Humicola insolens YH-8, a thermophilic fungus isolated from manure and compost heaps, produced a significant amount of thermostable cellulases in cultures on wheat bran medium (50°C, 4 days). The mold bran extract hydrolyzed Avicel, CMC and newsprint at 90%, 45% and 35%, respectively, to glucose. Then, Avicelase and CMCase were purified from the culture extract by adsorption onto Avicel, heat and acid treatment and consecutive column chromatographies to a homogeneous state on polyacrylamide gel disc electrophoresis. The purified cellulases, especially CMCase, was found highly thermostable. The optimal temperature of both enzymes was 50°C. Avicelase was stable after heating at 65°C for 5 min and CMCase retained 45% of the original activity after heating at 95°C for 5min.

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The thermophilic fungus, Humicola insolens YH-8 exhibited high β-glucosidase activity when grown in solid wheat bran medium. The β-glucosidase was purified from the culture extract by consecutive column chromatographies and found to be homogeneous on polyacrylamide gel disc electrophoresis. The molecular weight was estimated to be 250, 000 by SDS-gel electrophoresis, and the isoelectric point was at pH 4.23. The enzyme had an optimum pH of 5.0, an optimum temperature of 50°C, and showed significant resistance to urea, dimethyl sulfoxide and ethyl alcohol.

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Two types of reductive intermediates, linear and tricyclic forms, isolated from browning mixtures of triose reductone (TR) with guanine and its derivatives showed evident mutagenicity on Salmonella typhimurium TA 100 without S-9 mixture. The linear intermediates, N²-(3-oxo-2-hydroxypropenyl) compounds of guanine, guanosine, 2' (3')-guanylic acid and 5'-guanylic acid were more effective than the tricyclic one, 1, N²-(2-hydroxypropenylidene)guanine, though they were far less active than 4-nitroquinoline-N-oxide. No acceleration in mutagenicity was observed with Cu²⁺ and other metal ions. The reaction mixtures of TR and nucleic acid bases were also mutagenic on TA 100. Intermediates of TR with guanine and its derivatives did not have a lethal effect in Recassays with Bacillus subtilis.

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The amounts of amino-acid neurotransmitters and GAD activity were determined in different brain regions of microencephalic rats that were offspring of mother rats injected with methyl-azoxymethanol (MAM)-acetate on the 15th day of pregnancy.
The contents of GABA, taurine, aspartic acid, glutamic acid, and glycine in the cerebrum, the target organ of MAM, of 90 days old MAM-induced microencephalic rats were higher than those of control animals. No difference was noted in the contents of these neurotransmitters on postpartum day 21 except for aspartic acid. The postnatal contents of these amino-acid neurotransmitters in the cerebellum, which is not a target organ of MAM, were not affected by prenatal MAM-treatment, except that GABA in 21 days old MAM-induced microencephalic rats was lower than in control rats.
In microencephalic rats, the cerebral cortex, nucleus caudate-putamen, hippocampus, and thalamus-hypothalamus weighed 50% less than in control rats, but the colliculus-midbrain, and cerebellum had similar weights to control rats.
GAD activity per gram wet weight of cerebrum increased rapidly for 21 days after birth, and then increased gradually. No difference was found in GAD activity in MAM-induced micro-encephalic and control rats. The results are discussed in connection with the biochemical adapt-ability of neurons.

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A xyloglucan (MBXG) from the cell walls of etiolated mung bean hypocotyls was characterized by analyzing the fragment oligosaccharides from controlled degradation products of the polymer with acid and enzyme.
Cellobiose, cellotriose and cellotetraose were isolated from the partial acid hydrolyzate of MBXG. Isoprimeverose (6-O-α-D-xylopyranosyl-D-glucopyranose) and a pentasaccharide, α-L-fucosyI-(1→2)-β-D-gaIactosyI-(1→2)-α-D-xylosyI-(1→6)-β-D-glucosyl-(11→4)-D-glucose, were isolated from the hydrolyzate of MBXG with an Asp. oryzae enzyme preparation.

