• 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.

• KOREAN JOURNAL OF CROP SCIENCE
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• v.65 no.2
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• pp.113-123
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• 2020

Global climatic change and increasing climatic instability threaten crop productivity. Due to climatic change, drought stress is occurring more frequently in crop fields. In this study, we investigated the effect of treatment with hydrogen peroxide (H₂O₂) before leaf development on the growth and yield of sorghum for minimizing the damage of crops to drought. To assess the effect of H₂O₂ on the growth of sorghum plant, 10 mM H₂O₂ was used to treat sorghum leaves at the 3-leaf stage during growth in field conditions. Plant height, stem diameter, leaf length, and leaf width were increased by 7.6%, 9.6%, 8.3% and 11.5%, respectively. SPAD value, chlorophyll fluorescence (Fv/Fm), photosynthetic rate, stomatal conductance, and transpiration rate were increased by 3.0%, 4.9%, 26.0%, 23.4% and 12.7%, respectively. The amount of H₂O₂ in the leaf tissue of sorghum plant treated with 10 mM H₂O₂ was 0.7% of the applied amount after 1 hour. The level increased to approximately 1.0% after 6 hours. The highest antioxidant activity measured by the Oxygen Radical Absorbance Capacity assay was 847.3 µmol·g^-1 at 6 hour after treatment. However, in the well-watered condition, the concentration of H₂O₂ in the plant treated by the foliar application of H₂O₂ was 227.8 µmol·g^-1 higher than that of the untreated control. H₂O₂ treatment improved all the yield components and yield-related factors. Panicle length, plant dry weight, panicle weight, seed weight per plant, seed weight per unit area, and thousand seed weight were increased by 8.8%, 18.0%, 24.4%, 24.7%, 29.9% and 7.1%, respectively. Proteomic analysis showed that H₂O₂ treatment in sorghum increased the tolerance to drought stress and maintained growth and yield by ameliorating oxidative stress.

• BMB Reports
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• v.38 no.5
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• pp.584-590
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• 2005

Glucose 6-phosphate dehydrogenase (EC 1.1.1.49) was purified from Aspergillus aculeatus, a filamentous fungus previously isolated from infected tongue of a patient. The enzyme, apparently homogeneous, had a specific activity of $220\;units\;mg^{-1}$/, a molecular weight of $105,000{\pm}5,000$ Dal by gel filtration and subunit size of $52,000{\pm}1,100$ Dal by sodium dodecyl sulphate-polyacrylamide gel electrophoresis. The substrate specificity was extremely strict, with glucose 6-phosphate (G6P) being oxidized by nicotinamide adenine dinucleotide phosphate (NADP) only. At assay pH of 7.5, the enzyme had $K_m$ values of $6\;{\mu}m$ and $75\;{\mu}m$ for NADP and G6P respectively. The $k_{cat}$ was $83\;s^{-1}$. Steady-state kinetics at pH 7.5 produced converging linear Lineweaver-Burk plots as expected for ternary-complex mechanism. The patterns of product and dead-end inhibition suggested that the enzyme can bind NADP and G6P separately to form a binary complex, indicating a random-order mechanism. The enzyme was irreversibly inactivated by heat in a linear fashion, with G6P providing a degree of protection. Phosphoenolpyruvate (PEP), adenosinetriphosphate (ATP), and fructose 6-phosphate (F6P), in decreasing order, are effective inhibitors. Zinc and Cobalt ions were effective inhibitors although cobalt ion was more potent; the two divalent metals were competitive inhibitors with respect to G6P, with $K_i$ values of $6.6\;{\mu}m$ and $4.7\;{\mu}m$ respectively. It is proposed that inhibition by divalent metal ions, at low NADPH /NADP ratio, is another means of controlling pentosephosphate pathway.

• Korean Journal of Clinical Laboratory Science
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• v.44 no.3
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• pp.103-111
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• 2012

Low-Density Lipoprotein cholesterol (LDLC) is the most important marker for the treatment of hyperlipidemia in NCEP-ATP III(National Cholesterol Education Program-Adult Treatment Panel III) guideline. Therefore, LDL cholesterol is pathologically meaningful, accurate measurement should be a top priority. Currently, LDLC is directly measured in most cases, but, the estimate is still used in mass health examination or screening test. This study is about the comparison of LDL-Cholesterol direct measurement with the estimate using various formula (Friedewald: [LDL-F=TC-HDL-TG/5], Nakajima: [LDL-N=TC-HDL-TG/4], Hattori: [LDL-H =0.94TC-0.94HDL-0.19TG], Puavilai: [LDL-P=TC-HDL-TG/6], Carvalho: [LDL-C=3(TC-HDL)/4]) for calculating more accurate value. We analyzed total cholesterol (TC), try-glyceride (TG), high-density lipoprotein cholesterol (HDLC), and LDLC levels of 210 subjects between June and November in 2011. Until now, the Friedewald formula is the most commonly used estimate for the LDLC. When Friedewald formula was applied, the correlation coefficient (r) was 0.940, showing high correlation. But, the result of the direct method was significantly different, compared with those of the Friedewald formula in triglyceride levels ${\geq}400mg/dL$(p<0.05). There was the highest correlation when we used LDL-P formula(r=0.947) in triglyceride levels <400 mg/dl. Also there was the lowest mean difference regardless of triglyceride level. Therefore, the study showed that TG/6 is more precise means of calculation than TG/5. On the other hand, the calculation of LDL-Cholesterol was underestimated, compared with direct measurement. It is necessary to have more data and modified Friedewald formula should be used for the accurate calculation.

