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Influence of carbon source on alpha-amylase production by Aspergillus oryzae.

The influence of the carbon source on alpha-amylase production by Aspergillus oryzae was quantified in carbon-limited chemostat cultures. The following carbon sources were investigated: maltose, maltodextrin (different chain lengths), glucose, fructose, galactose, sucrose, glycerol, mannitol and acetate. A. oryzae did not grow on galactose as the sole carbon source, but galactose was co-metabolized together with glucose. Relative to that on low glucose concentration (below 10 mg/l), productivity was found to be higher during growth on maltose and maltodextrins, whereas it was lower during growth on sucrose, fructose, glycerol, mannitol and acetate. During growth on acetate there was no production of alpha-amylase, whereas addition of small amounts of glucose resulted in alpha-amylase production. A possible induction by alpha-methyl-D-glucoside during growth on glucose was also investigated, but this compound was not found to be a better inducer of a-amylase production than glucose. The results strongly indicate that besides acting as a repressor via the CreA protein, glucose acts as an inducer.

Aspergillus oryzae↗

Influence of ORP variation, carbon source and nitrate concentration on denitrifying phosphorus removal by DPB sludge from dephanox process.

The effect of added carbon source and nitrate concentration on the denitrifying phosphorus removal by denitrifying phosphorus removal bacteria sludge was systematically studied using batch experiments, at the same time the variation of ORP was investigated. Results showed that the denitrifying and phosphorus uptake rate in the anoxic phase increased with the high initial anaerobic carbon source addition. However, once the initial COD concentration reached a certain level, which was in excess of the PHB saturation of Poly-p bacteria, residual COD carried over to the anoxic phase inhibited the subsequent denitrifying phosphorus uptake. This was equal to supplementing the external carbon source to the anoxic phase, furthermore the higher the external carbon source concentration the more powerful the inhibition caused. High nitrate concentration in the anoxic phase increased the initial denitrifying phosphorus rate. Oncethe nitrate was exhausted, phosphate uptake changed to phosphate release. Moreover, the time of this turning point occurred later with the higher nitrate addition. On the other hand, through on-line monitoring the variation of the ORP with different initial COD concentration, it was found that ORP could be used as a control parameter for phosphorus release, but it is impossible to utilize ORP for controlling the dinitrification and anoxic phosphorus uptake operations.

Carbon↗

Denitrifying characteristics of the multiple stages enhanced biological nutrient removal process with external carbon sources.

This research investigated denitrifying activity of activated sludge with three external carbon sources (sodium acetate, methanol and glucose) via a series of batch experiments. Activated sludge used was cultivated in a multiple stages enhanced biological nutrient removal (EBNR) process that exhibited high removal efficiency of effective carbon, nitrogen, and phosphorus. Results showed type of external carbon source had a significant influence on specific nitrate utilization rate, nitrite accumulation, adaptive time of microorganisms, and nitrate removal efficiency. Sodium acetate addition resulted in high phosphate concentration in effluent; meanwhile methanol caused increasing turbidity and carbon breakthrough problem. When glucose was fed to be the external carbon source, accumulative nitrite concentration was higher than that with sodium acetate or methanol addition. When sodium acetate, methanol and glucose were used to be the electron donor, average dosages for nitrate elimination were 6.97, 5.85, and 5.65 mg-COD/mg-N, respectively. Because the final polyhydroxyalkanoates (PHAs) concentrations contained within the biomass were more than the original level and no phosphate re-release was observed, glycogen-accumulating organisms (GAOs) might exist in the multiple stages EBNR process and increased carbon dosage for further nitrate removal.

Biodegradation, Environmental↗

Enhanced post-denitrification without addition of an external carbon source in membrane bioreactors.

