 hr.
The Km value was 0.05% α-1, 4 polygalactosamine and the V was 0.154/μmol reducing sugar (galactosamine)/min/μg protein. This polygalactosaminidase was inhibited by Sn²⁺, Fe²⁺, Fe³⁺, Hg²⁺, Cu²⁺ ions and SDS. The enzyme did not hydrolyze oligo galactosamines (n < tetramer) or N-acetyl-polygalactosamines. It acted only on oligo galactosamine (n > trimer) and polygalactosamine endogeneously so far tested.

View full abstract

Identification of the T-cell recognition sites in β-lactoglobulin (β-LG), the major allergen in bovine milk, was attempted with its enzymatic digests. The peptides from both the tryptic and chymotryptic digests of reduced carboxymethylated (RCM) β-LG were separated by gel-filtration and reverse-phase HPLC. Co-culturing lymph node cells from β-LG-primed BALB/c mice with each fraction, T-cell proliferation was examined by uptake of ³H-labelled-thymidine. Three tryptic fragments (the residues ²¹Ser-⁴⁰Arg, ⁴¹Val-⁶⁰Lys, and ¹⁰²Tyr-¹²⁴Arg) and three chymotryptic fragments (the residues ²¹Ser-³²Leu, ⁵⁹Gln-⁸²Phe, and ¹²³Val-¹⁴⁰Leu) could stimulate the proliferation of the T cells. These T-cell antigenic sites seem to be located on the surface regions of the β-LG molecule. They didn't agree completely with the T-cell epitopes of β-LG predicted by the amphipathic periodicity hypothesis.

View full abstract

A component of the microcrystalline cellulose (Avicel)-hydrolyzing enzyme produced by a fungal strain, Y-94, tentatively called Avicelase II, was purified by several kinds of column chromatography followed by chromatofocusing with PolybufFer anion exchanger (PBE 94), Bio-Gel A0.5 m gel filtration, electrophoresis on polyacrylamide gel and disc isoelectric focusing (IEF). The molecular weight of the enzyme was estimated to be 68, 000 by the SDS-polyacrylamide gel method and 56, 000 by Bio-Gel A0.5 m gel filtration, and its isoelectric point was found to be around 4.4 by the chromatofocusing and disc IEF methods. The optimum pH and temperature for Avicel hydrolysis were 5.3 and 62°C, respectively. The enzyme was stable between pH 4.1 and 6.0 at 4°C for 24 hr, and up to 61°C for 10min. Heavy metal ions such as Cu²⁺ and Hg²⁺ strongly inactivated the enzyme. A highly purified enzyme preparation showed hydrolyzing activities towards Avicel, xylan, carboxylmethyl cellulose (CMC) (activity ratio of 100:50:20, respectively) and acidswollen cellulose. By the PAS method, staining glycoprotein with Shiffs reagent and periodic acid, the enzyme seemed to be a glycoprotein containing some carbohydrate in its structure. Avicelase II tended to show a higher affinity towards cellooligosaccharides of high molecular weight. The enzyme hydrolyzed Avicel and acid-swollen cellulose, and produced trace amounts of cellotriose and significant quantities of glucose. In addition, the enzyme is able to act on the internal glycosidic bonds of CMC and xylan. Judging from the results, it appears that Avicelase II should be classified as a member of the specific endoglucanases rather than as an exoglucanase.

View full abstract

The effects of prolactin (P), hydrocortisone (F), and insulin (I) on the formation of lipidsaccharide intermediates concerned in the N-linked glycoprotein biosynthesis of lactating bovine mammary explants in organ culture were examined using radioactive mannose (Man). The response of F was fast (<20 hr), while that of P and I was delayed (> 72 hr). F at concentrations varied from 10^-3 to 10μg/ml decreased the incorporation of radioactivity from [¹⁴C]Man into the lipid-linked saccharides (C/M), lipid-linked oligosaccharides (C/M/W), and protein fractions by about 25-30%. On the other hand, at the range of 10^-4 to 5μg/ml, I had a stimulatory effect and P did not affect the N-glycosylation of mammary explants. The stimulatory effect of I, however, was negated by with F. Partial inhibition of F was shown by even 10ng/ml. The inclusion of F into the pre-incubation medium significantly inhibited the [¹⁴C]Man incorporation in the C/M/W and protein fractions rather than in the C/M fraction. The results suggest that F inhibits the formation of the lipid-linked saccharide intermediates for N-glycosylated glycoprotein in lactating bovine mammary explants.

