Cellulase production by Trichoderma reesei when cultured on xylose-based media supplemented with sorbose

The ability of L-sorbose to stimulate cellulase production In shake flask culture of Trichoderma reesei was examined in mineral salts media (initial pH 5.0) containing either 1.0% D-xylose, 1.0% cellulose, and/or 0.1, 0.3, or 0.5% L-sorbose. When sorbose was the only carbon source, growth was limited, little substrate was utilized, pH increased, and cellulase activity was not apparent. The other carbon sources promoted good growth, pH dropped sharply to 2.5-3.0, substrate was utilized rapidly, and cellulase activity was detected. After three weeks of fermentation, twice as much cellulase activity was detected in the medium containing only cellulose as the carbon source, as compared to xylose as the carbon source. Cellulase activity was higher when media contained xylose supplemented with sorbose compared to xylose as the only carbon source. At 0.3 and 0.5% levels of sorbose supplementation of xylose-based media, cellulase activity was similar to that in cellulose-based media.     

The synthesis of yeast pyruvate decarboxylase is regulated by large variations in the messenger RNA level

The yeast PDC1 gene coding for the fermentative enzyme pyruvate decarboxylase was isolated. This DNA sequence was used to identify the corresponding messenger RNA by hybridization. It could be shown that the synthesis of pyruvate decarboxylase is efficiently regulated by variations in the amount of PDC1 mRNA. Very low levels of PDC1 mRNA were found in cells growing in a medium containing ethanol. Glucose addition to these cells leads to a rapid accumulation of PDC1 mRNA. The PDC1 mRNA levels found in different mutants and in cells growing in media containing carbon sources other than glucose or ethanol suggest that the amount of PDC1 mRNA in yeast cells is affected by a number of different factors.     

Petite-negative mutants of Saccharomyces cerevisiae

Summary A series of yeast mutants has been isolated with the inability to grow on fermentable carbon sources whilst growing normally on ethanol media. One of the mutants, namely MC16/206 lacks pyruvate decarboxylase activity and does not grow on glucose at 37°C but grows on both ethanol and glucose at 27°C. In this strain rho ^- petites are non-viable.     

Influence of readily metabolizable carbon on pentachlorophenol metabolism by a pentachlorophenol-degrading Flavobacterium sp.

The influence of high concentrations of pentachlorophenol (PCP) and readily metabolizable carbon on the activity and viability of a PCP-degrading Flavobacterium sp. was examined in a mineral salts medium. Lags preceding PCP removal by glutamate-grown Flavobacterium cells were greatly attenuated by the addition of glutamate, aspartate, succinate, acetate, glucose, or cellobiose. The effect of these supplementary carbon sources on the apparent lag was not mediated entirely through the stimulation of growth since PCP metabolism accompanied the onset of growth. The specific activity of PCP-degrading cells in the absence of supplementary carbon was 1.51 x 10(-13) +/- 0.08 x 10(-13) g of PCP per cell per h and in the presence of supplementary carbon was 0.92 x 10(-13) +/- 0.09 x 10(-13) g of PCP per cell per h. Glutamate in combination with glucose or cellobiose partially repressed PCP metabolism. PCP removal by PCP-induced, glutamate-grown cells suspended in the presence of 4 g of sodium glutamate per liter was sensitive to shock loads of PCP, with a Ki of about 86.8 micrograms/ml. Subsequent removal rates, however, were more resistant to PCP. Optimal stimulation of PCP removal by sodium glutamate required 3.0 g/liter, about the same concentration as that which saturated growth in the absence of PCP. PCP removal rates decayed within minutes following the transfer of PCP-induced, glutamate-grown cells to media containing PCP without supplementary carbon, and increasing PCP concentrations accelerated the decay.(ABSTRACT TRUNCATED AT 250 WORDS)     

Influence of readily metabolizable carbon on pentachlorophenol metabolism by a pentachlorophenol-degrading Flavobacterium sp

