• Journal of Microbiology and Biotechnology
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• v.18 no.12
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• pp.1958-1965
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• 2008

A denitrifying polyphosphate-accumulating bacterium (YKP-9) was isolated from activated sludge of a 5-stage biological nutrient removal process with step feed system. This organism was a Gram-negative, coccus-shaped, facultative aerobic chemoorganotroph. It had a respiratory type of metabolism with oxygen, nitrate, and nitrite as terminal electron acceptors. The 16S rRNA gene sequence of strain YKP-9 was most similar to the 16S rRNA gene sequence of Paracoccus sp. OL18 (AY312056) (similarity level, 97%). Denitrifying polyphosphate accumulation by strain YKP-9 was examined under anaerobic-anoxic and anaerobic-oxic batch conditions. It was able to use external carbon sources for polyhydroxyalkanoates(PHA) synthesis and to release phosphate under anaerobic condition. It accumulated polyphosphate and grew a little on energy provided by external carbon sources under anoxic condition, but did neither accumulate polyphosphate nor grow in the absence of external carbon sources under anoxic condition. Cells with intracellular PHA cannot accumulate polyphosphate in the absence of external carbon sources under anoxic condition. Under oxic condition, it grew but could not accumulate polyphosphate with external carbon sources. Based on the results from this study, strain YKP-9 is a new-type denitrifying polyphosphate-accumulating bacterium that accumulates polyphosphate only under anoxic condition, with nitrate and nitrite as the electron acceptors in the presence of external carbon sources.

• Journal of Microbiology and Biotechnology
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• v.34 no.2
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• pp.407-414
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• 2024

Phosphorus is an essential but non-renewable nutrient resource critical for agriculture. Luxury phosphorus uptake allows microalgae to synthesize polyphosphate and accumulate phosphorus, but, depending on the strain of algae, polyphosphate may be degraded within 4 hours of accumulation. We studied the recovery of phosphorus from wastewater through luxury uptake by an engineered strain of Synechocystis sp. with inhibited polyphosphate degradation and the effect of this engineered Synechocystis biomass on lettuce growth. First, a strain (∆phoU) lacking the phoU gene, which encodes a negative regulator of environmental phosphate concentrations, was generated to inhibit polyphosphate degradation in cells. Polyphosphate concentrations in the phoU knock-out strain were maintained for 24 h and then decreased slowly. In contrast, polyphosphate concentrations in the wild-type strain increased up to 4 h and then decreased rapidly. In addition, polyphosphate concentration in the phoU knockout strain cultured in semi-permeable membrane bioreactors with artificial wastewater medium was 2.5 times higher than that in the wild type and decreased to only 16% after 48 h. The biomass of lettuce treated with the phoU knockout strain (0.157 mg P/m²) was 38% higher than that of the lettuce treated with the control group. These results indicate that treating lettuce with this microalgal biomass can be beneficial to crop growth. These results suggest that the use of polyphosphate-accumulating microalgae as biofertilizers may alleviate the effects of a diminishing phosphorous supply. These findings can be used as a basis for additional genetic engineering to increase intracellular polyphosphate levels.

• Journal of Life Science
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• v.10 no.4
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• pp.397-402
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• 2000

Polyphosphate kinase catalyzes the formation of polyphosphate from ATP. To understand the mechanism of phosphate accumulation, the Serratia marcescens gene encoding ppk was cloned from the genomic library by the method of Southern hybridization. The hybridization positive DNA fragment region from pDH3 was subcloned into the expression vector. The ppk gene product, a polypeptide of 75 kDa, was confirmed by SDS-PAGE. Expression of the Serratia marcescens ppk is regulated by the catabolite repression system. The enzyme activity polyphosphate kinase was increased in the E. coli strain harboring plasmid pMH4 with ppk gene.

