Substances which restore impaired cell division in UV-irradiated E. coli B were surveyed among various bacteria. The active substance was found only in several genera of Gram-negative bacteria, i.e., Escherichia, Enterobacter, Salmonella and some species of Pseudomonas. The activity in the dialyzed cell extract of E. coli B/r was observed in the presence of β-NAD and was enhanced by Mg²⁺ and Mn²⁺. The active substance was very labile, but the activity was protected by 1 mM dithiothreitol in the process of purification. The activity of a fraction recovered through DEAE-cellulose column chromatography was stimulated by the presence of membrane fraction. Upon treatment with lipid-degrading enzymes and proteases, the division-stimulating activity was lost or reduced. It appears that the inactivation by lipase and phospholipase A₂ was due to the formation of lysophospholipids and that a proteinous substance participated in the recovery of impaired cell division of UV-irradiated E. coli B.

View full abstract

Two kinds of aldehyde dehydrogenases were found in the supernatant fraction of extracts of sweet potato roots. These enzymes had different Km values for acetaldehyde. The major enzyme, which had a low Km value, was partially purified from fresh sweet potato roots by several steps, including phenyl-Sepharose column chromatography. Purification increased the specific activity 240-fold compared to that at the ammonium sulfate fractionation step. The molecular weight was calculated to be close to 150, 000. In potassium phosphate buffer the optimal pH was approximately 8.0, and, in Tris-HCl buffer, it was approximately 8.8. The Km value for acetaldehyde was 7.0 μM. According to the data obtained from phenyl-Sepharose column chromatography, the enzyme was rather amphipathic in spite of its being a soluble enzyme.

View full abstract

Lysis of commercial baker's yeast cells was examined using Zymolyase. The lysis was stimulated by the addition of sodium sulfite or potassium chloride or both. The effect of potassium chloride was less than that of sodium sulfite, but the two compounds acted synergistically. The cells were effectively lysed by Zymolyase in the presence of 0.1 M sodium sulfite and 0.8 M potassium chloride. The extent of lysis was similar to that of brewery yeast cells obtained from a brewhouse. Cells pretreated with sodium sulfite did not show much of an increase in susceptibility to Zymolyase, but were effectively lysed by the enzyme in the presence of potassium chloride. Potassium chloride stimulated lysis only in the presence of Zymolyase. Yeast cells treated with cupric ions in the presence of sodium sulfite became highly susceptible to Zymolyase, suggesting irreversible destruction of the sodium sulfite-sensitive and potassium chloride-sensitive structure of the cell wall.
Cells of Saccharomyces cerevisiae prepared under various culture conditions were completely lysed by Zymolyase in the presence of sodium sulfite or potassium chloride or both.

View full abstract

Acid phosphatase bound to the cell wall of potato tubers consists of six isozymes. They were separated from one another by chromatography on DEAE-cellulose, phosphocellulose and Sephadex G-200 columns. No significant differences were observed among these isozymes in pH or temperature dependence of activity and thermal stability, but they were distinct from each other in substrate specificity and sensitivity to several compounds. The isozymes were of non-specific acid phosphatase, and characteristic of high affinity for adenosine 5'-monophosphate as compared with the cytoplasmic type of enzyme. Sugar phosphates or glycerophosphate showed lower affinity, and bis-(p-nitrophenyl)phosphate was only slightly hydrolyzed. The molecular weights of the isozymes were estimated to be 240, 000-400, 000, based on gel filtration. The reassociation of each isozyme with the cell wall preparation resulted in no change in substrate specificity. The degree of association was affected by the presence of the substrate or hydrolysis product and depended on their concentrations.

View full abstract

Adenosine-, guanosine-, cytidine- and uridine-5'-di(tri)-phosphate-3'-diphosphates were enzymatically prepared by the use of Streptomyces adephospholyticus ATP nucleotide pyrophosphokinase (E.C. 2.7.6.4). Their regulatory effects were then investigated on translation of rat liver, rabbit globin and silkworm pupae chorion mRNAsby the wheat germ lysate system in vitro. They were found to exert base-specific and mRNA species-specific stimulatory or inhibitory effects, respectively. In particular, cytidine 5'-diphosphate-3'-diphosphate stimulated chorion mRNA translation 3-fold. The significance and limitations of the findings were discussed in relation to natural occurrence and possible regulatory functions of the nucleotides in eucaryotes.