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Three kinds of oligosaccharides, hepta-, nona- and deca-saccharides, were obtained by hydrolysis with Trichoderma viride cellulase from mung bean xyloglucan (MBXG) prepared from cell walls of etiolated mung bean hypocotyls. Structural formulas were proposed for the oligosaccharides from the results of methylation analyses and fragmentation analyses with Aspergillus oryzae β-glucosidase to be as follows:_??_

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L-Leucine-pyruvate transaminase activity increased 6- to 20-fold in 3 hr when Gluconobacter suboxydans cells grown on yeast extract-medium were transferred to and incubated in a nitrogenfree medium. The increase in enzyme activity was influenced remarkably by the age and concentration of cells used. The phenomenon depended upon de novo synthesis of enzyme protein.
The enzyme activity in cell-free extracts of cells incubated under a nitrogen-free condition decreased remarkably after heat treatment at 50°C (pH 6.0) or after freezing and thawing. The level of such enzyme inactivation was high in extracts of cells in the early stages of induction and low in later stages.

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Arthrobacter simplex was screened as an α-keto-δ-guanidinovalerate (ketoarginine) assimilating organism. A characteristic feature was its growth on ketoarginine as a carbon source; it began to grow after an extremely long lag. Its growth was stimulated by addition of 0.02% yeast extract to the medium.
The results indicated the transamination of arginine-α-ketoglutarate (α-KGA) and the hydrolyzing reaction of ketoarginine into α-keto-δ-aminovalerate and urea. Two intermediates, ketoarginine and α-keto-δ-aminovalerate, were isolated and identified by various procedures. Coupling of the two reactions was demonstrated in cell-free extracts of arginine-grown cells; ketoarginine formed from arginine by transamination with α-KGA was hydrolyzed directly to α-keto-δ-aminovalerate and urea. The metabolic routes of arginine in microorganisms were discussed.

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Phospholipase A activity of Pseudomonas aeruginosa was found only in the membrane fraction under normal conditions, but a part of that activity was converted into soluble form after polymyxin B treatment. This soluble phospholipase A was further activated by addition of 80% saturated (NH₄)₂SO₄ or incubation at 70±5°C for 5 minutes. The phenomenon was confirmed in terms of kinetic parameters. Phospholipase A activity in the sonicates of P. aeruginosa was also markedly enhanced by (NH₄)₂SO₂ at 80% saturated concentration. The mechanism of the activation is discussed.

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The properties of creatinine deiminase (EC 3.5.4.21) were characterized with a crystalline preparation from Corynebacterium lilium ATCC 15990. The molecular weight was determined to be 195, 000 by the sedimentation equilibrium method, and the isoelectric point was found to be 4.2 by isoelectric focusing. The enzyme was relatively thermostable and had a broad pH optimum of 7.5 to 10.0. It was specific for creatinine and showed a Km value of 1.27mM. A compound from creatinine was isolated, with the release of ammonia, and identified as N-methylhydantoin. The enzyme activity was inhibited by heavy metal ions and p-chloromercuribenzoate. The enzyme may be useful in determinations of serum and urinary creatinine.

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Amine oxidase of Aspergillus niger was inactivated by ethylenediamine under aerobic conditions, but not under anaerobic conditions. In the presence of ethylenediamine, the oxidized form of the enzyme did not react with phenylhydrazine under anaerobic conditions, but reacted slowly under aerobic conditions. These findings and previous study [Suzuki et al., J. Biochem. 69, 1065 (1971)] suggest that the oxidized form of the enzyme develops an inactive Schiff base between the carbonyl group of the enzyme and the amino group of ethylenediamine under aerobic conditions. The circular dichroic (CD) spectra in the near-ultraviolet region indicated that the structure around the inactive Schiff base was slightly different from that of the reduced form of the enzyme.

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Ribosomal particles of E. coli were examined by using a heat leakage scanning calorimeter. Remarkable changes were observed in thermograms of 70S ribosomes and their subunits when the Mgt²⁺ concentration was raised from 1 rum to 10mM. It was suggested that ribosomal subunits exist in more than one conformation, and changes in their conformation might be the primary cause of the association-dissociation process of ribosomes. Comparisons of thermograms of RNase- and chymotrypsin-treated, as well as non-treated 50S and 30S subunits suggest that conformational changes in each subunit may be ascribed to changes in rRNA.

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Investigations were conducted on the adsorption equilibrium between the air bubbles and some organic materials in waste water. Langmuir's adsorption isotherm held for the adsorption of soluble or insoluble material in single-component solution or suspension, while the extended adsorption isotherm held in the multi-component system. Among organic materials tested, isoelectric protein coagula were most efficiently bubble-separated, although the efficiency of separation was greatly affected by the coexistence of soluble hydrophobic molecules due to competitive adsorption between coagulated protein and small hydrophobic molecules. Pretreatment was suggested of protein-rich waste water without the usual coagulants.

