Nitrogen source regulates expression of alanine dehydrogenase isoenzymes in Streptomyces avermitilis in a chemically defined medium

1997 ◽  
Vol 43 (2) ◽  
pp. 189-193 ◽  
Author(s):  
Jan Novák ◽  
Jan Kopecký ◽  
Zdenko Vaněk
Keyword(s):  
Nitrogen Source ◽  
Ammonium Sulfate ◽  
Alanine Aminotransferase ◽  
Specific Activity ◽  
Defined Medium ◽  
Streptomyces Avermitilis ◽  
Chemically Defined Medium ◽  
Ammonium Ions ◽  
Alanine Dehydrogenase ◽  
Dehydrogenase Reaction

Ammonium ions and alanine influence production of the macrolide avermectin in Streptomyces avermitilis. L-Alanine dehydrogenase and alanine aminotransferase are the primary enzymes responsible for regulating the intracellular concentration of alanine and also of ammonium ions. In cultures of S. avermitilis in a chemically defined medium with ammonia or L-alanine as the only nitrogen source, specific activities of both enzymes increased during growth. The alanine dehydrogenase specific activity increased more than 86-fold after the culture was supplemented with 0.2% L-alanine and 5-fold after addition of 0.5% ammonium sulfate, whereas alanine aminotransferase specific activity increased 3- to 4-fold with either substrate. Five isoenzymes of alanine dehydrogenase were detected histochemically in S. avermitilis after native gel electrophoresis. Isoenzyme 1 was induced by alanine and temporarily repressed by high concentrations of ammonium sulfate. The presence of isoenzyme 1 was also related to changes in the kinetic properties of the alanine dehydrogenase reaction measured in crude desalted extracts. A nonlinear double-reciprocal plot was obtained in initial velocity studies using L-alanine as a substrate in the sample induced with L-alanine. The nonlinearity was caused by both substrate inhibition and allosteric regulation (positive cooperativity) by L-alanine. In contrast, the sample induced by ammonium sulfate showed a linear double-reciprocal plot.Key words: isoenzymes, L-alanine dehydrogenase, Streptomyces avermitilis, avermectin.

scholarly journals l-Alanine Auxotrophy of Lactobacillus johnsonii as Demonstrated by Physiological, Genomic, and Gene Complementation Approaches

2004 ◽  
Vol 70 (3) ◽  
pp. 1869-1873 ◽  
Author(s):  
Hengameh van der Kaaij ◽  
Frank Desiere ◽  
Beat Mollet ◽  
Jacques-Edouard Germond
Keyword(s):  
Bacillus Subtilis ◽  
Amino Acid ◽  
Genetic Analysis ◽  
Heterologous Expression ◽  
Defined Medium ◽  
Chemically Defined Medium ◽  
Lactobacillus Johnsonii ◽  
Gene Complementation ◽  
Alanine Dehydrogenase ◽  
Comparative Genetic Analysis

ABSTRACT Using a chemically defined medium without l-alanine, Lactobacillus johnsonii was demonstrated to be strictly auxotrophic for that amino acid. A comparative genetic analysis showed that all known genes involved in l-alanine biosynthesis are absent from the genome of L. johnsonii. This auxotrophy was complemented by heterologous expression of the Bacillus subtilis l-alanine dehydrogenase.

UTILIZATION OF CARBOHYDRATES BY ACTINOBACILLUS MALLEI

1966 ◽  
Vol 12 (4) ◽  
pp. 625-639 ◽  
Author(s):  
D. H. Evans
Keyword(s):  
Methylene Blue ◽  
Nitrogen Source ◽  
Ammonium Chloride ◽  
Carbon Sources ◽  
Defined Medium ◽  
Ammonium Citrate ◽  
Chemically Defined Medium ◽  
Negative Results ◽  
Proteose Peptone ◽  
Positive Results

Twenty-three substrains representing colonial variants of 11 strains of Actinabacillus mallei were examined for their ability to attack carbohydrates. Tests conducted in a basal liquid complex medium, containing yeast extract and proteose peptone No. 3 with bromcresol purple as indicator, showed that all strains tested produced acid from arabinose, glucose, fructose, galactose, mannose, and trehalose, while five substrains gave positive results with lactose, one with sucrose, and two with maltose. Eosin methylene blue agar of the same basal composition gave positive results for most of the strains grown on arabinose, glucose, fructose, galactose, mannose, and trehalose, and negative results for all strains grown on xylose, lactose, sucrose, and maltose. In a chemically defined medium containing ammonium chloride as nitrogen source and bromcresol purple as indicator, acid was produced by eight substrains of five of these strains from glucose, galactose, mannose, and trehalose, and by several strains from fructose and sucrose. The ability of these five selected strains to utilize carbohydrates as sole carbon sources for growth was tested in a chemically defined medium containing ammonium citrate as nitrogen source. All strains were able to grow on glucose, galactose, mannose, and trehalose, and most were able to grow on fructose. Arabinose, xylose, lactose, sucrose, and maltose did not support the growth of any of the strains tested.