This study investigates a post-denitrification process without the addition of an external carbon source combined with an enhanced biological phosphorus removal (EBPR) in a membrane bioreactor (MBR). Three trial plants, with two different process configurations, were operated on two different sites, and a variety of accompanying batch tests were conducted. It was shown that even without dosing of an external carbon source, denitrification rates (DNR) much above endogenous rates could be obtained in post-denitrification systems. Furthermore, the anaerobic reactor located ahead of the process had a positive impact on the DNR. Given these surprising results, the project team decided to identify the carbon source used by the microorganisms in the post-denitrification process. Batch tests could demonstrate that lysis products do not play a major role as a C-source for post-denitrification. The following hypothesis was proposed to explain the observations: the glycogen, internally stored by the substrate accumulating bacteria, if anaerobic conditions are followed by aerobic conditions could act as carbon source for denitrification in post-denitrification system. First exploratory batch tests, where the glycogen evolution was monitored, corroborate this assumption.

Bacteria, Aerobic↗

Sporulation synchrony of Saccharomyces cerevisiae grown in various carbon sources.

Sporulation of several strains of Saccharomyces cerevisiae grown in a variety of carbon sources that do not repress the tricarboxylic acid cycle enzymes was more synchronous than the sporulation of cells grown in medium containing dextrose which does repress those enzymes. Dextrose-grown cells showed optimal sporulation synchrony when inoculated into sporulation medium from early stationary phase when the dextrose in the medium is exhausted. Logarithmic-phase cells grown in either non-fermentable carbon sources (acetate and glycerol) or a fermentable carbon source that does not repress tricarboxylic acid cycle enzymes (galactose) sporulated more synchronously than the early stationary-phase dextrose cells. Attempts were made to sporulate cells taken from both complex and semidefined media. The semidefined acetate medium failed to support the growth of a number of strains. However, cells grown in the complex acetate medium, as well as both complex and semidefined glycerol and galactose media, sporulated with better synchrony than did the dextrose-grown cells.

Acetates↗

Effect of carbon source on pyrimidine formation in Pseudomonas fluorescens ATCC 13525.

The effect of carbon source on the regulation of the de novo pyrimidine biosynthetic enzymes in the type strain of Pseudomonas fluorescens was investigated. The de novo pyrimidine biosynthetic enzyme activities were measured in P. fluorescens ATCC 13525 cells and in cells from an auxotroph deficient for orotate phosphoribosyltransferase activity where glucose or succinate served as the carbon source. Pyrimidine supplementation to the culture medium affected the biosynthetic enzyme activities in ATCC 13525 cells. Pyrimidine Limitation of the mutant strain indicated transcriptional regulation of the biosynthetic pathway by pyrimidines that was influenced by carbon source. Transcriptional regulation of pyrimidine synthesis in P. fluorescens appeared to be more highly regulated than was observed for the taxonomically related species Pseudomonas aeruginosa or Pseudomonas putida.

Aspartate Carbamoyltransferase↗

Mutagenesis by 9-aminoacridine in Salmonella typhimurium: inhibition by glucose and other PTS class A carbon sources.

Reversion of the hisC3076 frameshift marker of Salmonella typhimurium has been measured following treatment of cells in growth and non-growth media with 9-aminoacridine (9AA). By varying the carbon source present in a defined medium, it has been shown that mutagenesis is reduced close to the spontaneous level in the presence of glucose whilst significant reductions are also observed with glucosamine, mannose, mannitol, fructose or glucose 6-phosphate. Intermediate mutant yields are observed when lactic acid or glycerol are present, whereas any one of a further group of carbon sources (gluconate, arabinose, ribose, succinate or casein hydrolysate) permit relatively large numbers of mutants to be recovered. Interestingly, when any one of these "high yield" carbon sources is supplemented with glucose the strong inhibitory effect characteristic of glucose is again observed. On the basis of these results, it can be concluded that inhibition of 9AA-induced reversion by a carbon source is not an exclusive property of glucose, although when more than one carbon source is present the inhibitory effect of glucose predominates. Possible explanations for these findings include the active exclusion of 9AA from cells as a direct consequence of glucose transport across the cell membrane. To address this possibility, cells were pre-grown in verapamil, a calcium channel antagonist which is known to increase the mutagenicity of various 9-anilinoacridine derivatives in S. typhimurium. We found that glucose inhibition of 9AA-induced mutagenesis was not relaxed to any significant extent following treatment with verapamil. In a further experiment, two glucose analogues (2'-deoxyglucose and methyl-D-glucoside) known to be actively transported into the cell but not metabolised past the first phosphorylation step were used. These analogues inhibit the transport into the cell of several types of molecules, but since they do not significantly depress 9AA mutagenesis it seems unlikely that blockage of 9AA transport across the cell membrane can be invoked to explain the inhibitory effect of glucose on 9AA mutagenesis. An alternative explanation based on glucose-mediated repression of an error-prone, mutation-generating, DNA-repair process is presented.