View full abstract

Autotrophic growth was observed in a strain of Hyphomicrobium sp. capable of utilizing methanol. The organism was grownon a mineral salt mediumunder the gas phase of H₂-O₂-CO₂ (66% : 21% : 13%). As nitrogen source, NH₄⁺ was essential and NO₃^- was not useful for autotrophic growth. The specific growth rate μ was measured at various concentration of O₂, and growth was observed at high O₂ concentration (about 35-40%). Hydrogenase activity was assayed by colorimetric and electrochemical methods. Methylene blue and benzylviologen were reduced by the colorimetric method in the presence of a cell free extract with H₂ gas. The electric current in redox dye reduction was detected by an enzyme-fuel-cell. The pH dependency of the hydrogenase activity was observed in both assay methods.

View full abstract

Two kinds of cytochrome c oxidase were partially purified from iron-grown T. ferrooxidans. The first type (cytochrome c oxidase I) was easily solubilized without a detergent and had a pH-optimum at 3.0. The other (cytochrome c oxidase II) which was bound tightly to the cell membrane and solubilized with sodium dodecyl sulfate had a pH-optimum at 5.2. Each type was heat-sensitive and inhibited by cyanide and azide. Since the pH level of the bacterial iron oxidizing activity corresponded closely with the pH of cytochrome c oxidase I but not cytochrome c oxidase II, it was proposed that the former may play an important role in the iron oxidizing system.

View full abstract

Purine nucleoside phosphorylase from Enterobacter cloacae KY3074 was partially purified by ammonium sulfate fractionation, column chromatography on DEAE-cellulose and DEAE-Sephadex A-50, and gel nitration on Sephadex G-100 and Sepharose 4B. The molecular weight of the enzyme was calculated to be about 87, 000 by a gel nitration method on Sephadex G-200. The enzyme was found to be most active at pH 7.5 to 8.5 and 50°C, stable between pH 7.0 and 7.3, and the activity was nearly lost above 70°C. The enzyme split 2'-deoxyinosine and ribonucleosides. Lineweaver-Burk plots for phosphate were non-linear, showing substrate activation. The breakdown of inosine approached an equilibrium when approximately 14% of inosine was phosphorylated.

View full abstract

Strain No. 14 that grew best in medium containing o-cresol at high concentration (0.01 %) as the sole carbon source was selected out of 22 strains of Aureobasidium spp. and its allied organisms, "Black Yeasts." The strain was investigated morphologically and taxonomically, and the conclusion was that it belonged to Aureobasidium pullulans (de Bary) Arnaud. The strain utilized phenol rapidly at a concentration as high as 0. 1 %, even when 3% methanol or 0.008% formaldehyde was present in medium. Based upon these results, strain No. 14 can probably be applied effectively to treat phenolic waste water, especially from the synthetic resin industry.

View full abstract

Multiple forms of lysozyme found in the rat liver were isolated and characterized from cellular organelles.
Isolation of the enzymes was achieved by Sephadex gel filtration and chromatography on CM cellulose-column. The purity of the preparations was examined by electrophoresis on polyacrylamide gel. The nuclear lysozyme moved as a single band indicating homogeneity, whereas other subcellular lysozymes appeared heterogeneous due to presence of more than one band, thus showing partial purity. Although the subcellular lysozymes were similar with respect to enzymatic properties, pH, buffer molarity optima and electrophoretic mobility, differences were observed in elution patterns, responses to nuclear inhibitor and heat sensitivity. Nuclear lysozyme was distinctly different by these criteria as compared to other subcellular lysozymes.