The influence of high concentrations of pentachlorophenol (PCP) and readily metabolizable carbon on the activity and viability of a PCP-degrading Flavobacterium sp. was examined in a mineral salts medium. Lags preceding PCP removal by glutamate-grown Flavobacterium cells were greatly attenuated by the addition of glutamate, aspartate, succinate, acetate, glucose, or cellobiose. The effect of these supplementary carbon sources on the apparent lag was not mediated entirely through the stimulation of growth since PCP metabolism accompanied the onset of growth. The specific activity of PCP-degrading cells in the absence of supplementary carbon was 1.51 x 10(-13) +/- 0.08 x 10(-13) g of PCP per cell per h and in the presence of supplementary carbon was 0.92 x 10(-13) +/- 0.09 x 10(-13) g of PCP per cell per h. Glutamate in combination with glucose or cellobiose partially repressed PCP metabolism. PCP removal by PCP-induced, glutamate-grown cells suspended in the presence of 4 g of sodium glutamate per liter was sensitive to shock loads of PCP, with a Ki of about 86.8 micrograms/ml. Subsequent removal rates, however, were more resistant to PCP. Optimal stimulation of PCP removal by sodium glutamate required 3.0 g/liter, about the same concentration as that which saturated growth in the absence of PCP. PCP removal rates decayed within minutes following the transfer of PCP-induced, glutamate-grown cells to media containing PCP without supplementary carbon, and increasing PCP concentrations accelerated the decay.(ABSTRACT TRUNCATED AT 250 WORDS)     

Novel selective and non-selective optical detection of micro-organisms

A new instrument, capable of detecting metabolic changes due to microbiological activity, is described. Optical changes in growth media are monitored in a semi-fluid zone that separates the liquid medium containing the sample. Data demonstrate that common media can be utilized in conjunction with this rapid automated technology. Nutrient broth with the pH dye indicator bromocresol purple was suitable for total counts. Selective media containing dyes were utilized to assess the presence or absence of specific groups of organisms. Biochemical reactions, such as lysine decarboxylase activity, were identified by the unique generated patterns, and specific enzymatic cleavage reactions with chromogenic substrates, such as 5-bromo-4 chloro-3 indolyl-β- D -glucuronic acid (X-GLUC), were monitored.     

Effect of culture medium composition on the activity of extracellular lectins of Lentinus edodes

The time course of lectin production in culture liquid of the basidial fungus Lentinus edodes strain F-249 in different media under the conditions of submerged culture was studied. The activity of agglutinins depended on the ratio between carbon and nitrogen sources and pH of culture medium. The activity of lectin in culture medium was maximal when the fungus was grown in a medium containing L-arabinose as a source of carbon and L-asparagine as a source of nitrogen (C : N ratio, (9.5-12): 1)) on the day 15-18 of culturing at pH 8-9.     

Effect of carbon sources on the synthesis of pectinase by Aspergilli

The synthesis of pectinase is investigated using six species of Aspergillus, with five media differing either in their carbon sources or level of carbon source(s). Five of the six species used, synthesized appreciable amounts of pectinase in the media containing sugars. Pectinase synthesis was highest for A. niger, NCIM 548, with all the sugar containing media. A. foetidus, NCIM 510, was the only one among the organisms studied, that responded well to the medium containing pectin in the absence of additional sugars supplied in the medium.     