scholarly journals The Growth Of Rhizobium in Synthetic Media

1961 ◽  
Vol 14 (3) ◽  
pp. 349 ◽  
Author(s):  
FJ Bergersen
Keyword(s):  
Amino Acids ◽  
Nitrogen Source ◽  
Exponential Growth ◽  
Defined Medium ◽  
Complete Medium ◽  
Good Growth ◽  
Chemically Defined Medium ◽  
Sodium Glutamate ◽  
Synthetic Media

A chemically defined medium for the growth of Rhizobium is described in which populations of up to 5 x 109 cells/ml were obtained. For the six strains of bacteria studied the complete medium supported exponential growth for two to five generations. The concentrations of biotin giving best growth varied ith strain between 125 and 250 f'g/l when the nitrogen source was sodium glutamate. NHt, NOs, and other amino acids, singly or in combination, did not upport as good growth as did sodium glutamate.

Myoinositol biosynthesis by Sertoli cells, and levels of myoinositol biosynthetic enzymes in testis and epididymis

1979 ◽  
Vol 57 (6) ◽  
pp. 962-967 ◽  
Author(s):  
Ranga Robinson ◽  
Irving B. Fritz
Keyword(s):  
Sertoli Cells ◽  
Specific Activity ◽  
Primary Cultures ◽  
Defined Medium ◽  
Seminiferous Tubules ◽  
Chemically Defined Medium ◽  
Dibutyryl Cyclic Amp ◽  
Old Rats ◽  
Specific Activities ◽  
Germinal Cells

Levels of glucose-6-phosphate cyclase (myoinositol-1-phosphate synthase, EC 5.5.1.4) and myoinositol-1-phosphate phosphatase (myoinositol-1-phosphatase, EC 3.1.3.25) were determined in extracts of testes from 10-, 20-, and 30-day-old rats, and in extracts of Sertoli cells, germinal cells, and epididymides. The specific activity of the cyclase was approximately [Formula: see text] that of the phosphatase in all extracts found to contain either enzyme. Among cells in the testis examined, Sertoli cells had highest levels of enzymes required for inositol biosynthesis from glucose, while spermatocytes and round spermatids did not have detectable activity. Spermatozoa from the epididymis also had no detectable cyclase or phosphatase activity. In contrast, extracts of washed epididymides contained exceedingly high specific activities of these enzymes. Primary cultures of Sertoli cells, maintained in a chemically defined medium without added inositol, released inositol into the medium during three successive 24-h periods. The amounts released were greater in cells stimulated by dibutyryl cyclic AMP. Results were interpreted to indicate that inositol in the fluid of seminiferous tubules most probably originates from Sertoli cells, which synthesize inositol from glucose. Additional inositol in the fluid of epididymal tubules could readily be provided by metabolism of glucose by epididymal epithelial cells.

UTILIZATION OF SODIUM CASEINATE BY LACTIC STREPTOCOCCI AND ENTEROCOCCI

1956 ◽  
Vol 2 (6) ◽  
pp. 607-610 ◽  
Author(s):  
I. J. McDonald
Keyword(s):  
Nitrogen Source ◽  
Sodium Caseinate ◽  
Defined Medium ◽  
Starter Cultures ◽  
Sole Nitrogen Source ◽  
Chemically Defined Medium ◽  
Casein Hydrolyzate ◽  
Selection Of

The ability of lactic streptococci to utilize unhydrolyzed sodium caseinate as the nitrogen source in an otherwise chemically defined medium seems to be a strain rather than a species characteristic. This characteristic therefore appears to have little value as an aid in classification although it might have some importance in selection of starter cultures for cheese manufacture. None of the enterococci examined grew with unhydrolyzed sodium caseinate as the sole nitrogen source. However, Streptococcus liquefaciens and Streptococcus zymogenes appeared to utilize sodium caseinate when small amounts of casein hydrolyzate were added to the medium.

Ammonium ions repress δ-(L-α-aminoadipyl)-L-cysteinyl-D-valine synthetase in Streptomyces clavuligerus NRRL 3585

1989 ◽  
Vol 35 (3) ◽  
pp. 399-402 ◽  
Author(s):  
Jinyou Zhang ◽  
Saul Wolfe ◽  
Arnold L. Demain
Keyword(s):  
Streptomyces Clavuligerus ◽  
Defined Medium ◽  
Antibiotic Biosynthesis ◽  
Chemically Defined Medium ◽  
Ammonium Ions ◽  
Rate Limiting Step ◽  
Limiting Step ◽  
Negative Effect ◽  
Major Factors ◽  
Rate Limiting

