Growth and production of rifamycin oxidase byCurvularia lunata

Curvularia lunata could neither grow nor produce rifamycin oxidase in synthetic media without peptone and yeast extract. Mycelia grown on complex media were tested for the ability to produce rifamycin oxidase in synthetic media. The optimum concentrations of peptone and yeast extract were in the range of 7.5–10 g/L. Five percent inoculum size was found to be optimum for good growth and enzyme production. Addition of metal ions to the cultivation medium increased the enzyme activity.     

The fermentation of L-sorbose by Gluconobacter melanogenus. I. General characteristics of the fermentation

Growing cultures, washed cells, and cell-free preparations of Gluconobacter melanogecnus IFO 3293 converted L -sorbose to 2-keto-L -gulonic acid, to D -sorbitol (which was metabolized further) and to 5-keto-D -fructose.     

Conversion of L-sorbose to L-sorbosone by Gluconobacter melanogenus

Growing cultures, washed cells, and cell-free extracts of Gluconobacter melanogenus IFO 3293 were found to convert L -sorbose to L -sorbosone. The product was identified by thin layer chromatography of the 2, 4-dinitrophenylhydrazone, and by paper partition chromatography using chemically prepared materials as standards. Factors influencing the conversion included incubation temperature and composition of the growth medium. Addition of betaine or choline to the growing cultures stimulated conversion of L -sorbose to L -sorbosone.     

Frammenti cotiledonari di Aesculus hippocastanum L. coltivati in vitro. Primi dati sul comportamento dell'amido e dell'escina

Abstract

Fragments of Aesculus hippocastanum L. cotyledons grown in vitro. First results about starch and aescin characteristic features.—Cotyledon fragments of Aesculus hippocastanum grown in vitro in different media have been able to form callus and roots. The starch granules in the new cells are compound in structure and morphologically different from the simple cotyledon granules, whereas they are similar to the granules of the other parts of the plant in toto. Moreover, the callus has no aescin even though it originates from the cotyledor tissues.     

Growth of baby hamster kidney cells in media containing neuraminidase

An established line of baby hamster kidney cells, BHK 21/C13 grows in media containing high concentrations of Vibrio cholerae neuraminidase at the same initial exponential rate as control cells grown in the absence of the enzyme. Glycoprotein fractions removed from the surface of cells grown with neuraminidase contain less than 4% of the sialic acid present in similar fractions removed from control cells. The significance of these results are discussed in relation to the role suggested for sial glycoproteins in growth control. A small but significant increase was observed in the density of confluent cells in media containing neuraminidase compared with control cultures.     

Effect of Cultivation Conditions on the Growth and Activities of Sulfur Metabolism Enzymes and Carboxylases of Sulfobacillus thermosulfidooxidans subsp. asporogenes Strain 41

The moderately thermophilic acidophilic bacterium Sulfobacillus thermosulfidooxidans subsp. asporogenes strain 41 is capable of utilizing sulfides of gold–arsenic concentrate and elemental sulfur as a source of energy. Growth in the presence of S⁰ under auto- or mixotrophic conditions was less stable than in media containing iron monoxide. The enzymes involved in the oxidation of sulfur inorganic compounds—thiosulfate-oxidizing enzyme, tetrathionate hydrolase, rhodanase, adenylyl phosphosulfate reductase, sulfite oxidase, and sulfur oxygenase—were determined in the cells of the sulfobacilli grown in mineral medium containing 0.02% yeast extract and either sulfur or iron monoxide and thiosulfate. Cell-free extracts of the cultures grown in the medium with sulfur under auto- or mixotrophic conditions displayed activity of the key enzyme of the Calvin cycle—ribulose bisphosphate carboxylase—and several other enzymes involved in the heterotrophic fixation of carbon dioxide. Activities of carboxylases depended on the composition of the cultivation media.     

