34 results on '"Kyung-Hwan Jung"'
Search Results
2. Continuous Synthesis of Chlorphenesin Galactoside
- Author
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Kyung-Hwan Jung
- Subjects
chemistry.chemical_compound ,chemistry ,Biochemistry ,medicine ,medicine.disease_cause ,Galactoside ,Escherichia coli - Published
- 2019
3. Enzymatic Synthesis of 1, 2-Hexanediol Galactoside by Whole Cells of β-Galactosidase-containing Recombinant Escherichia coli
- Author
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Kyung-Hwan Jung and Yi-Ok Kim
- Subjects
0106 biological sciences ,Electrospray ,Chromatography ,Silica gel ,Sodium ,chemistry.chemical_element ,medicine.disease_cause ,01 natural sciences ,Galactoside ,0104 chemical sciences ,Adduct ,010404 medicinal & biomolecular chemistry ,Hydrolysis ,chemistry.chemical_compound ,chemistry ,Biochemistry ,010608 biotechnology ,Galactose ,medicine ,Escherichia coli - Abstract
side (HD-G) by a transgalactosylation reaction using β-galactosidase (β-gal)-containing recombinant Escherichia coli cells. The transgalactosylation reaction was carried out under high-lactose conditions for 24 hr. After 12 hr had elapsed, a new spot was identified by thin-layer chromatography (TLC) analysis, and we presumptively designated this new spot as HD-G. The n, we carried out the purification of the presumptive HD-G spot from the reaction mixture by using silica gel chromatography, and its mass was measured by electrospray ionization-mass spectrometry. The purified new spot on the chromatograph was identified a sodium adduct ion ([M+Na] + , m/z = 303.1423) of HD-G. In addition, when purified HD-G was hydrolyzed using commercially available E. coli β-gal, it was observed that a galactose molecule was released from HD-G. That is, it was demonstrated that HD-G is a galactoside derivative of HD. Finally, we confirmed that HD-G was enzymatically synthesized by E. coli β -gal as a new molecular entity. In the future, we plan to determine the minimum inhibitory concentrations of HD-G against different bacterial species. The cytotoxicity of HD-G against human skin cells will also be examined. It is expected hopefully that the galactosylation of HD would improve the functionality of HD-G.
- Published
- 2016
4. Ethanol Production from Glycerol using Pachysolen tannophilus in a Surface-aerated Fermentor
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Woon-Yong Choi, Kyung-Hwan Jung, Do-Hyung Kang, Yi-Ok Kim, and Hyeon-Yong Lee
- Subjects
chemistry.chemical_compound ,Ethanol ,chemistry ,Biochemistry ,Glycerol ,Yeast extract ,Ethanol fuel ,Fermentation ,Food science ,Aeration ,Yeast ,Fed-batch culture - Abstract
We investigated ethanol production from glycerol after screening of the yeast Pachysolen tannophilus ATCC 32691. For yeast to produce ethanol form glycerol, it is important that aeration is finely controlled. Therefore, we attempted to produce ethanol using a surface-aerated fermentor. When 880 ml of YPG medium (1% yeast extract, 2% peptone, 2% glycerol) was used to produce ethanol, the optimal aeration conditions for ethanol production were a surface aeration rate and agitation speed of 500 ml/min and 300 rpm, respectively. In a fed-batch culture, the maximum ethanol production and the maximum ethanol yield from glycerol (Ye/g) was 5.74 g/l and 0.166, respectively, after 90 hr using the surface-aerated fermentor.