Production of cephems (predominantly cephamycin C) by Streptomyces clavuligerus grown in chemically defined medium supplemented with 120 mM NH4Cl was sharply reduced. This concentration of ammonium ions in the medium repressed δ-(L-α-aminoadipyl)-L-cysteinyl-D-valine (ACV) synthetase formation by about 75%. Of the other cephalosporin synthases, cyclase was repressed by 70%, expandase by 50%, and epimerase only to a very small extent. Inhibition of the action of ACV synthetase was only slight in the presence of 100 mM NH4Cl. Repression of ACV synthetase, cyclase, and expandase appears to be the major factors contributing to the negative effect of ammonium on S. clavuligerus NRRL 3585. ACV synthetase is probably the rate-limiting step of cephalosporin biosynthesis in this strain.Key words: β-lactam biosynthesis, antibiotic biosynthesis, cephalosporins, Streptomyces clavuligerus, ACV synthetase.

scholarly journals Aspartate Biosynthesis Is Essential for the Growth of Streptococcus thermophilus in Milk, and Aspartate Availability Modulates the Level of Urease Activity

2007 ◽  
Vol 73 (18) ◽  
pp. 5789-5796 ◽  
Author(s):  
Stefania Arioli ◽  
Christophe Monnet ◽  
Simone Guglielmetti ◽  
Carlo Parini ◽  
Ivano De Noni ◽  
...  
Keyword(s):  
Aspartic Acid ◽  
Phosphoenolpyruvate Carboxylase ◽  
Parent Strain ◽  
Urease Activity ◽  
Streptococcus Thermophilus ◽  
Defined Medium ◽  
Lactobacillus Delbrueckii ◽  
Chemically Defined Medium ◽  
Ammonium Ions ◽  
Carboxylase Activity

ABSTRACT We investigated the carbon dioxide metabolism of Streptococcus thermophilus, evaluating the phenotype of a phosphoenolpyruvate carboxylase-negative mutant obtained by replacement of a functional ppc gene with a deleted and inactive version, Δppc. The growth of the mutant was compared to that of the parent strain in a chemically defined medium and in milk, supplemented or not with l-aspartic acid, the final product of the metabolic pathway governed by phosphoenolpyruvate carboxylase. It was concluded that aspartate present in milk is not sufficient for the growth of S. thermophilus. As a consequence, phosphoenolpyruvate carboxylase activity was considered fundamental for the biosynthesis of l-aspartic acid in S. thermophilus metabolism. This enzymatic activity is therefore essential for growth of S. thermophilus in milk even if S. thermophilus was cultured in association with proteinase-positive Lactobacillus delbrueckii subsp. bulgaricus. It was furthermore observed that the supplementation of milk with aspartate significantly affected the level of urease activity. Further experiments, carried out with a p ureI -gusA recombinant strain, revealed that expression of the urease operon was sensitive to the aspartate concentration in milk and to the cell availability of glutamate, glutamine, and ammonium ions.

scholarly journals A buffered media system for yeast batch culture growth

2021 ◽  
Vol 21 (1) ◽  
Author(s):  
Rianne C. Prins ◽  
Sonja Billerbeck
Keyword(s):  
Nitrogen Source ◽  
Ammonium Sulfate ◽  
Batch Culture ◽  
Biomass Production ◽  
Buffering Capacity ◽  
Drop Out ◽  
Batch Cultures ◽  
Media System ◽  
The Media ◽  
Media Types

Abstract Background Fungi are premier hosts for the high-yield secretion of proteins for biomedical and industrial applications. The stability and activity of these secreted proteins is often dependent on the culture pH. As yeast acidifies the commonly used synthetic complete drop-out (SD) media that contains ammonium sulfate, the pH of the media needs to be buffered in order to maintain a desired extracellular pH during biomass production. At the same time, many buffering agents affect growth at the concentrations needed to support a stable pH. Although the standard for biotechnological research and development is shaken batch cultures or microtiter plate cultures that cannot be easily automatically pH-adjusted during growth, there is no comparative study that evaluates the buffering capacity and growth effects of different media types across pH-values in order to develop a pH-stable batch culture system. Results We systematically test the buffering capacity and growth effects of a citrate-phosphate buffer (CPB) from acidic to neutral pH across different media types. These media types differ in their nitrogen source (ammonium sulfate, urea or both). We find that the widely used synthetic drop-out media that uses ammonium sulfate as nitrogen source can only be effectively buffered at buffer concentrations that also affect growth. At lower concentrations, yeast biomass production still acidifies the media. When replacing the ammonium sulfate with urea, the media alkalizes. We then develop a medium combining ammonium sulfate and urea which can be buffered at low CPB concentrations that do not affect growth. In addition, we show that a buffer based on Tris/HCl is not effective in maintaining any of our media types at neutral pH even at relatively high concentrations. Conclusion Here we show that the buffering of yeast batch cultures is not straight-forward and addition of a buffering agent to set a desired starting pH does not guarantee pH-maintenance during growth. In response, we present a buffered media system based on an ammonium sulfate/urea medium that enables relatively stable pH-maintenance across a wide pH-range without affecting growth. This buffering system is useful for protein-secretion-screenings, antifungal activity assays, as well as for other pH-dependent basic biology or biotechnology projects.

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