View full abstract

Tunicamycin resistant mutants (TM^R)were isolated and characterized from Chinese hamster ovary cells. One feature of this TM^R mutants was a marked decrease in incorporation of radioactive glucosamine, both into membraneglycoproteins and G protein of vesicular stomatitis virus.
The cellular uptake and incorporation into acid insoluble materials of various radioactive substances, including glucosamine, galactosamine, mannose, 2-deoxyglucose and leucine, was examined for the purpose of determination whether the reduced incorporation of radioactive glucosamine into glycoproteins was due to a defect in the glycosylation step or decreased uptake of glucosamine by cells.
While incorporation of glucosamine and 2-deoxyglucose into acid insoluble fractions was
reduced strikingly in the mutants, the incorporation of mannoseand leucine were the same as in the parent cells.
The uptake of glucosamine in TM^R cells was lower than that in the wild type cells, and the Km value for glucosamine uptake differed between the mutants and wild type cells. There was no obvious difference in the uptake of 2-deoxyglucose and mannose.

View full abstract

ent-15α-Hydroxykaurenoic acid (8) was synthesized and fed to a myceliumsuspension of Gibberella fujikuroi in the presence of 1-n-decylimidazole, a gibberellin biosynthesis inhibitor. The metabolites included 15β-hydroxy GA₂₄, GA₄₅ (GA of Pyrus communis), 15β-hydroxy GA₁₅ and 15β-hydroxy GA₂₅. Microbial production of 12α-hydroxy GAs from ent-12β-hydroxykaurene is also described.

View full abstract

A comparative study of selective cytotoxicity between non-transformed and SV40-transformed cells has been undertaken in combination with antitumor agents having different mechanisms of action. The selective in vitro cytotoxicity assay (N/T assay) measured the decrease in cell proliferation under the microscope. SV40-transformed cells had higher sensitivity than non-transformed cells against these antitumor agents at low concentration; but an apparent dissimilar pattern of sensitivity was observed in some antimetabolites, such as cytosine arabinoside, 6-mercaptopurine, 5-fluorouracil, and in some inhibitors of energy-transfer. The results can be interpreted in terms of a possible difference in cytotoxicity between non-transformed and SV40-transformed cell lines. The N/T assay may be considered a convenient system for evaluating selective cytotoxicity for in vitro-in vivo comparisons.

View full abstract

Sixteen kinds of flavonoids including five anthocyanins, two flavones and nine flavone glycosides were found to be present in mature dark-red leaves and seeds of Perilla frutescens var. crispa. In seeds, apigenin and luteolin were present in a ratio of about 1 : 1. Both flavones and nine flavone glycosides were found in the leaves. In addition, the leaves contained five kinds of anthocyanins including cyanidin 3, 5-diglucoside and its esters with cinnamic acid derivatives. Amongthese flavonoids, the 3-p-coumarylglucoside-5-glucoside of cyanidin (shisonin) and the 7-caffeylglucosides of apigenin and luteolin were the major component in the leaves.

View full abstract

This paper reports on a total synthesis of racemic δ-cadinene (2), which had been obtained previously in optically active form by acid-catalyzed cyclization of (-)-germacrene D. The Robinson annelation using cyclohexenone enamine (12) proceeded stereoselectively to form δ-cadinenone (3), whose oxygen was removed by the thioketal-Raney Ni method to produce δ-cadinene.

View full abstract

The functional relationship between bacteriophage φX174 and SI3 gene products was investigated using an in vitro DNA synthesis system in two stages: (a), replication of the closed circular duplex form DNA (RF to RF) and (b) synthesis of single-stranded DNA from RF and formation of the intermediate 50S complex in phage morphogenesis (RF to 50S complex). Partially purified gene A products of φX174 and S13 were able to function reciprocally in the stage from RF to RF replication. In the stage of RF to 50S complex, extracts from Escherichia coli infected with φX174 catalyzed the formation of 50S complex from S13 RF DNA. Extracts from E. coli infected with SI3 also had the ability to catalyze the formation of 50S complex from φX174 RF DNA. Thus, all gene products required for DNA replication and phage morphogenesis were found to function reciprocally between φX174 and S13.

View full abstract

Twelve strains of Chlorella which lack the sporopollenin layer in their cell wall were treated with polysaccharide degrading enzyme mixtures. Osmotically labile protoplasts were obtained from two of them (C. ellipsoidea C-87 and C. saccharophila C-211). The absence of the cell wall was demonstrated by the Calcofluor stain and by electron microscopies. Some protoplasts adhered to each other; it seemed like a cell fusion. Protoplasts of C. ellipsoidea C-87 were able to regenerate the cell wall and to grow on a regeneration medium.

View full abstract