Adenosylmethionine decarboxylase from various organisms: Relation of the putrescine activation of the enzyme to the ability of the organism to synthesize spermine

S-adenosyl-L-methionine decarboxylase (EC 4.1.1.50) from most eukaryotic organisms is activated by putrescine whereas the corresponding enzyme from bacterial sources shows a stringent requirement for magnesium ions. Adenosylmethionine decarboxylase from lower eukaryotes such as protozoa, however, is not influenced by diamines, neither are any metals needed for its maximal activity. A common characteristic of those organisms containing putrescine-insensitive adenosylmethionine decarboxylase appeared to be either a total absence or very low intracellular content of spermine. While extracts of all organisms containing putrescine-activated adenosylmethionine decarboxylase (animal tissues and yeast) exhibited easily measurable spermine synthase activity, no such activity was detected in cells of Tetrahymena pyriformis, Escherichia coli or Pseudomonas aeruginosa all containing adenosylmethionine decarboxylase insensitive to putrescine and other diamines.The activation of adenosylmethionine decarboxylase by putrescine, the immediate precursor of spermidine, may thus assure the availability of sufficient amounts of decarboxylated adenosylmethionine (S-methyladenosyl-cysteamine) for the synthesis of spermidine even in the presence of a spermine synthesizing system competing for the same precursor (decarboxylated adenosylmethionine).     

Endoglucanase Activity of Compost-Dwelling Fungus Paecilomyces inflatus is Stimulated by Humic Acids and Other Low Molecular Mass Aromatics

Summary Paecilomyces inflatus isolated from municipal waste compost was found to have cellulolytic activity in several solid and liquid media. This study was done to reveal the multifarious effects of municipal waste compost on endoglucanase activity of P. inflatus. The highest enzyme activities under the conditions of solid-state fermentation were measured in authentic compost samples compared with wood, straw and bran substrates. In surface liquid cultures glucose, cellobiose, xylan, Avicel cellulose, carboxymethylcellulose (CM-cellulose), starch and citrus pectin were used as carbon sources. All carbon sources supported the growth of P. inflatus. However, only CM-cellulose, cellobiose and pectin noticeably stimulated endoglucanase (EG) activity. Further stimulation of EG activity was obtained in cultures containing 1% CM-cellulose as a carbon source by supplementation with low-molecular mass aromatic compounds vanillin, veratric acid and benzoic acid, and with soil humic acid (SHA). SHA and veratric acid were found to be the most efficient elicitors of the cellulolytic activity. P. inflatus was able to utilize nitrate and ammonium as pure nitrogen sources in media containing cellulose.     

Evidence for cycloisomaltooligosaccharide production from starch by Bacillus circulans T-3040

Bacillus circulans T-3040 produces cycloisomaltooligosaccharide glucanotransferase (CITase) and cycloisomaltooligosaccharides (cyclodextrans, CIs) when it is grown in media containing dextran as the carbon source. To investigate the effects of carbon sources on CITase activity, B. circulans T-3040 was cultured with glucose; sucrose; a mixture of isomaltose, isomaltotriose, and panose (IMOs); a mixture of maltohexaose and maltoheptaose (G67); dextrin (average degree of polymerization?=?36); dextran 40; and soluble starch. In addition to dextran 40, CIs were produced when the T-3040 strain was grown in media containing soluble starch as the sole carbon source. CITase production was induced by dextran 40, IMOs, and soluble starch but not by G67 or dextrin, which suggests that α-1,6 glucosidic linkages are required for CITase induction. Although CITase was induced by IMOs, no CIs were produced in the culture. CI-producing activity in the presence of soluble starch as the substrate (SS-CITase activity) was observed only in cultures containing dextran 40 or soluble starch. The production of CITase was significantly unaffected by glucose addition, but SS-CITase activity almost completely disappeared after glucose addition. A 135-kDa protein was found to contribute to CI formation from starch in the presence of CITase. This protein had a disproportionation activity with maltooligosaccharides, and its induction and inhibition system may be different from those of CITase.     

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.     

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 a 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.     