Aminacrine↗

Effects of alternative carbon sources on biological transformation of nitrophenols.

The removal of nitrophenols under denitrifying conditions was studied in bench-scale upflow anaerobic sludge blanket (UASB) reactors (R1, R2, R3 and R4) using three different carbon sources. Initially acetate was used as carbon source (substrate) in all the four reactors followed by glucose and methanol. Reactor R1 was kept as control and R2, R3, R4 were fed with 30 mg/l concentration of 2-nitrophenol (2-NP), 4-nitrophenol (4-NP), and 2,4-dinitrophenol (2,4-DNP), respectively. Throughout the study the hydraulic retention time (HRT) and COD/NO3-N ratio were kept as 24 h and 10, respectively. 2-Aminophenol (2-AP), 4-aminophenol (4-AP) and 2-amino,4-nitrophenol (2-A,4-NP) were found as the major intermediate metabolites of 2-NP, 4-NP and 2,4-DNP degradation, respectively. Methanol was found to be a better carbon source for 4-NP and 2,4-DNP degradation as compared to acetate and glucose, while 2-NP degradation was not influenced much by the change of substrate. Nitrate nitrogen removal was always more than 99%. COD removal efficiency of the nitrophenol fed reactors varied from 85.7% to 97.7%. The oxidation-reduction potential (ORP) inside the reactors dropped, up to -300 mv, with glucose as carbon source. As the reactors were switched over to methanol, ORP increased to -190 mv. The granular sludge developed inside the reactors was light brown in colour when acetate and glucose were used as substrate, which turned dark brown to black at the end of methanol run. Biomass yield in terms of volatile suspended solids was observed as 0.15, 0.089 and 0.14 g per gram of COD removal for acetate, glucose and methanol, respectively.

Acetates↗

Carbon-source-dependent nitrogen regulation in Escherichia coli is mediated through glutamine-dependent GlnB signalling.

The P(II) signal transduction proteins GlnB and GlnK are uridylylated/deuridylylated in response to the intracellular glutamine level, the primary signal of the cellular nitrogen status. Furthermore, GlnB was shown to be allosterically regulated by 2-oxoglutarate, and thus GlnB was suggested to integrate signals of the cellular carbon and nitrogen status. Receptors of GlnB signal transduction in Escherichia coli are the NtrB/NtrC two-component system and GlnE, an enzyme which adenylylates/deadenylylates glutamine synthetase. In this study, the authors investigated the effect of different carbon sources on the expression of the NtrC-dependent genes glnA and glnK and on the uridylylation status of GlnB and GlnK. With glutamine as nitrogen source, high levels of glnA and glnK expression were obtained when glucose was used as carbon source, but expression was strongly decreased when the cells were grown with poor carbon sources or when cAMP was present. This response correlated with the uridylylation status of GlnB, suggesting that the carbon/cAMP effect was mediated through GlnB uridylylation, a conclusion that was confirmed by mutants of the P(II) signalling pathway. When glutamine was replaced by low concentrations of ammonium as nitrogen source, neither glnAglnK expression nor GlnB uridylylation responded to the carbon source or to cAMP. Furthermore, glutamine synthetase could be rapidly adenylylated in vivo by the external addition of glutamine; however, this occurred only when cells were grown in the presence of cAMP, not in its absence. Together, these results suggest that poor carbon sources, through cAMP signalling, favour glutamine uptake. The cellular glutamine signal is then transduced by uridylyltransferase and GlnB to modulate NtrC-dependent gene expression.