• Journal of Life Science
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• v.16 no.1
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• pp.131-135
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• 2006

Oligopeptide is known to be an essential nitrogen nutrient for bacterial growth. Oligopeptide can be transported into cytoplasm by a specific transport system, Opp system. Opp system is composed of five proteins, which are transcribed by an operon. These are responsible for oligopeptide binding protein (OppA), permease (OppB and OppC) and energy generation system (OppD and OppF), respectively. Previously, we isolated the opp operon from Vibrio fluvialis and constructed the oppA mutant by allelic exchange method. In this study, we investigated the growth pattern and biofilm production under the different growth condition. When the cells were cultivated using brain heart infusion(BHI) medium, the wild type was faster than the mutant in growth during the exponential phase. However, it showed that the growth pattern of two strains in M9 medium is very similar. The growth of wild type showed better than that of the mutant grown at pH 8. At pH 7, there was no an obvious difference in growth. After 5 mM $H_2O_2$ was treated to the cells $(OD_{600}=1.2)$, the cell survival was examined. The oppA mutation did not affect in survivability. In the presence of $10{\mu}g/ml$ polymyxin B, the biofilm production of the oppA mutant was higher than that of the wild type.

• Korean Journal of Soil Science and Fertilizer
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• v.2 no.1
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• pp.53-68
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• 1969

The nutriophysiological response of rice plant to root environment was investigated with eye observation of root development and rhizosphere in situation. The results may be summarized as follows: 1) The quick decomposition of organic matter, added in low yield soil, caused that the origainal organic matter content was reached very quickly, in spite of it low value. In high yield soil the reverse was seen. 2) In low yield soil root development, root activity and T/R value were very low, whereas addition of organic matter lowered them still wore. This might be contributed to gas bubbles around the root by the decomposition of organic matter. 3) Varietal difference in the response to root environment was clear. Suwon 82 was more susceptible to growth-inhibitine conditions on low-yield soil than Norin 25. 4) Potassium uptake was mostly hindered by organic matter, while some factors in soil hindered mostly posphorus uptake. When the organic matter was added to such soil, the effect of them resulted in multiple interaction. 5) The root activity showed a correlation coeffieient of 0.839, 0.834 and 0.948 at 1% level with the number of root, yield of aerial part and root yield, respectively. At 5% level the root-activity showed correlation-coefficient of 0.751, 0.670 and 0.769 with the uptake of the aerial part of respectively. N, P and K and a correlation-coefficient of 0.729, 0.742 and 0.815 with the uptake of the root of respectively N.P. and K. So especially for K-uptake a high correlation with the root-activity was found. 6) The nitrogen content of the roots in low-yield soil was higher than in high-yield soil, while the content in the upper part showed the reverse. It may suggest ammonium toxicity in the root. In low-yield soil Potassium and Phosphorus content was low in both the root and aerial part, and in the latter particularly in the culm and leaf sheath. 7) The content of reducing sugar, non-recuding sugar, starh and eugar, total carbohydrates in the aerial part of plants in low yield soil was higher than in high yield soil. The content of them, especially of reducing sugar in the roots was lower. It may be caused by abnormal metabolic consumption of sugar in the root. 8) Sulfur content was very high in the aerial part, especially in leaf blade of plants on low yield soil and $P_2O_5/S$ value of the leaf blade was one fifth of that in high yield soil. It suggests a possible toxic effect of sulfate ion on photophosphorization. 9) The high value of $Fe/P_2O_5$ of the aerial part of plants in low yield soil suggests the possible formation of solid $Fe/PO_4$ as a mechanical hindrance for the translocation of nutrients. 10) Translocation of nutrients in the plant was very poor and most nutrients were accumulated in the root in low yield soil. That might contributed to the lack of energy sources and mechanical hindrance. 11) The amount of roots in high yield soil, was greater than that in low yield soil. The in high-yield soil was deep, distribution of the roots whereas in the low-yield soil the root-distribution was mainly in the top-layer. Without application of Nitrogen fertilizer the roots were mainly distributed in the upper 7cm. of topsoil. With 120 kg N/ha. root were more concentrated in the layer between 7cm. and 14cm. depth. The amount of roots increased with the amount of fertilizer applied.