Glycine decarboxylase in Rhodopseudomonas spheroides and in rat liver mitochondria

1. Glycine decarboxylase and glycine–bicarbonate exchange activities were detected in extracts of Rhodopseudomonas spheroides and in rat liver mitochondria and their properties were studied. 2. The glycine decarboxylase activity from both sources is stimulated when glyoxylate is added to the assay system. 3. Several proteins participate in these reactions and a heat-stable low-molecular-weight protein was purified from both sources. 4. These enzyme activities increase markedly when R. spheroides is grown in the presence of glycine, glyoxylate, glycollate, oxalate or serine. 5. All the enzymes required to catalyse the conversion of glycine into acetyl-CoA via serine and pyruvate were detected in extracts of R. spheroides; of these glycine decarboxylase has the lowest activity. 6. The increase in the activity of glycine decarboxylase on illumination of R. spheroides in a medium containing glycine, and the greater increase when ATP is also present in the medium, probably accounts for the increased incorporation of the methylene carbon atom of glycine into fatty acids found previously under these conditions (Gajdos, Gajdos-Török, Gorchein, Neuberger & Tait, 1968). 7. The results are compared with those obtained by other workers on the glycine decarboxylase and glycine–bicarbonate exchange activities in other systems.     

Glyoxylate cycle in Mucor racemosus.

The dimorphic phycomycete Mucor racemosus was grown in media containing acetate, glutamate, and peptone as carbon sources. The component enzymes of the glyoxylate bypass, isocitrate lyase and malate synthase, were present under these conditions throughout the growth cycles. Highest specific activities for each enzyme were found in media with acetate as the carbon source. In an enriched peptone medium containing glucose, neither activity was detected until glucose was exhausted from the medium. Treatment of acetate-grown cells with glucose resulted in a rapid decline in the specific activities of both enzymes. The importance of this cycle in acetate-grown cells was indicated by the ability of itaconic acid (100 mM) to inhibit the growth of M. racemosus in acetate but not glutamate media. Itaconate was also shown to be a potent inhibitor of isocitrate lyase activity in vitro.     

Rat uterine polyamine biosynthetic decarboxylase activities following multiple injections of estradiol-17 beta and/or estriol

A single injection of 0.5 micrograms estradiol-17 beta (E2) plus 0.5 micrograms estriol (E3) stimulated a different pattern in 22-24 day-old rat uterine ornithine decarboxylase (ODC) and S-adenosyl methionine decarboxylase (SAMDC) activities than was induced by either a single injection of 0.5 micrograms E2 or multiple injections of 0.5 micrograms E3. Differences included alterations in enzyme activity peak timing as well as activity duration. Every 3 hour injections of 0.05 micrograms E2 induced maximum uterine ODC activity at 4, 24, 32, and 40 hours, intermediate activity at 48, 64, and 72 hours as well as a small peak by 56 hours. When 0.05 micrograms E2 plus 0.05 micrograms E3 were injected simultaneously every 3 hours, the ODC activity pattern was similar except that activity fell to intermediate levels by 40 hours. It is suggested that E3 alterations of E2 induced uterine enzyme activities (when monitored at frequent intervals) could be physiological alterations in uterine growth responses due to E2-E3 hormone interactions. However, there appeared to be no differences between E2 or E2 plus E3 induction of DNA synthesis and luminal epithelial cell height and cross-sectional area or ODC and SAMDC activities when measured at 24, 48, or 72 hours.     

A simple plate-assay for the screening of l-malic acid producing microorganisms

A simple plate-assay has been developed to screen microorganisms for L-malic acid production. Acid producing organisms were identified, after microbial colony growth on media containing glucose or fumaric acid as sole carbons sources, by formation of a dark halo of formazan. The halo was observed when the plate was covered with a soft agar overlay containing NAD(+)-malate dehydrogenase, NAD+, phenazine methosulfate (PMS) and 3-[4,5-dimethylthiazol-2-yl]-2,5-diphenyltetrazolium bromide (MTT). The assay developed is simple, specific for L-malic acid and therefore can be used to identify L-malic acid producing filamentous fungi using glucose as carbon source (e.g. Aspergillus strains). The assay is also applicable for screening bacteria with high fumarase activity, able to convert fumaric acid to L-malic acid.