Bacterial Proteins↗

Carbon-source-dependent transcriptional control involved in the initiation of cellular differentiation in Streptomyces griseus.

Carbon source is one of the environmental factors that affects cellular differentiation of Streptomyces. We have identified the craA gene as a putative negative regulator involved in the carbon-source-dependent initiation of cellular differentiation in Streptomyces griseus. Carbon-source-dependent transcriptional repression of craA, which is caused by binding of a putative repressor protein to its promoter region, is proposed to result in the initiation of aerial mycelium formation. The presence of a craA homologue in the chromosome of Streptomyces coelicolor A3(2) implicates the existence of a similar regulatory mechanism in this organism. The repression of craA-promoter activity in glucose media could be alleviated not only by replacing glucose with maltose but also by supplying copper, which suggests that the stimulatory effect of copper on cellular differentiation in Streptomyces is excerted via abolishment of glucose-repression of craA.

Amino Acid Sequence↗

Effect of different carbon sources on lipase production by Candida rugosa.

Different carbon sources affecting growth and lipase production in Candida rugosa were studied by using batch cultures on defined medium. Carbohydrates and acids non-related to fats did not induce lipase production. The highest yields of enzyme were obtained with lipids or fatty acids as carbon sources. Tween 80 stimulated lipase biosynthesis and secretion outside the cell. Combinations of two types of substrates, carbohydrates and fatty acids, did not improve lipase production, and in some cases, their consumption was produced in a sequential pattern. Glucose presented a repressing effect on lipase production. Moreover, glucose was found to be effective in stimulating lipase secretion by cells with a high level of cell-bound lipase activity because of their previous growth in oleic acid.

Journal Article↗

Carbon source dependence of transposable element-associated gene activation in Saccharomyces cerevisiae.

Seven cis-dominant mutations leading to the overproduction of the glucose-repressible alcohol dehydrogenase isozyme ADHII (structural gene, ADH2) in Saccharomyces cerevisiae have previously been shown to be due to insertion of a transposable element, Ty, in the 5' regulatory region of the ADH2 gene. We showed that although mating-competent cells (a, alpha, a/a, or alpha/alpha cells) overproduced both ADHII enzyme and ADH2 mRNA, mating-incompetent cells (a/alpha or ste-cells) produced much less ADHII enzyme and ADH2 mRNA. This mating type effect on ADH2 expression was greatest in the presence of a normally derepressing carbon source, glycerol, and much less apparent in the presence of a repressing carbon source, glucose. In addition, Ty insertion led to an aberrant carbon source response in mating-incompetent cells--the normally glucose-repressible ADHII becomes glycerol repressible. The mating type effect and aberrant carbon source response in mating-incompetent cells was specific for Ty-associated mutations in the 5' flanking region of the ADH2 gene in that a non-Ty mutation in the same region did not show these effects. Finally, Ty1 RNA levels also showed a/alpha, suppression, which was apparent only during growth on a nonfermentable carbon source such as glycerol. This suggests that Ty-mediated gene expression is subject to regulation by both mating competence and carbon catabolites.

Alcohol Dehydrogenase↗

Contribution of different carbon sources to isoprene biosynthesis in poplar leaves.

This study was performed to test if alternative carbon sources besides recently photosynthetically fixed CO2 are used for isoprene formation in the leaves of young poplar (Populus x canescens) trees. In a 13CO2 atmosphere under steady state conditions, only about 75% of isoprene became 13C labeled within minutes. A considerable part of the unlabeled carbon may be derived from xylem transported carbohydrates, as may be shown by feeding leaves with [U-13C]Glc. As a consequence of this treatment approximately 8% to 10% of the carbon emitted as isoprene was 13C labeled. In order to identify further carbon sources, poplar leaves were depleted of leaf internal carbon pools and the carbon pools were refilled with 13C labeled carbon by exposure to 13CO2. Results from this treatment showed that about 30% of isoprene carbon became 13C labeled, clearly suggesting that, in addition to xylem transported carbon and CO2, leaf internal carbon pools, e.g. starch, are used for isoprene formation. This use was even increased when net assimilation was reduced, for example by abscisic acid application. The data provide clear evidence of a dynamic exchange of carbon between different cellular precursors for isoprene biosynthesis, and an increasing importance of these alternative carbon pools under conditions of limited photosynthesis. Feeding [1,2-13C]Glc and [3-13C]Glc to leaves via the xylem suggested that alternative carbon sources are probably derived from cytosolic pyruvate/phosphoenolpyruvate equivalents and incorporated into isoprene according to the predicted cleavage of the 3-C position of pyruvate during the initial step of the plastidic deoxyxylulose-5-phosphate pathway.