View full abstract

The properties of endo-β-N-acetylglucosaminidase isolated from a Flavobacterium sp. were examined with native hen ovalbumin as a substrate for the first step in preparing a large amount of the deglycosylated glycoprotein with the native structure. This enzyme cleaved the oligosaccharide chain in native ovalbumin not only for the A₁ ovalbumin containing two phosphoryl residues per molecule, but also for the A₂ and A₃ ovalbumins, containing one and no phosphoryl residues, respectively. With polyacrylamide gel electrophoresis, glycosylated ovalbumin of the native form was separated from the deglycosylated variety, which was then transferred to a vinylidene fluoride membrane and stained with concanavalin A and peroxidase to confirm the deglycosylation. The apparent Km value was about 2.67mM at pH 6.0 when native ovalbumin was used as the substrate, the optimum pH for the enzyme reaction with native ovalbumin being around 5.0. This enzyme was not inhibited by its end product.

View full abstract

The denaturation of rabbit skeletal myosin caused by freezing and thawing was measured by the decrease in Ca²⁺-ATPase activity and by the increase of turbidity in 0.5 M KC1 solution. The changes in both activity and turbidity were great at concentrations of myosin less than 0.5mg/ml. Denaturation was accelerated at freezing temperatures close to 0°C. The myosin was protected from denaturation by non-ionic surfactants, Tween 20 at 2% completely preventing denaturation. For the same degree of prevention as 2% Tween 20, the concentration of glycerol needed was 20%. Glycerol and Tween 20 acted synergistically in preventing denaturation. The use of Tween 20 at 0.1% and glycerol at 1% resulted in 90% of the original Ca²⁺-ATPase activity after freezing and thawing.

View full abstract

A taxonomic study below the generic or at the specific level was performed as to the electrophoretic patterns of seven enzymes in twenty-six strains of Lipomyces and Waltomyces species. The seven enzymes were glucose-6-phosphate dehydrogenase, 6-phosphogluconate dehydrogenase, isocitrate dehydrogenase, malate dehydrogenase, hexokinase, phosphoglucomutase and fumarase. The five species, L. anomalus, L.kononenkoae, L. starkeyi, L. tetrasporus and W. lipofer constituted separate clusters from each other, the similarity values being all 0%. The two strains of L. kononenkoae were quite dissimilar in their electrophoretic enzyme patterns (s = 0%). The unidentified Lipomyces strain, CBS 8113, was very similar to the type strain of L. kononenkoae (s= 100%). The five strains of L. starkeyi were divided into two groups with a similarity value of 0%. The unidentified Lipomyces strain, CBS 8064, was a member of the second L. starkeyi group (s = 43%). The eleven strains of L. tetrasporus were divided into three separate groups with similarity values of less than 30%. The four strains of W. lipofer had identical electrophoretic enzyme patterns (s= 100%). The unidentified Lipomyces strain, CBS 8114, constituted a separate cluster, which was linked to the four strains of W. lipofer with a similarity value of only 14%. These data are discussed from the taxonomic point of view.

View full abstract

To analyze the T cell reactivity to ovomucoid (OM), one of the potent allergens in hen's eggs, the antigen-specific T cell proliferation assay was done. In the case of C3H/He and BALB/c mice, proliferative responses of OM-primed T cells to all domains (DI, DII, Dili) of OM were observed, but the response to DI was inferior to the other domains. The T cells from C57BL/6 responded to Dili and DII, but not at all to DI. These results suggest that each domain has common site(s) recognized by T cells, reflecting their homologies. Furthermore, the T cell reactivity to reduced carboxymethylated OM (RCM-OM), which has a conformation distinct from native OM, was examined. RCM-OM did not stimulate the T cells immunized with native OM, implying that OM-specific T cells recognize the conformational epitopes dependent on the tertiary structure of OM. This unique profile of the T cell reactivity to OM should be useful for analyzing the mechanism of the antigen processing and presentation of APC and the subsequent T cell antigen recognition.

View full abstract

One of the DNAs, Charon 10-gE1A-2, pSV2-gpt-gE1A, or Adl2 E1A, was microinjected separately into pronuclei of fertilized mouse eggs and fourteen transgenic mouse lines were obtained. The structures of integrated DNAs in these transgenic mice were analyzed by Southern blot hybridization. Head-to-head and tail-to-tail structures were found, although the injected molecules had a tendency to join each other in head-to-tail tandem forms when multiple copies were integrated. The analyses of the mice containing pSV2-gpt-gE1A showed that most of the restriction sites at the end of the injected molecules were restored at the tandem junctions. Furthermore, the integrated DNAs at the junctions to the host chromosomal DNA were deleted and/or rearranged.