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The adsorption equilibrium between air bubbles and isoelectric coagula of casein was affected by the superficial air velocity and particle size of coagula. An increase in the superficial air velocity V_s increased the affinity between the air bubbles and casein particles, but decreased the maximum surface concentration. On the other hand, an increase in particle size d_p decreased the affinity, but increased the maximum surface concentration. The optimum operating conditions of V_s and d_p existed because of the conflicting effects of them. These results also explain the selective separation of particles of different sizes.

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A callus tissue culture was established from leaves and stems of Mentha spicata L. Lipid constituents isolated from callus tissues were composed of fatty acids, fatty acid methyl esters, triglycerides, squalene, stigmasterol, sitosterol, oleanolic acid (1), ursolic acid (2) and pomolic acid (3). These liquid constituents did not contain monoterpenoids.

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An RNA preparation containing only. 3'-5' phosphodiester linkages was investigated. Such RNA was extracted from yeast packed together with a filter aid (Celite 545) into a column with diluted alkali-salt solution in temperature below 40°C. The RNA from the cells immediately neutralized the surrounding alkali, so that a neutral solution of RNA was obtained as extract. The RNA yield was higher by column extraction than by usual flask extraction with hot salt solution. The enzymatic degradation of RNA during column extraction was prevented by preheating the yeast at 100°C for 2 hr or by extraction in temperature below 0°C, in the presence of high concentration of salt.

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A possible mechanism of fermentation conversion is described from polyalcohol fermentation to ethanol fermentation by Pichia miso. Little alcohol dehydtogenase activity was found in polyalcohol-producing cells, whereas higher enzyme activity was induced by ethanol-producing cells. The fermentation conversion may be caused by the different levels of alcohol dehydrogenase activity between polyalcohol- and ethanol-producing cells. It was also shown that yeast growth was inhibited and that yeast cells were lysed by ethanol (at 6g/100ml) that accumulated in 24hr.

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A bacterial strain capable of assimilating gaseous n-alkanes was newly isolated from activated sludge by enrichment culture technique using n-butane as the sole carbon source. The strain was identified as Pseudomonas butanovora sp. nov. It utilized n-alkanes of C₂_??_C₉, primary alcohols and carboxylic acids for growth, but did not utilize sugars and C₁ compounds. The cell yields on gaseous n-alkanes, such as ethane, propane and n-butane, were 80% or more. The maximum specific growth rate on n-butane was 0.22 hr^-1 at 30°C, pH 7.0. Dried cells of this new isolate grown on n-butane contained 73% pure protein.

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Phenol utilizing yeasts were isolated from soil. The relationship were examined between distribution of phenol uptake rate using intact cells and distribution of the activities of catechol 1, 2-oxygenase which is one of the key enzymes in phenol metabolism. Two of the isolates showed catechol 1, 2-oxygenase activity even when grown in glucose medium, though the enzyme activity was about 1% of the full activity induced by phenol. Partially constitutive mutants for catechol 1, 2-oxygenase were obtained by mutagenesis of an inducible strain. The level of mutant enzyme activity was close to that of the isolated constitutive strain. One isolate, Trichosporon cutaneum, preferentially utilized phenol to glucose in medium containing both phenol (200ppm) and glucose (0.1%), until the concentration of phenol decreased to 10_??_20ppm.

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Cilia regeneration with deciliated Tetrahymena pyriformis W was tested to determine the antitubulinic activities of ansamitocins and related compounds in a microbial system. Various factors interfered with the regeneration process, i.e. excess shearing force in deciliation procedure, high temperature (32°C or above), high or low pH (pH 9 or 5), and exogenous divalent cations, such as Zn²⁺, Mn²⁺, Cu²⁺ and Co²⁺. Under suitable conditions, cilia regeneration was completed in about 60 min of incubation, and a number of newly formed cilia were observed around the cell surface. When ansamitocins were added to the recovery solution, cilia regeneration was completely suppressed without alterations in cell shape or the surface structure of deciliated Tetrahymena. In this assay system, the inhibitory activity of ansamitocins was slightly stronger than that of maytansine.