Biological Transport↗

Growth of Metarhizium anisopliae on non-preferred carbon sources yields conidia with increased UV-B tolerance.

Conidia of the insect-pathogenic fungus Metarhizium anisopliae var. anisopliae produced on different growth substrates (culture media or insect cadavers) demonstrate reproducibly altered tolerance to UV-B radiation [Rangel, D.E.N., Braga, G.U.L., Flint, S.D., Anderson, A.J., Roberts, D.W., 2004. Variations in UV-B tolerance and germination speed of M. anisopliae conidia produced on artificial and natural substrates. J. Invertebr. Pathol. 87, 77-83]. In the current study, the fungus was grown on potato dextrose agar with yeast extract (PDAY), on minimal medium [(MM)=Czapek medium without saccharose], or on MM with one of 16 different carbon sources. The conidia produced on these media were exposed to UV-B radiation. Great amplitude in phenotypic plasticity for UV-B tolerance was demonstrated, viz., conidia produced under nutritive stress [MM or MM supplemented with non-preferred carbon sources (e.g., fructose, galactose, lactose etc.)] had at least two times higher tolerance than conidia produced on the rich medium (PDAY). Endogenous trehalose and mannitol accumulated at least two times more in conidia produced on MM (or MM with lactose, a non-preferred carbon source), as compared to conidia from MM plus glucose. High accumulations of these two carbohydrates in fungal spores are known to protect them against a wide range of stresses. Sporulation, however, was most profuse on PDAY, second best on MM plus d-mannose and least on MM or MM containing non-preferred carbon sources. Taken together, the results illustrate that nutritive stress generated by MM or MM plus a non-preferred carbon source greatly improved UV-B tolerance, but reduced conidial yield; while, on the other hand, preferred carbon sources improved conidial yield, but reduced UV-B tolerance.

Animals↗

Dual influence of the carbon source and L-methionine on the synthesis of sulphur compounds in the cheese-ripening yeast Geotrichum candidum.

The effect of the carbon source and l-methionine on the ability of Geotrichum candidum to produce volatile sulphur compounds (VSC) was studied. This yeast was cultivated in a synthetic medium supplemented with various carbon sources and l-methionine at different concentrations. Both glycerol and glucose significantly increased VSC production by G. candidum. Unlike the effect on the l-methionine- and 4-methylthio-2-oxobutyric acid-demethiolating activities, the supply of a carbon source had a dramatic effect on the activity of aminotransferase, a key enzyme in l-methionine catabolism. An increase in the initial concentration of l-methionine resulted in a rise in the production of sulphur compounds (VSC, 4-methylthio-2-oxobutyric acid) but had limited effect on l-methionine-catabolising enzyme activities. Evidence for the existence of a dual effect of the carbon source and l-methionine on VSC biosynthesis was obtained in this study.

Butyrates↗

Changes in the protein expression of yeast as a function of carbon source.

