• KSBB Journal
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• v.14 no.3
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• pp.384-388
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• 1999

We have used high pH anion-exchange chromatography to analyze low level (below $20{\mu}M$) $\alpha$-D-glucose-1-phosphate (G-1-P) that can be used as a cytostatic compound, an antibiotic, and immunosuppressive drug. Our chromatographic method afforded excellent peak resolution and seletivity for glucose-6-phosphate and various maltooligosaccharides as well as G-1-P. The pulsed amperometric detector yielded linear response on G-1-P ranging from 2 - $20{\mu}M$, giving slope of $4.8{\times}10^4$(peak area/${\mu}M$). The detection limit was $2{\mu}M$. This method was applied to the purification of thermophilic $\alpha$-glucan phosphorylase from Thermus caldophilus. The technique will be extremely useful in future studies concerning carbohydrate metabolism in living organisms.

• The Korean Journal of Pharmacology
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• v.26 no.2
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• pp.121-126
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• 1990

The contractility of isolated rat atria, suspended in Krebs-Ringer bicarbonate medium containing 5.5mM glucose, was depressed approximately 50% by 0.1 mM of lidocaine. Partial recovery of the lidocaine-depressed contractility was achieved by the metabolizable substrates pyruvate, acetate, and fructose, but not by addition of glucose. Glucose produced the dose-dependent increase in the force of contraction of normal atria, whereas pyruvate, acetate, and fructose produced no significant effect in the contractile activity of the normal atria. In the absence of exogenous glucose lidocaine produced more marked depression of atrial contractility than that in the presence of exogenous glucose. The results of this study may confirm that the utilization of cardiac glycogen is also inhibited by lidocaine at sites of the glucose phosphate isomerase step or step between glycogen to glucose-6-phosphate.

• BMB Reports
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• v.30 no.2
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• pp.157-161
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• 1997

The present paper reports characteristics and specificity of the inhibitory action of $N^{\alpha}-tosyl-L-lysine-chloromethyl\;ketone$ (TLCK) and $N^{\alpha}-tosyl-L-phenylalanine-chloromethyl\;ketone$ (TPCK) on the glucose6-phosphate transporter of rat liver microsomes. The TLCK-induced inhibition was pH dependent. The inhibition constants for TPCK were determined by following pseudo-Lst order reaction mechanism. The inhibition was protected by preincubation with excess amount of glucose-6-phosphate. The results proved that (a) TLCK inactivates the microsomal glucose-6-phosphate transporter, (b) the inhibition results from the modification of sulfhydryl groups of the transporter.

• Journal of Microbiology
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• v.42 no.1
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• pp.20-24
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• 2004

Cloned myo-inositol-1-phosphate synthase (INOS) of Drosophila melanogaster was expressed in Escherichia coli, and purified using a His-affinity column. The purified INOS required NAD$\^$+/ for the conversion of glucose-6-phosphate to inositol-1-phosphate. The optimum pH for myo-inositol-1-phosphate synthase is 7.5, and the maximum activity was measured at 40$^{\circ}C$. The molecular weight of the native enzyme, as determined by gel filtration, was approximately M$\_$r/ 271,000${\pm}$15,000. A single subunit of approximately M$\_$r/ 62,000${\pm}$5,000 was detected upon SDS-polyacrylamide gel electrophoresis. The Michaelis ($K_{m}$) and dissociation constants for glucose-6-phosphate were 3.5 and 3.7 mM, whereas for the cofactor NAD$\^$+/ these were 0.42 and 0.4 mM, respectively.

• Journal of Microbiology and Biotechnology
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• v.29 no.4
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• pp.577-586
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• 2019

The engineered Aspergillus oryzae has a high NADPH demand for xylose utilization and overproduction of target metabolites. Glucose-6-phosphate dehydrogenase (G6PDH, E.C. 1.1.1.49) is one of two key enzymes in the oxidative part of the pentose phosphate pathway, and is also the main enzyme involved in NADPH regeneration. The open reading frame and cDNA of the putative A. oryzae G6PDH (AoG6PDH) were obtained, followed by heterogeneous expression in Escherichia coli and purification as a his6-tagged protein. The purified protein was characterized to be in possession of G6PDH activity with a molecular mass of 118.0 kDa. The enzyme displayed maximal activity at pH 7.5 and the optimal temperature was $50^{\circ}C$. This enzyme also had a half-life of 33.3 min at $40^{\circ}C$. Kinetics assay showed that AoG6PDH was strictly dependent on $NADP^+$ ($K_m=6.3{\mu}M$, $k_{cat}=1000.0s^{-1}$, $k_{cat}/K_m=158.7s^{-1}{\cdot}{\mu}M^{-1}$) as cofactor. The $K_m$ and $k_{cat}/K_m$ values of glucose-6-phosphate were $109.7s^{-1}{\cdot}{\mu}M^{-1}$ and $9.1s^{-1}{\cdot}{\mu}M^{-1}$ respectively. Initial velocity and product inhibition analyses indicated the catalytic reaction followed a two-substrate, steady-state, ordered BiBi mechanism, where $NADP^+$ was the first substrate bound to the enzyme and NADPH was the second product released from the catalytic complex. The established kinetic model could be applied in further regulation of the pentose phosphate pathway and NADPH regeneration of A. oryzae to improve its xylose utilization and yields of valued metabolites.