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The attachment site of the carbohydrate moiety to the peptide chain of normal _K-casein was investigated with _K-casein component P-6 containing the most carbohydrates. Three short glycopeptides 6-IB₁, 6-IB_2-1, and 6-IB_2-2 were prepared by cyanogen bromide cleavage and digestion of proteases (pronase P and thermolysin). Glycopeptides 6-IB₁, 6-IB_2-1, and 6-IB_2-2 corresponded to residues 128_??_139 (Gly-Glu-Pro-Thr-Ser-Thr-Pro-Thr-Thr-Glu-Ala-Val), residues 128_??_132 (or 127_??_131) and residues 135_??_139 of _K-casein A, respectively, and contained threonine and or serine, but not asparagine. Glycopeptide 6-IB₁ was considered to have three carbohydrate chains because it contained three galactosamine residues. The results of alkali treatment of 6-IB₁, under reduction condition excluded serine residue as the binding site, and confirmed the existence of three binding sites in the carbohydrate moieties. The carbohydrate moiety was shown to attach to threonine residue No. 131 from analysis of 6-IB_2-1 and to threonine residue No. 135 (or 136) from analysis of 6-IB_2-2. It was concluded that the carbohydrate chains attached to threonine residues No. 131, 133 and 135 (or 136).

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The neutral constituent sugars of antibiotic K-52B and their glycosidic linkages were examined by methylation analysis and Smith degradation. After partial acid hydrolysis of K-52B, neutral oligosaccharides I, II and III were isolated, and the constituent sugars of each oligosaccharide and their glycosidic linkages were similarly examined. K-52B was found to contain α-D-glc_p, (1→4)-D-gal_p-(1→4)-L-fuc and L-ara_f-(1→4)-D-gal-(1→as neutral sugar fragments.

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A mutant of E. colt (PE4LA) excreted approximately 15% of total cellular protein without cell lysis. The materials in the culture supernatant of the mutant were precipitated with 5% cold TCA. Protein, lipopolysaccharide (LPS), and phospholipid were found in a ratio of approximately 5:6:1. In electrophoretical analyses, exoproteins appeared to contain both periplasmic and outer membrane proteins.
An electron microscopic study showed that PE4LA cells had many blebs around the cell surface and that these blebs were surrounded by double track layers. Some vesicles were also observed as free forms of blebs, while the parent cells had neither blebs nor vesicles. The vesicles appeared to be rich in LPS and lacked phosphatidylglycerol, compared to the outer membrane.
The physiological and morphological data suggested alterations in the PE4LA cell surface, but what was altered remains obscure. It was concluded that PE4LA cells do not have a substantial increase in permeability, but rather have some defect in the cell envelope organization, which causes the formation of blebs with periplasmic proteins.

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An endo-(1→6)-β-D-glucanase capable of hydrolyzing octasaccharide to two tetrasaccharides was isolated from cells of Flavobacterium M64. The octasaccharide represents the repeating unit of succinoglycan (SG-D). One tetrasaccharide was composed of D-glucose, succinic acid and pyruvic acid (4:1:1, molar ratio), and the other was composed of D-glucose and D-galactose (3:1, molar ratio). This enzyme hydrolyzed the (1→6)-β-D-glucosidic linkage adjacent to the (1→6)-linked β-D-glucose residue in the octasaccharide repeating unit of succinoglycan and also hydrolyzed the octasaccharide repeating units of similar polysaccharides produced by many strains of Agrobacterium and Rhizobium species.

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Xylanase induction by β-xyloside was investigated in non-growing conditions using non-induced mycelia of Streptomyces sp. No. 3137 harvested from glucose medium. The mycelia started to produce xylanase without lag time when β-xyloside was added. The rate of xylanase synthesis was dependent on the concentration of β-xyloside added to the inducing culture medium. The induction constants of various β-xylosides were calculated from the Lineweaver-Burk plots; those of methyl-, isopropyl-, butyl- and ethylencyanohydrin-β-D-xylosides were 10.53mM, 3.83mM, 0.55mM and 0.25mM, respectively. Some α-xylosides repressed xylanase synthesis. The rate of xylanase synthesis decreased suddenly after the addition of α-xyloside. The inhibition constants of methyl-, ethyl- and isopropyl-α-D-xylosides were 8.80mM, 12.50mM and 33.33mM, respectively. The xylanase induction was also repressed by glucose. However, this repression was completely restored after consuming additional glucose.