- Published
- 2013
5. Long-term Repeated-Batch Operation of Immobilized Escherichia coli Cells to Synthesize Galactooligosaccharide
- Author
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Kyung Hwan Jung, Sang-Eun Lee, and Ji Hyeon Yeon
- Subjects
Inclusion Bodies ,Chromatography ,Galactooligosaccharide ,Escherichia coli Proteins ,Thermophile ,Galactose ,Oligosaccharides ,General Medicine ,Cells, Immobilized ,beta-Galactosidase ,medicine.disease_cause ,Applied Microbiology and Biotechnology ,chemistry.chemical_compound ,Bioreactors ,chemistry ,Biochemistry ,Reactor system ,Microbial enzymes ,Escherichia coli ,medicine ,Glutaraldehyde ,Batch operation ,Operational stability ,Biotechnology - Abstract
In this study, we investigated whether galactooligosaccharide (GOS) can be stably and steadily synthesized using immobilized beta-galactosidase (β-gal) inclusion body (IB)- containing E. coli cells during long-term repeated-batch operation. To improve the operational stability of this enzyme reactor system, immobilized E. coli cells were crosslinked with glutaraldehyde (GA) after immobilization of the E. coli. When we treated with 2% GA for E. coli crosslinking, GOS production continued to an elapsed time of 576 h, in which seven batch runs were operated consecutively. GOS production ranged from 51.6 to 78.5 g/l (71.2 ± 10.5 g/l, n = 7) during those batch operations. In contrast, when we crosslinked E. coli with 4% GA, GOS production ranged from 31.5 to 64.0 g/l (52.3 ± 10.8, n = 4), and only four consecutive batch runs were operated. Although we did not use an industrial β-gal for GOS production, in which a thermophile is used routinely, this represents the longest operation time for GOS production using E. coli β-gal. Improved stability and durability of the cell immobilization system were achieved using the crosslinking protocol. This strategy could be directly applied to other microbial enzyme reactor systems using cell immobilization to extend the operation time and/or improve the reactor system stability.
- Published
- 2012
6. Immobilization of Yeast Pichia stipitis for Ethanol Production
- Author
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Ji-Eun Lee, Sang-Eun Lee, Kyung-Hwan Jung, Do-Hyung Kang, Eunjin Kim, Joon Ho Choi, Woon Yong Choi, and Hyeon-Yong Lee
- Subjects
Chromatography ,Ethanol ,Materials science ,biology ,Lignocellulosic biomass ,Xylose ,biology.organism_classification ,Hydrolysate ,Yeast ,chemistry.chemical_compound ,chemistry ,Biochemistry ,Biofuel ,Ethanol fuel ,Pichia stipitis - Abstract
In this study, DEAE-cotton [derivatized by 2-(diethylamino)ethyl chloride hydrochloride (DEAE·HCl)] was prepared as a carrier for immobilized Pichia stipitis for ethanol production. When cotton was derivatized with 0.5 M DEAE·HCl, the yeast cell suspension was adsorbed at 100% of the initial cell OD600. The adsorbed yeast cells were estimated to be 101.8 mg-dry cells/g-DEAE-cotton. In particular, when a flask culture using the immobilized yeast cells was conducted in a glucose and xylose-containing medium, the yeast cells on the DEAE-cotton gradually produced ethanol, according to glucose and xylose consumption; the ethanol yield was approximately 0.33 g-ethanol/g-monosaccharide. Because DEAE-cotton was successfully used as a carrier for ethanol production from a glucose and xylose-containing medium, we expect that this bioethanol production process may be used for the bioethanol production process from the hydrolysate of lignocellulosic biomass. All the results of DEAE-cotton were compared with those of DEAE-cellulose as a carrier for immobilization.