• Korean Journal of Microbiology
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• v.23 no.2
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• pp.90-100
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• 1985

The present study was designed to investigate cellular regulation of phosphate metabolism between catabolically repressed and derepressed states in yeast (Saccharomyces uvarum). The activities of various phospatases and the contents of phosphate compounds were detected according to the culture phase and various phosphate concentrations. As the results, Saccharomyces uvarum derepressed many phosphate metabolizing enzymes such as alkaline phosphatase, acid phosphatase and ATPase more than ten fold simultaneously during catabolic repression (phospgate and sugar starvation). At the same state, the amounts of orthophosphate, nucleotidic labile phosphate and acid soluble polypgosphate were increased, compared to basal levels of normally cultivated cells. $Mg^{++}-stimulated$ type among all phospatases was appeared to have most of the enzyme activity. It could be postulated that $K^+ -stimulated$ alkaline phosphatase was directly or indirectly correlated with the synthesis of acid insoluble polyphosphate $Mg^{++}-stimulated$ phosphatase with the degradation of polyphosphates. In case of cultivation in the medium supplemented with sugar and phosphate (catabolic derepression), phospgatase activities except for alkaline phosphatase were decreased rapidly through the progressive batch culture, After 12 hrs culture, at early exponential phase, the cellular accumulation of acid insoluble polyphosphate increased about 5 fold, compared to those of the starved cells. Under catabolic repression, it could be postulated that intracellular phosphate metabolism was regulated by derepressions of phosphatases. The function of polyphosphate system was shown to compensate the ATP/ADP system as phosphate donor and energy source especially during catabolic repression.

• Journal of Microbiology and Biotechnology
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• v.13 no.3
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• pp.385-393
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• 2003

Microbial communities were analyzed in an anaerobic/aerobic sequencing batch reactor (SBR) fed with glucose as a sole carbon source. Scanning electron microscopy (SEM) showed that tetrad or cuboidal packet bacteria dominated the microbial sludge. Quinone, slot hybridization, and 165 rRNA gene sequencing analyses showed that the Proteobacteria beta subclass and the Actinobacteria group were the main microbial species in the SBR sludge. However, according to transmission electron microscopy (TEM), the packet bacteria did not contain polyphosphate granules or glycogen inclusions, but only separate coccus-shaped bacteria contained these, suggesting that coccus-shaped bacteria accumulated polyphosphate directly and the packet bacteria played other role in the enhanced biological phosphorus removal (EBPR). Based on previous reports, the Actinobacteria group and the Proteobacteria beta subclass were very likely responsible for acid formation and polyphosphate accumulation, respectively, and their cooperation achieved the EBPR in the SBR operation which was supplied with glucose.

• The Korean Journal of Mycology
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• v.13 no.4
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• pp.235-241
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• 1985

The present study was designed to investigate biosynthetic patterns of polysaccharides under catabolic repression and derepression in Saccharomyces uvarum. Correlation coefficients between polysaccharide synthesis and polyphosphate accumulation were examined, according to the culture phase and under various phosphate concentrations (free, limited, sufficient). During catabolic derepression, biosynthesis of glycogen was enhanced. rapidly and highly in the cells grown on minimal medium, compared with those grown on the complete medium. Acid soluble glycogen type was the main component of total glycogen and alkali soluble glycogen was synthesized in small amount, after 24 hr culture, at the time of almost exhaustion of sugar in the medium. Total glycogen was accumulated highly in proportion to the amount of phosphate added to the medium. It could be postulated that type 'C' isoenzyme among ALPase was directly or indirectly correlated with the glucan synthesis. Mannan synthesis indicated maximal amount at the early exponential phase and stationary phase, and also acid soluble sugars at the stationary phase. Correlation coefficient between the mannan synthesis and poly-p-'C' accumulation, and also between mannan synthesis and phospholipid content indicated 0.866 and 0.726, respectively.

• The Korean Journal of Mycology
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• v.13 no.3
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• pp.141-148
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• 1985

To investigate cellular regulation of phosphate metabolism between catabolically repressed and derepressed states in Saccharomyces uvarum, the activities of polyphosphatases, the analysis of polyphosphate and cytochemical observation of volutin granules were examined according to the culture phase and under various phosphate concentrations. As the results, tripolyphosphatase activity was increased more than six-fold during catabolic repression as compared with those of catabolic derepression and the polyphosphatase activity increased at the time of maximal accumulation of acid insoluble polyphosphate 'B'. Of the low molecular weight polyphosphates, tripolyphosphate was mainly detected by thin layer chromatography. When the synthesis of volutin granules in derepressed cells was observed cytochemically, acid insoluble polyphosphate localizing at the cell wall was primarily synthesized and then transferred into the cytoplasm, nucleus and/or vacuole.