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Pseudo- and hybrid-11S globulins were reconstituted from native acidic and basic subunits of soybean and broad bean 11S globulins. The subunit structures of these two globulins are known to be similar to each other. Pseudo-11S globulins were formed in combinations between glycinin acidic subunit (G-AS₁₊₂) and glycinin basic subunit (G-BS) and between legumin acidic subunit (L-ASII) and legumin basic subunit (L-BS). Hybrid-11S globulins were formed in combinations between G-AS₁₊₂ and L-BS and between L-ASII and G-BS. The yields of the reconstituted 11S components of G-AS₁₊₂+G-BS and G-AS₁₊₂+L-BS were lower than those of L-ASII+G-BS and L-ASII+L-BS. These pseudo- and hybrid-11S globulins were similar to native 11S globulins; they all consisted of reconstituted intermediary subunits which were composed of acidic and basic subunits linked by disufide bridges and had molecular weights similar to those of native 11S globulins. However, the dissociation-association behaviors of pseudo-glycinin and hybrid-11S globulins were different from those of native 11S globulins.

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The presence of α-ketoglutarate (α-KG) dehydrogenase complex in the glutamate-producing bacteria was demonstrated for the first time with Brevibacterium flavum. The partially purified enzyme, which was specific to KG and NAD⁺ with the usual requirements for other co-factors, was labile and stabilized by glycerol, Mgt²⁺, and thiamine pyrophosphate. The enzyme showed an optimum pH of 7.6 and K_ms of 80, 86, and 61μM for KG, NAD⁺, and CoA, respectively. cis-Aconitate, succinyl-CoA, NADPH, NADH, pyruvate, and oxalacetate strongly inhibited the activity, while it was activated by acetyl-CoA, but not by AMP. Various inorganic and organic salts also inhibited the activity. When cells were cultured in glucose and acetate media, the specific activity of the cell extracts increased markedly and reached to a maximum at the late-logarithmic phase. Then, it decreased to the basal level. The addition of glutamate stimulated the synthesis of the enzyme.

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One-dimensional unsteady heat conduction was studied on gels of soy protein (Tofu), gelatin, egg-albumin, milk casein and wheat gluten. These heterogeneous foodstuffs could be treated as homogeneous materials as for the unsteady heat conduction by using the “effective” thermal diffusivity. The effective thermal diffusivity of soy protein gel depended not only on its water content but also on its fat content. The effective thermal diffusivities of non-fat protein gels with the same water content differed among different kinds of proteins.

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Wheat gluten (WG) and fish protein concentrate (FPC) were subjected to alkali treatment under various conditions. The optimum pH for forming lysinoalanine (LAL) was found to be in the neighborhood of 13 in WG and higher in FPC. LAL formation was maximized at 70°C in WG and 90°C in FPC. LAL formation in WG increased at a rapid rate during the initial course of heat treatment. The rate slowed reaching a plateau at a heating time of 2 hr. In FPC, however, LAL formation continued to increase over a longer period of heating. In both cases the amount of LAL formed was not accounted for simply by the amount of cystine or cysteine consumed, and it was estimated that some second factor great contributed as another precursor of dehydroalanine (DHA).
A significant amount of LAL was found to appear by simultating alkali-treatment of performic acid-oxidized WG, which lacks half-cystine. Analysis of the present data indicated that LAL was formed in a severer condition when the second factor acted as a DHA precursor than when cystine and/or cysteine was the precursor.

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Ethylene production and respiratory rate were examined in acid citrus fruits such as yuzu, seedless yuzu and daidai, and wase satsuma mandarin. A large amount of ethylene was produced from four varieties of citrus fruits harvested from May to July but not after September. A rise in ethylene production did not always coincide with a rise in respiratory rate. Excised tissues of fruits contained the ability of ethylene production throughout the developmental stages. The endogenous ethylene level at the ripening stage. reached the maximum when the color changed from green to yellow.

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When DNA-dependent RNA polymerase from Ehrlich ascites tumor cells (E-cells) was directly incubated with 2-ethylsulfonyl-7-methyl-5H-1, 3, 4-thiadiazolo [3, 2-a] pyrimidin-5-one (TPSO₂-2) and dialyzed to remove the chemical, its transcriptive activity was suppressed. The inhibition of enzyme activity was not restored by the addition of substrates. TPSO₂-2 alkylated alcohols in the presence of amines. An alkylthio derivative which had no repressing effect on E-cell propagation did not react with alcohols. TPSO₂-2 showed a high inhibitory effect against “SH enzyme, ” such as yeast alcohol dehydrogenase, but was scarcely inhibited by an alkylthio derivative. TPSO₂-2 reacted with L-cysteine. It is presumed that the reactivity of the 2-position of 1, 3, 4-thiadiazolo [3, 2-a]-pyrimidines is responsible for biological activity and that RNA polymerase is inactivated by the alkylation of the SH or OH group with TPSO₂-2.

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