- Published
- 2012
7. Galactooligosaccharide Synthesis by Active β-Galactosidase Inclusion Bodies-Containing Escherichia coli Cells
- Author
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Hye Ji Kim, Kyung-Hwan Jung, Hyeon-Beom Seo, Sang-Eun Lee, and Ji-Hyeon Yeon
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Oligosaccharides ,Lactose ,medicine.disease_cause ,Applied Microbiology and Biotechnology ,Hydrolysate ,chemistry.chemical_compound ,Escherichia coli ,medicine ,Beta-galactosidase ,Trisaccharide ,Chromatography, High Pressure Liquid ,Inclusion Bodies ,chemistry.chemical_classification ,Chromatography ,biology ,Galactooligosaccharide ,Temperature ,General Medicine ,Hydrogen-Ion Concentration ,beta-Galactosidase ,Culture Media ,Enzyme ,chemistry ,Biochemistry ,Spectrometry, Mass, Matrix-Assisted Laser Desorption-Ionization ,Galactose ,biology.protein ,Biotechnology - Abstract
In this study, galactooligosaccharide (GOS) was synthesized using active β-galactosidase (beta-gal) inclusion bodies (IBs)- containing Escherichia coli (E. coli) cells. Analysis by MALDI-TOF (matrix-assisted laser desorption/ionizationtime of flight) mass spectrometry revealed that a trisaccharide was the major constituent of the synthesized GOS mixture. Additionally, the optimal pH, lactose concentration, amounts of E. coli β-gal IBs, and temperature for GOS synthesis were 7.5, 500 g/l, 3.2 U/ml, and 37 °C, respectively. The total GOS yield from 500 g/l of lactose under these optimal conditions was about 32%, which corresponded to 160.4 g/l of GOS. Western blot analyses revealed that β-gal IBs were gradually destroyed during the reaction. In addition, when both the reaction mixture and E. coli β-gal hydrolysate were analyzed by high-performance thin-layer chromatography (HP-TLC), the trisaccharide was determined to be galactosyl lactose, indicating that a galactose moiety was most likely transferred to a lactose molecule during GOS synthesis. This GOS synthesis system might be useful for the synthesis of galactosylated drugs, which have recently received significant attention owing to the ability of the galactose molecules to improve the drugs solubility while decreasing their toxicity. β-Gal IB utilization is potentially a more convenient and economic approach to enzymatic GOS synthesis, since no enzyme purification steps after the transgalactosylation reaction would be required.
- Published
- 2011
8. Repeated-Batch Operation of Immobilized β-Galactosidase Inclusion Bodies-Containing Escherichia coli Cell Reactor for Lactose Hydrolysis
- Author
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Ji Hyeon Yeon and Kyung Hwan Jung
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Tris ,Cell ,Lactose ,medicine.disease_cause ,Applied Microbiology and Biotechnology ,Inclusion bodies ,Hydrolysis ,chemistry.chemical_compound ,Bioreactors ,Escherichia coli ,medicine ,Beta-galactosidase ,Inclusion Bodies ,Chromatography ,biology ,Escherichia coli Proteins ,General Medicine ,Enzymes, Immobilized ,beta-Galactosidase ,medicine.anatomical_structure ,Biochemistry ,chemistry ,Batch Cell Culture Techniques ,Biocatalysis ,biology.protein ,Biotechnology - Abstract
In this study, we investigated the performance of an immobilized β-galactosidase inclusion bodies-containing Escherichia coli cell reactor, where the cells were immobilized in alginate beads, which were then used in repeated-batch operations for the hydrolysis of o-nitrophenyl-β-D-galactoside or lactose over the long-term. In particular, in the Tris buffer system, disintegration of the alginate beads was not observed during the operation, which was observed for the phosphate buffer system. The o-nitrophenyl-β-D-galactoside hydrolysis was operated successfully up to about 80 h, and the runs were successfully repeated at least eight times. In addition, hydrolysis of lactose was successfully carried out up to 240 h. Using Western blotting analyses, it was verified that the beta-galactosidase inclusion bodies were sustained in the alginate beads during the repeated-batch operations. Consequently, we experimentally verified that β-galactosidase inclusion bodies-containing Escherichia coli cells could be used in a repeated-batch reactor as a biocatalyst for the hydrolysis of o-nitrophenyl-β-D-galactoside or lactose. It is probable that this approach can be applied to enzymatic synthesis reactions for other biotechnology applications, particularly reactions that require long-term and stable operation.