Organogenesis in Callus from Shoot Apices of Pisum sativum

Shoot formation was observed in callus from apical cells of pea (Pisum sativum L. cv. Century). Shoot apices from 4-day-old plants were macerated and the resulting cell masses grown on agar media. The callus formation and shoot production occurred within 4 to 6 weeks in defined media containing 0.2 to 5.0 μM benzyladenine and 1 μM naphthaleneacetic acid. While most callus produced one or more shoots at high frequency, root formation did not occur regularly. Plants obtained by these procedures were grown to maturity producing flowers and pods.     

Adventitious shoot formation in cultures of 30-year-old Larix decidua,L. leptolepis,L. eurolepis,and L. laricina trees

Primordial shoot explants excised from buds of one Larix decidua tree, about 30 years old, produced more adventitious buds, elongating into shoots, when grown on half strength Litvay medium than when grown on other basal media. Thidiazuron and N₆-benzyladenine (BA) were equally effective in adventitious bud induction. In a comparative study of 30-year-old L. decidua, L. leptolepis, L. eurolepis, and L. laricina trees, explants from L. eurolepis and L. decidua produced a high number of cultures with adventitious buds that elongated into shoots; those from L. leptolepis were less productive, and those from L. laricina failed to form adventitious buds. The highest response was obtained with material collected in August and September, and in March and April; the lowest response occurred in explants from the October collection.     

Effects of external osmolality on polyamine metabolism in HeLa cells

The polyamine content of Escherichia coli is inversely related to the osmolality of the growth medium. The experiments described here demonstrate that a similar phenomenon occurs in mammalian cells. When grown in media of low NaCl concentration, HeLa cells and human fibroblasts were found to contain high levels of putrescine, spermidine, and spermine. The putrescine content of HeLa cells was a function of the osmolality of the medium, as shown by growing cells in media containing mannitol or additional glucose. External osmolality per se had no effect on the contents of spermidine and spermine. For all media, the total cellular polyamine content could be correlated with the activity of ornithine decarboxylase, the first enzyme in polyamine biosynthesis. Different levels of enzyme activity appear to result solely from variations in the rate of enzyme degradation.A sudden increase in NaCl concentration produced rapid loss of ornithine decarboxylase activity and a gradual loss of putrescine and spermidine. A sudden decrease in NaCl concentration led to rapid and substantial increases in ornithine decarboxylase activity and putrescine.     

Potassium Depletion and Regulation of Angiotensin II Receptors in Vascular Smooth Muscle Cells

Abstract

Mesenteric artery smooth muscle cells were grown in culture media containing high, normal, or low concentrations of potassium to study the effects on angiotensin II (Ang II) receptor regulation. Cell growth was similar among cells grown in the different culture media. Cells grown in high potassium media (K=5.8 mEq/L) had an equilibrium dissociation constant, K_d, of 1.59 ± 0.2 nM, whereas those grown in normal potassium media (K=4.1 mEq/L) had a K_d of 1.79 ± 0.2 nM and those grown in a low potassium media (K=2.9 mEq/L) had a K_d of 1.19 ± 0.12 nM (not significantly different, NS). Binding capacity of smooth muscle cells grown in high potassium media was 81 ± 16.7 fmol/mg prot, 95.1 ± 12.4 fmol/mg prot in those grown in normal potassium media and those grown in low potassium media 86.4 ± 24.1 fmol/mg prot (NS). Binding of radiolabelled Ang II was reduced by approximately 70% in cells exposed to unlabelled Ang II for 30 or 60 minutes. However, this effect of exposure to Ang II to reduce subsequent binding of Ang II was identical in cells grown in high and low potassium medium. Therefore, we were unable to identify a direct effect of low potassium to induce changes in Ang II receptor binding affinity or binding capacity. Previously observed changes in these Ang II binding parameters in potassium-depleted rats was probably a consequence of other factors which were simultaneously altered by potassium deficiency.     

Purification and Properties of Uricase from Enterobacter cloacae

Uricase activity was found in Enterobacter cloacae KY3074 grown on guanine, hypoxanthine, uric acid, and xanthine media. The enzyme was purified from cells grown on uric acid as a source of nitrogen. The purification procedure included ammonium sulfate fractionation, gel filtration on Sephadex G-150, and column chromatography on DEAE-cellulose and DEAE-Sephadex. The enzyme had a molecular weight of about 105,000 and was specific for uric acid. The optimum pH was around 9.5, and the activity was inhibited by the presence of potassium cyanide, Ag⁺ or Cu²⁺. This uricase can be used for estimation of uric acid.     