View full abstract

Ricin D was denatured in frozen acidic solutions accompanied by the insolubilization. Kinetic studies indicated that the insolubilization of ricin D occurred during the incubation of the frozen solution at - 51°C--5°C, but not in the freezing process at - 80°C or the thawing process. Such an insolubilization of ricin D in the frozen system was accelerated by lowering the pH below pH 4.0, and the optimum temperature was found at - 29°C. The rate of the insolubilization of ricin D decreased on the addition of NaCl or on elevation of the temperature from - 29°C to - 5°C, or on increasing the protein concentration above 0.15%. The freeze-denaturation of ricin D was prevented by saccharides, and lactose, with a higher affinity for ricin D than galactose, suppressed the insolubilization of ricin D more effectively than galactose. A slight suppressive effect of lactose was found for insolubilization of a derivative of ricin D in which the saccharide-binding ability of the low affinity saccharide-binding site was destroyed by N-bromosuccinimide-oxidation. It is suggested that the denaturation of ricin D in the frozen acidic solution is principally due to the concentration of protons and solutes into liquid regions surrounded by ice crystals. Spectroscopic data also suggest that the protonation of the ionizable group(s) with pK_a = 4-5 may be closely associated with the denaturation of ricin D in the frozen state below pH 4.0.

View full abstract

The effects of urea concentration on the dithiothreitol-dependent gelation of conalbumin (ovotransferrin) were investigated. In the presence of a high concentration of urea, conalbumin gelation was induced at lower molar ratios of dithiothreitol to protein than in the absence of the denaturant. Sodium dodecyl sulfate-polyacrylamide gel electrophoresis revealed that in the presence, but not in the absence of urea, intermolecular disulfide bonds were mainly involved in the gelation process. Under a scanning electron microscope, the gel formed in the presence of urea was composed of more fibrous networks than that formed in the absence of the denaturant.

View full abstract

The great usefulness of the previously reported "stopped-flow time difference analysis" technique for improving enzymatic analyses based on dehydrogenase-catalyzed reactions is demonstrated by combining the technique with the conventional enzymatic method for pyruvate determination. Reagent economy was accomplished by using a micro-stopped-flow apparatus designed by one of the authors. The running speed and sensitivity were improved by about 100 and 7 times the figures for the conventional enzymatic method, respectively. Furthermore, as procedural blanks were unnecessary, a simpler method could be developed. The interference by several other keto acids was easily eliminated by analyzing the reaction curve, and the apparent first-order rate constant proved to be useful for qualitative analysis.

View full abstract

An inulinase was highly purified from the culture broth of Penicillium purpurogenum by chromatographies on DEAE-Sepharose CL-6B, Toyopearl HW-65, and Bio-Gel P-100. The enzyme was homogeneous by disc electrophoretic analysis. The molecular weight was 6.4 × 10⁴ by SDS-disc electrophoresis and gel filtration on Bio-Gel P-150. The isoelectric point was pH 3.6 by isoelectric focusing. The enzyme hydrolyzed inulin rapidly, but did not affect sucrose. By paper chromatography analysis, the major products from inulin were tri-, tetra-, penta-, and hexasaccharides. The substrate specificity of the enzyme on hydrolyses of fructo-oligosaccharides[1^F(1-β-D-fructofuranosyl)_nsucrose (n = 1 to 6 and n (average of polymerization degree) = 8)] were examined. The Km values and relative maximum velocities for the hydrolyses of inulin and fructooligosaccharides (GF_n, n = 2 to 7 and n = 9) were as follows: inulin, (DP = 35) 0.21 mM and 100; GF₉, 0.24mM and 86.5; GF₇, 0.33 mM and 132; GF₆, 0.85 mM and 71.2; GF₅, 3.8 HIM and 25.4; GF₄, 2.8mM and 28.8; GF₃, (nystose) 16mM and 0.8; GF₂ (1-kestose), 8.4mM and 0.2. The molecular activities for the hydrolyses of fructo-oligosaccharides (GF_n, n = 2 to 6) were increased depending on the degree of polymerization of fructosyl residues, and were nearly constant if the polymerization degree was over seven. These results strongly suggested that the endo-type inulinase from Penicillium purpurogenum had a subsite structure consisting of at least seven subsites.

View full abstract

A monoclonal antibody (MAb 1C3) to bovine β-casein was prepared, and a peptide (from 184 to 202 residues of β-casein) that could bind this monoclonal antibody was separated from a tryptic digest of β-casein. Other β-casein fragments, the peptide from 193 to 202 and the one from 1 to 189 residues, were also prepared for comparison. MAb 1C3 belongs to the IgG₁ subclass with a K chain and its pi was 5.9. The binding affinity of β-casein and β-casein fragments to the monoclonal antibody was investigated by a competitive radioimmunoassay, and the order of affinity was β-casein peptide from 193 to 202 > β-casein > β-casein peptide from 184 to 202 > β-casein peptide from 1 to 189. On the basis of the results, the mechanism for the interaction between MAb 1C3 and β-casein fragments is discussed.