- Published
- 2011
9. Preparation of Branched Dextran Microspheres of Soluble Interferon Interferon-alpha and its Activity In Vitro and In Vivo
- Author
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Hua Hong, Jun Tack Hong, Kyung-Hwan Jung, Byeong-Churl Jang, Sun Kyun Yoo, Jeong Rang Jo, and Ji-Hyeon Yeon
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Glycidyl methacrylate ,Time Factors ,Alpha interferon ,Applied Microbiology and Biotechnology ,Cell Line ,Plasma ,Animal data ,chemistry.chemical_compound ,In vivo ,Animals ,Humans ,Immunologic Factors ,Drug Carriers ,Chromatography ,Activator (genetics) ,Interferon-alpha ,Dextrans ,General Medicine ,Blood proteins ,Microspheres ,In vitro ,Rats ,Dextran ,Biochemistry ,chemistry ,Delayed-Action Preparations ,Hepatocytes ,Protein Binding ,Signal Transduction ,Biotechnology - Abstract
The study objective was to prepare biodegradable branched dextran microspheres encapsulated with His-tagged interferon-alpha (BDM-hIFN-alpha) and evaluate its activity in vitro and in vivo. The glycidyl methacrylate derivatized dextrans (Dex-GMA) as precursor was primarily synthesized by substituting hydroxyl groups of either the branched or linear type of dextran with GMA. Dex-GMA microspheres loaded with hIFN-alpha was then prepared by the water-in-water emulsion technique. In vitro release and Western blotting experiments demonstrated the retained activity of hIFN-alpha released from branched dextran microspheres at 24 h by inducing phosphorylation of signal transducer and activator transcription-1 (STAT-1), a down-stream effector of IFN-alpha, in HepG2 cells. Animal data further revealed a peak of plasma levels of IFN-alpha in rats injected intravenously with BDM-hIFN-alpha at 10 min post-injection, but a sharp decline at 2 h. High plasma levels of neopterin, a plasma protein induced by IFN-alpha, were also detected in rats injected with BDM-hIFN-alpha at 10 min post-injection. Notably, plasma levels of neopterin remained high at 4 h, but largely declined thereafter.
- Published
- 2011
10. Change in compactness of inclusion bodies of recombinant β-galactosidase expressed in the araBAD promoter system of Escherichia coli
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Kyung-Hwan Jung and Ji-Hyeon Yeon
- Subjects
Chemistry ,Biomedical Engineering ,Catabolite repression ,Repressor ,Bioengineering ,medicine.disease_cause ,Trypsin ,Applied Microbiology and Biotechnology ,Inclusion bodies ,law.invention ,chemistry.chemical_compound ,Biochemistry ,law ,medicine ,Recombinant DNA ,Inducer ,Guanidine ,Escherichia coli ,Biotechnology ,medicine.drug - Abstract
We investigated the relevance of the relationship between the compactness of β-galactosidase inclusion bodies (β-gal IBs) and their enhanced enzymatic activity with or without the addition of D-fucose (inducer analog) or methyl α-D-glucopyranoside (α-MG, catabolite repressor) after induction in the araBAD promoter system of Escherichia coli. Experiments conducted to evaluate the solubilization of β-gal IBs in guanidine hydrochloride as well as their trypsin degradation and temperature stability revealed that β-gal IBs expressed in response to the addition of D-fucose or α-MG had a looser structure. Additionally, β-gal IBs expressed when D-fucose or α-MG was added were more quickly solubilized in guanidine hydrochloride or degraded by trypsin-treatment than those produced when these compounds were not added. Moreover, the activity of β-gal IBs expressed when D-fucose or α-MG were added was less stable at various temperatures. Consequently, we deduced that the looser structure of β-gal IBs resulted in enhanced enzymatic activity of β-gal IBs upon addition of D-fucose or α-MG after induction.
- Published
- 2010
11. Aeration alleviates ethanol inhibition and glycerol production during fed-batch ethanol fermentation
- Author
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Hyeon-Beom Seo, Do-Hyung Kang, Ji-Hyeon Yeon, Myung Hoon Jeong, Kyung-Hwan Jung, and Hyeon-Yong Lee
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Ethanol ,Chemistry ,Biomedical Engineering ,Bioengineering ,Ethanol fermentation ,Applied Microbiology and Biotechnology ,chemistry.chemical_compound ,Biochemistry ,Biofuel ,Yield (chemistry) ,Glycerol ,Ethanol fuel ,Food science ,Industrial and production engineering ,Aeration ,Biotechnology - Abstract
In this study, we investigated the effects of aeration on ethanol inhibition and glycerol production during fed-batch ethanol fermentation. When aeration was conducted at 0.13, 0.33, and 0.8 vvm, the ethanol productivity, specific ethanol production rate, and ethanol yield in the presence of greater than 100 g/L of ethanol were higher than when aeration was not conducted. In addition, estimation of the parameters (α and β) in a model equation of ethanol inhibition kinetics indicated that aeration alleviated ethanol inhibition against the specific growth rate and the specific ethanol production rate. Specifically, when aeration was conducted, the glycerol yield and specific glycerol production rate decreased approximately 50 and 70%, respectively. Finally, the results of this study indicated that aeration during fed-batch ethanol fermentation may improve the ethanol concentration in the final culture broth, as well as the ethanol productivity.