Effect of carbon and nitrogen growth limitation upon nutrient uptake and metabolism in batch cultures of Catharanthus roseus (L) G. Don

Cell suspension cultures of the Madagascan Periwinkle, Catharanthus roseus (L). G. Don were grown as batch cultures in two different types of media; in one medium the limiting nutrient was inorganic nitrogen, and in the other it was carbon. The response of the cells to these growth-limiting conditions was monitored by measuring cellular fresh weight, dry weight and protein accumulation, cell viability, medium sugar and nitrate levels, and the activities of certain intracellular enzymes throughout growth in batch culture. The enzymes investigated were glucose-6-phosphate dehydrogenase (EC 1.1.1.49), 6-phosphogluconate dehydrogenase (EC 1.1.1.44), hexokinase (EC 2.7.1.40), phosphofructokinase (EC 2.7.1.11), nitrate reductase (EC 1.6.6.1), glutamate dehydrogenase (EC 1.4.1.2) and glutamine synthetase (EC 6.3.1.2). The effect of culturing the cells under different nutritional regimes was apparent in all aspects of growth; only some enzyme activities were unaffected. Cell viability remained at a high level for several days after growth limitation in both types of culture. The possibility that protein degradation in nitrogen-limited batch cultures is under very stringent control is discussed.     

Synthesis of L-asparaginase bySerratia marcescens (Nima)

The production of L-asparaginase by two mutants ofSerratia marcescens grown on 14 different media was studied. The enzyme content increased from trace levels to 2.4 international units per ml when the organisms were grown in glycerol-peptone yeast extract medium. Glucose was the best carbon source under aerobic conditions. The enzyme content increased when L-asparagine was present in the growth medium.     

Extracellular Chitinases of Fluorescent Pseudomonads Antifungal to Fusarium oxysporum f. sp. dianthi Causing Carnation Wilt

Vascular wilt of carnation caused by Fusarium oxysporum f. sp. dianthi (Prill. & Delacr.) W. C. Synder & H.N. Hans inflicts substantial yield and quality loss to the crop. Mycolytic enzymes such as chitinases are antifungal and contribute significantly to the antagonistic activity of fluorescent pseudomonads belonging to plant-growth-promoting rhizobacteria. Fluorescent pseudomonads antagonistic to the vascular wilt pathogen were studied for their ability to grow and produce chitinases on different substrates. Bacterial cells grown on chitin-containing media showed enhanced growth and enzyme production with increased anti-fungal activity against the pathogen. Furthermore, the cell-free bacterial culture filtrate from chitin-containing media also significantly inhibited the mycelial growth. Both the strains and their cell-free culture filtrate from chitin-amended media showed the formation of lytic zones on chitin agar, indicating chitinolytic ability. Extracellular proteins of highly antagonistic bacterial strain were isolated from cell-free extracts of media amended with chitin and fungal cell wall. These cell-free conditioned media contained one to seven polypeptides. Western blot analysis revealed two isoforms of chitinase with molecular masses of 43 and 18.5 kDa. Further plate assay for mycelial growth inhibition showed the 43-kDa protein to be antifungal. The foregoing studies clearly established the significance of chitinases in the antagonism of fluorescent pseudomonads, showing avenues for possible exploitation in carnation wilt management.     

The effect of alkanes on the physiology and metabolism of the entomopathogenic fungus,Isaria fumosorosea

The influence of different alkanes on spore morphology, glyoxlate pathway enzyme activities, total lipid contents and fatty acid composition of Isaria fumosorosea were investigated under laboratory conditions. Fungal spores grown on different alkanes showed higher germination and mycelial growth when compared to control. A strong induction of glyoxlate cycle enzymes in cell-free extracts was observed for cells grown on different alkanes when compared to glucose and control. Higher activities of glyoxlate cycle enzymes were observed for cells grown on alkanes when compared to other treatments. Even numbered fatty acids accounted for the majority of fatty acid production with a significant increase in relative amounts of linoleic acid and palmatic acid observed for conidia grown on alkanes. These results indicate that addition of alkanes to culture media can be a tool to pre-induce metabolic adaptations that can facilitate successful infection of insect host by entomopathogenic fungi.     