• BMB Reports
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• v.32 no.4
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• pp.363-369
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• 1999

The orf8(chlD) gene cloned from Streptomyces antibioticus T$\"{u}$99 was overexpressed using an E. coli system to confirm its biological function. Induction of the E. coli strain transformed with recombinant plasmid pRFJ 1031 containing orf8 resulted in the production of a 43,000 dalton protein. Glucose-1-phosphate thymidylyltransferase activity of the cell extract obtained from the transformed strain was 4-5 times higher than that of the control strain. The expressed protein was purified 18-fold from E. coli cell lysate using three chromatographic steps with a 17% overall recovery to near homogeneity. The N-terminal amino acid sequence of the purified protein agrees with the nucleotide sequence predicted from the orf8 gene. The SDS-PAGE estimated subunit mass of 43,000 dalton agrees well with that calculated from the amino acid composition deduced from the nucleotide sequence of the orf8 gene (43,000 Da). Also, the native enzyme has a monomeric structure with a molecular mass of 43,000 dalton. The purified protein showed glucose-1-phosphate thymidylyltransferase activity catalyzing a reversible bimolecular group transfer reaction, and was highly specific for dTTP and ${\alpha}$-D-glucose 1-phosphate as substrates in the forward reaction, and for dTDP-D-glucose and pyrophosphate in the reverse reaction.

• Environmental Analysis Health and Toxicology
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• v.18 no.4
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• pp.305-310
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• 2003

A rapid, inexpensive enzymatic method is proposed for indirect water quality testing in terms of heavy metal toxicity. The activity of glucose-6-phosphate dehydrogenase was applied for heavy metal toxicity test as an effective criterion in water quality. The toxicity of Pb (lead) and Cd (cadmium) for water flea, Moina macrocopa, were evaluated for 2-8 days with variables of mobilization ability. And the reproduction impairment of Moina macrocopa were investigated as the parameter of chronic toxicity test for Pb and Cd. As a result, the EC$_{50}$ for immobilization of Moina macrocopa were Pb and Cd were 1.6749 and 0.4683, respectively. The values of reproductive impairment to Moina macrocopa for Pb and Cd were 9.5938 and 8.3264 in EC$_{50}$ A significant alteration of G6PDH (Glucose-6-phosphate dehydrogenase) activity of Moina macrocopa was observed when Cd and Pb were treated in media. The results obtained indicate that G6PDH activity of Moina macrocopa can be used as an indicative parameter in aquatic toxicity tests for heavy metals.als.

• KSBB Journal
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• v.4 no.3
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• pp.259-265
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• 1989

In a batch fermentation process using carrot cell suspension culture, the effect of initial concentration of limiting nutrients(glucose and phosphate) on the specific growth rate and cell yield was investigated. The period of exponential growth is about 2 days and the consumption of glucose and phosphate in culture medium was very small when the initial concentrations of glucose and phosphate are 1.49g/1 ~ 3.01g/l and 0.08 ~ 0.32mM respectively. The specific growth rate of cells ranged from TEX>$0.15\;day^{-1}$ to $0.3\;day^{-1}$ irregularly. And the ratio of the initial concentration of glucose to phosphate did not affect the specific growth rate and the cell yield. The increase on cells had linear relationship with the consumption of limiting nutrients. Therefore, the increase of cells was found to be more influenced by the concentration of glucose than that of phosphate.

• The Korean Journal of Zoology
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• v.17 no.1
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• pp.17-22
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• 1974

The G-6-PD (glucose-6-phosphate dehydrogenase) activity, the first step in the pentose phosphate shunt, of rat livers during prenatal and pstnatal development in different sexes was studied. The enzyme activity is very high (54.2 units) at 16 days of embryo ad then decreases to a low level (17.6 units) at 13days after birth. There are significant increase between 13 and 15 days of age and continuously increases to the level of 53.2 units at 19days of age. The G-6-PD activity in female rat livers was slightly higher than in males.

• Journal of Microbiology and Biotechnology
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• v.27 no.4
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• pp.844-855
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• 2017

Phosphate-solubilizing bacteria (PSB) have the ability to dissolve insoluble phosphate and enhance soil fertility. However, the growth and mineral phosphate solubilization of PSB could be affected by exogenous soluble phosphate and the mechanism has not been fully understood. In the present study, the growth and mineral phosphate-solubilizing characteristics of PSB strain Burkholderia multivorans WS-FJ9 were investigated at six levels of exogenous soluble phosphate (0, 0.5, 1, 5, 10, and 20 mM). The WS-FJ9 strain showed better growth at high levels of soluble phosphate. The phosphate-solubilizing activity of WS-FJ9 was reduced as the soluble phosphate concentration increased, as well as the production of pyruvic acid. Transcriptome profiling of WS-FJ9 at three levels of exogenous soluble phosphate (0, 5, and 20 mM) identified 446 differentially expressed genes, among which 44 genes were continuously up-regulated when soluble phosphate concentration was increased and 81 genes were continuously down-regulated. Some genes related to cell growth were continuously up-regulated, which would account for the better growth of WS-FJ9 at high levels of soluble phosphate. Genes involved in glucose metabolism, including glycerate kinase, 2-oxoglutarate dehydrogenase, and sugar ABC-type transporter, were continuously down-regulated, which indicates that metabolic channeling of glucose towards the phosphorylative pathway was negatively regulated by soluble phosphate. These findings represent an important first step in understanding the molecular mechanisms of soluble phosphate effects on the growth and mineral phosphate solubilization of PSB.