• Journal of Microbiology and Biotechnology
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• v.33 no.10
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• pp.1370-1375
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• 2023

In this study, we aimed to enhance the accumulation of chorismate (CHR) and anthranilate (ANT), key intermediates in the shikimate pathway, by modifying a shikimate over-producing recombinant strain of Corynebacterium glutamicum [19]. To achieve this, we utilized a CRISPR-driven genome engineering approach to compensate for the deletion of shikimate kinase (AroK) as well as ANT synthases (TrpEG) and ANT phosphoribosyltransferase (TrpD). In addition, we inhibited the CHR metabolic pathway to induce CHR accumulation. Further, to optimize the shikimate pathway, we overexpressed feedback inhibition-resistant Escherichia coli AroG and AroH genes, as well as C. glutamicum AroF and AroB genes. We also overexpressed QsuC and substituted shikimate dehydrogenase (AroE). In parallel, we optimized the carbon metabolism pathway by deleting the gntR family transcriptional regulator (IolR) and overexpressing polyphosphate/ATP-dependent glucokinase (PpgK) and glucose kinase (Glk). Moreover, acetate kinase (Ack) and phosphotransacetylase (Pta) were eliminated. Through our CRISPR-driven genome re-design approach, we successfully generated C. glutamicum cell factories capable of producing up to 0.48 g/l and 0.9 g/l of CHR and ANT in 1.3 ml miniature culture systems, respectively. These findings highlight the efficacy of our rational cell factory design strategy in C. glutamicum, which provides a robust platform technology for developing high-producing strains that synthesize valuable aromatic compounds, particularly those derived from the shikimate pathway metabolites.

• Applied Biological Chemistry
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• v.43 no.3
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• pp.163-168
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• 2000

To remove phosphate accumulated in the soil and water, Acinetobacter lwoffi PO8 possessing a high ability to accumulate phosphate was isolated from a active sludge. Bacterium was cultured in the liquid medium containing $150\;{\mu}g/mL$ of phosphate at $30^{\circ}C$ in different culture conditions to examine intracellular phosphate uptake. The initial pH in the range of $7.5{\sim}8.5$ was effective on the growth and phosphate uptake of the strain. Glycerol and arabinose used as a carbon sources showed 93 and 91% the phsphate uptake, respectively. Among the nitrogen sources, ammonium salt such as $NH_4NO_3$ and $(NH_4)_2SO_4$ was effectively utilized on the phosphate uptake compared with amino compounds. The rate of phosphate uptake of $NH_4NO_3$, and $(NH_4)_2SO_4$, was 95 and 96%, respectively The growth and Phosphate uptake ability in the strain were significantly promoted when metal ions were added in the medium; $Co^{2+}$, however, was not utilized by the strain. The capacity of phosphate uptake was enhanced to $10{\sim}20%$ when arginine, methionine, or lysine was added. Using $^{32}P$ to examine the uptake Pattern of intracellular phosphate, experiment result showed that polyphosphate was largely found in the fraction of intracellular inorganic phosphate of Acinetobacter lwoffi PO8.

• The Journal of Natural Sciences
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• v.11 no.1
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• pp.55-63
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• 1999

In this study, the stabilizer, PH and lysis method for optimum condition of S. uvarum protoplast formation were investigated, and also enzyme activity and poly-P formation of intact cell and protoplast mere compared. Upon protoplast formation, incubation time of 5 hours in snail gut enzyme and 3 hours in drisielase were reignited. 0.8 Mole mannitol and 6 mole KCl were apt to protoplast formation. Protoplast was contained less 22-27 percentage in ALPase, 4-15 percentage in ACPase than intact cell. Accumulation of inorganic polyphosphate did not increase significently in protoplast compared with intact cell.