Characterization of a yeast regulatory mutant constitutive for synthesis of inositol-1-phosphate synthase

Summary The enzyme inositol-1-phosphate synthase is repressed at least 50-fold in wild type yeast grown in inositol-supplemented media. Mutants which synthesize this enzyme constitutively have been isolated using a selection procedure based on excretion of inositol into the growth medium by putative mutants. Biochemical analysis of one of the mutants (opi1-1) confirmed that the nature of the mutations is regulatory, and not in the structural gene for the enzyme. Immunoprecipitation of crude extracts with antibody directed against purified inositol-1-phosphate synthase showed that a protein which reacts with the antibody is present in the mutant grown under both repressing and derepressing conditions, in contrast to the wild type which synthesizes the enzyme only when derepressed. Assay of inositol-1-phosphate synthase activity in crude extracts of the mutant verified synthase activity in cells grown under both repressing and drepressing conditions. Synthase purified from this mutant was characterized with respect to molecular weight, thermolability and affinity for substrates glucose-6-phosphate and NAD. These analyses indicated that purified mutant synthase was similar to the wild type enzyme.     

Transitory pulse-like treatment with IAA solution more effectively induces xylogenesis in callus culture than permanent presence of the auxin

Differentiation of Acerpseudoplatanus L. cells into tracheary elements (TE) and an increase in the number of TE inHaplopappus gracilis (Nutt.) A. Gray calli were observed after pulse-treatment of the cultures grown in glass tubes with indole-3-acetic acid (IAA) solutions. The effect was enhanced if the treatment was repeated in three subsequent days. The manner of IAA application which caused a wave-like pattern of IAA flow through the callus culture appeared to be more important than the IAA concentration. Induction of differentiation of A.pseudoplatanus cells and an increase in the number of TE inH. gracilis callus did not occur when the calli were grown on agar media supplemented with increased concentrations of IAA.     

Potentiality of leaf sheath cells for regeneration of rice (Oryza sativa L.) plants

Summary Regeneration of rice plantlets (Oryza sativa L.) from calli originated from leaf sheath cells was made possible. This was possible in tissues initially grown in media containing 2.4-D (2.4-dichlorophenoxyacetic acid) at low temperature and illumination. The slow growing tissues were subsequently subjected to growth conditions at an elevated temperature and higher illumination with addition of kinetin and IAA and without 2.4-D. The suitability of leaf sheath cells for protoplast technology is indicated by this success.     

Control of Tryptophan Biosynthesis in Plant Tissue Cultures: Lack of Repression of Anthranilate and Tryptophan Synthetases by Tryptophan

Tobacco (cv. Xanthi and cv. Wisconsin 38), rice, carrot, tomato, and soybean tissue cultures were grown in liquid media containing L-tryptophan. The addition of tryptophan increased the cellular tryptophan levels greatly (12–2500 fold), but did not lower appreciably the levels of two tryptophan biosynthetic enzymes, anthranilate synthetase and tryptophan synthetase. However, the addition of 50 μM tryptophan to the crude enzyme extract completely inhibited the anthranilate synthetase activity while 1 mM tryptophan inhibited the tryptophan synthetase activity by only 10–20°/o. This information indicates that tryptophan biosynthesis is controlled by the feedback inhibition of anthranilate synthetase by tryptophan and not by repression of enzyme synthesis. All of the species had significant enzyme levels. Anthranilate synthetase activity could not be detected in extracts from cells grown on tryptophan unless the extracts were first passed through two G-25 Sephadex columns with a short 30 °C warming step in between, a procedure shown to remove an inhibitor of the enzyme.