Protein expression trends in yeast were monitored as a function of carbon source (glucose versus galactose) using multidimensional high performance liquid chromatography (HPLC) coupled to gas-phase fractionation, using relative intensity triggering (GPFri). Size exclusion HPLC was used to separate whole cell lysates, and following proteolysis of these fractions, each was separated by reversed phase HPLC, which was coupled on-line via electrospray to an ion trap mass spectrometer. The GPFri technique increased the dynamic range of proteins detected by increasing the number of peptide ions subjected to low energy collision induced dissociation to the 24 most intense ions in each of the survey scans. No protein or peptide labeling was used; instead, the number of SEQUEST identifications for each peptide (previously termed "hits") were used as a semiquantitative means of assessing both the direction (increase vs decrease) and significance of change in protein abundance. None of the traditional SEQUEST filters, e.g., Xcorr, DelCn, Sp, Rsp, etc., were employed in this study. Instead, a Student's t-test was used to distinguish those proteins that significantly and reproducibly changed between carbon sources from those that did not. This relied on the SEQUEST misassignments occurring in equal proportion between treatments and thereby negating each other; statistically significant changes in SEQUEST assignments were nonrandom events by definition and therefore reflective of correct identifications. This method of data analysis showed a large degree of concordance with results reported by other groups in similar transcriptional profiling and proteomic experiments. In all, 176 and 231 (fold-change > or = 1.1; p < or = 0.05) proteins were identified as being increased in peptide hit number when the yeast cells' source of carbon was changed between glucose and galactose, respectively.

Algorithms↗

Succinic acid production with reduced by-product formation in the fermentation of Anaerobiospirillum succiniciproducens using glycerol as a carbon source.

Succinic acid was produced by fermentation of Anaerobiospirillum succiniciproducens using glycerol as a carbon source. When cells were anaerobically cultured in a medium containing 6.5 g/L glycerol, a high succinic acid yield (133%) was obtained while avoiding the formation of by-product acetic acid. The gram ratio of succinic acid to acetic acid was 25.8:1, which is 6.5 times higher than that obtained using glucose (ca. 4:1) as a carbon source. Therefore, succinic acid can be produced with much less by-product formation by using glycerol as a carbon source, which will facilitate its purification. When glucose and glycerol were cofermented with the increasing ratio of glucose to glycerol, the gram ratio of succinic acid to acetic acid and succinic acid yield decreased, suggesting that glucose enhanced acetic acid formation irrespective of the presence of glycerol. Glycerol consumption by A. succiniciproducens required unidentified nutritional components present in yeast extract. By intermittently feeding yeast extract along with glycerol, a high succinic acid yield (160%) could be obtained while still avoiding acetic acid formation. This resulted in the highest ratio of succinic acid to acetic acid (31.7:1).

Acetic Acid↗

Biosurfactant production by Pseudomonas aeruginosa A41 using palm oil as carbon source.

Biosurfactant production by Pseudomonas aeruginosa A41, a strain isolated from seawater in the gulf of Thailand, was examined when grown in defined medium containing 2% vegetable oil or fatty acid as a carbon source in the presence of vitamins, trace elements and 0.4% NH(4)NO(3), at pH 7 and 30 degrees C with 200 rpm-shaking for 7 days. The yield of biosurfactant steadily increased even after a stationary phase. Under such conditions the surface tension of the medium was lowered from 55-70 mN/m to 27.8-30 mN/m with every carbon source tested. However, types of carbon sources were found to affect biosurfactant yield. The yields of rhamnolipid biosurfactant were 6.58 g/L, 2.91 g/L and 2.93 g/L determined as rhamnose content when olive oil, palm oil and coconut oil, respectively, were used as a carbon source. Among them, biosurfactant obtained from palm oil was the best in lowering surface tension of the medium. Increase in biosurfactant activities in terms of oil displacement test and rhamnose content were observed to be higher with shorter chain fatty acids than that of the longer chains (C12>C14>C16). In addition, we found that C18:2, highly unsaturated fatty acid, showed higher oil displacement activity and rhamnose content than that of C18:1. The optimal oil displacement activity was found at pH 7-9 and in the presence of 0.5-3% NaCl. The oil displacement activity was stable to temperatures up to 100 degrees C for 15 h. Surface tension reduction activity was relatively stable at pH 2-12 and 0-5% of NaCl. Emusification activity tested with various types of hydrocarbons and vegetable oils showed similarity of up to 60% stability. The partially purified biosurfactant via TLC and silica gel column chromatography gave three main peaks on HPLC with mass spectra of 527, 272, and 661 m/z respectively, corresponding to sodium-monorhamnodecanoate, hydroxyhexadecanoic acid and an unknown compound, respectively.

Glycolipids↗