View full abstract

Pectic substances were extracted from the vegetables with oxalate buffer of pH 4.25 and, after saponification, fractionated into two components, weakly acidic pectic polysaccharide (WAP) and pectic acid, by DEAE-cellulose and Sephadex G-100 chromatographies. The galacturonic acid content (17.3-25.8%) of WAPs was much lower than that of pectic acids, though the neutral sugar compositions of both pectic substances were almost the same. The arabinose-galactose side chains were found to be very long or highly branched in WAPs compared with those in pectic acids.
All the WAPs were appreciably hydrolyzed by exo- and endopolygalacturonases. The limiteddegradation products (the residual polysaccharides; i.e., the rhamnogalacturonan segments) obtained by endopolygalacturonase from both WAPs and pectic acids showed a similar behavior on Sephadex G-100 and Sepharose CL-4B gel filtrations; each of the rhamnogalacturonan segments was eluted in the void volume of the Sephadex G-100 column. From these results, we concluded that WAPs are probably an inherent pectic component of the cell walls of the vegetables.

View full abstract

We searched for a new aggregation factor, and found one we named 3315-AF in the culture filtrate of Streptomyces sp. strain No. A-3315. 3315-AF was purified by active carbon treatment, ethanol precipitation, gel filtration on Sepharose 2B, ether extraction, silica gel chromatography and gel filtration on Sephadex LH-20. 3315-AF was found to be a triglyceride which consists of myristic acid, pentadecanoic acid, and palmitic acid. The aggregation activity of 3315-AF was maximum around pH 8.0 at 30°C and the activity increased by addition of metallic ions such as calcium and cobalt. Hyaluronic acid, ovalbumin, BSA, and casein inhibited the aggregation activity. 3315-AF aggregated Proteus vulgaris and HeLa cells as well as Serratia marcescens and weakly aggregated Saccharomyces cerevisiae, Candida albicans, C. neoformans, and Leukemia P388, but it was inert to human erythrocytes and Sarcoma 180.

View full abstract

1. An aromatic amine-dependent NAD(P)H-oxidizing enzyme was purified from tomato fruit to homogeneity as judged by SDS-polyacrylamide gel electrophoresis. NAD(P)H was oxidized under aerobic conditions in the presence of aromatic amines such as N, N-dimethylaniline, N-methylaniline, and aniline. The amines were essential for the reaction, but were not metabolized.
2. The enzyme had a monomer molecular weight of 48, 000. On the basis of its spectral characteristics and the inhibition by KCN, NaN₃, and catalase, the enzyme was identified as a protoheme peroxidase.

View full abstract

In our previous papers, ¹⁾ syntheses of the derivatives of nicotine (1) have been investigated. The pyrolysis of 1 was one of the methods to prepare nicotine analogues which possessed unsaturation in the pyrrolidine ring. Nicotyrine (2) and myosmine (3) were reported to be prepared via pyrolysis of 1 over quartz sand at high temperature ( ?? 570°C).²⁾ However, their yields were very low, and several by-products were detected in the reaction mixture.
In this paper, we report the selective preparation of 2 by catalytic dehydrogenation of 1 over hydrous zirconium oxide and its analogues.

View full abstract

D-Gluconate dehydrogenase isolated from Pseudomonas fluorescens was immobilized on the surfaces of carbon and gold electrodes by irreversible adsorption. The electrodes with the adsorbed enzyme produced anodic currents in solutions containing D-gluconate. The currents were attributable to the electro-enzymic oxidation (direct bioelectrocatalytic oxidation) of D-gluconate; the electrochemical system required no external redox molecules serving as mediators of electron transfer between the electrode and the adsorbed enzyme. A model of the direct bioelectrocatalysis at the enzyme-modified electrodes is presented.

View full abstract

A newly found methanol-using bacterium, Mycobacterium gastri MB 19, is a facultative methylotroph which assimilates methanol via the ribulose monophosphate pathway. 3-Hexulose phosphate synthase was purified from the organism and characterized. This enzyme was found to use glycolaldehyde (Km =4.3 mM) and methylglyoxal (Km=5.7 mM) as well as formaldehyde (Km=1.4 mM) in the presence of D-ribulose 5-phosphate as an acceptor. The product of the condensation of glycolaldehyde with D-ribulose 5-phosphate was isolated by ionexchange chromatography. The dephosphorylated product was tentatively identified as a heptulose with the molecular formula C₇H₁₄O₇ from its spectrophotometric properties and GC-MS results.

View full abstract