- Published
- 2009
12. Improvement of soluble recombinant interferon-α expression by methyl α-D-glucopyranoside in araBAD promoter system of Escherichia coli
- Author
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Sun Kyun Yoo, You-Jin Lee, Ji-Hyeon Yeon, Kyung-Hwan Jung, and Byeong-Churl Chung
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Transcription rate ,Recombinant interferon ,Cell growth ,Biomedical Engineering ,Repressor ,Bioengineering ,Biology ,medicine.disease_cause ,Applied Microbiology and Biotechnology ,Molecular biology ,Inclusion bodies ,law.invention ,Biochemistry ,Interferon ,law ,medicine ,Recombinant DNA ,Escherichia coli ,Biotechnology ,medicine.drug - Abstract
To improve the soluble expression of recombinant human interferon-α that was directed by the araBAD promoter system of Escherichia coli, we attempted to control the overall protein expression rate via the addition of a repressor, methyl α-D-glucospyranoside (α-MG). Recombinant interferon-α was usually expressed as an inclusion body at the end of DO (dissolved oxygen)-stat fed-batch culture. However, the addition of 0.0025 to 0.01% α-MG after 0.5% L-arabinose induction effectively inhibited a tendency towards the formation of inclusion bodies, in which 67.6 to 73.1% of the expressed interferon-α was found in the soluble fraction. It was likely that the addition of a repressor after L-arabinose induction partially modulated the transcription rate from the araBAD promoter system and changed the ratio of soluble and insoluble interferon-α expression. This modulation might be considered as a method that can improve the soluble expression level of recombinant protein at the optimal temperature for cell growth.
- Published
- 2009
13. Optimization of an Extracellular Dextranase Production from Lipomyces starkeyi KCTC 17343 and Analysis of Its Dextran Hydrolysates
- Author
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Jung Hee Shin, Sun-Kyun Yoo, Kyung-Hwan Jung, Joong-Hyun Yeom, Yoon-Hyuck Chang, and Byung Chul Chang
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chemistry.chemical_classification ,Dextranase ,Chromatography ,biology ,Hydrolysate ,Enzyme assay ,Hydrolysis ,chemistry.chemical_compound ,Enzyme ,Dextran ,chemistry ,Biochemistry ,Extracellular ,biology.protein ,Fermentation - Abstract
We optimized dextranase culture conditions by batch fermentation using Lipomyces starkeyi KCTC 17343. Furthermore, dextranase was purified by an ultra-membrane, and then dextran hydrolyzates were characterized. Cell growth and dextranase production varied depending on the initial culture pH and temperature. The conditions of optimal dextranase production were met in a pH range of 4-5 and temperature between 25-30℃. At optimal fermentation conditions, total enzyme activity and specific enzyme activity were about 4.85 IU/㎖ and 0.79 IU/g cells, respectively. The specific growth rate was examined to be 0.076 hr-¹. The production of dextranase in culture broth was very stably maintained after mid-log phase of growth. The enzyme hydrolyzed dextran into DP (degree of polymerization) 2 to 8 oligodextran series. Analysis of the composition of hydrolysates suggested that the enzyme produced is an endo-dextranase.
- Published
- 2009
14. Process Optimization of Dextran Production by Leuconostoc sp. strain YSK. Isolated from Fermented Kimchi
- Author
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Jun-Taek Hong, Jung-Hee Shin, Sung-Paal Yim, Byung Chul Chang, Kyung-Hwan Jung, Seung-Kyun Hwang, Kyung-Suk Hwang, and Sun-Kyun Yoo
- Subjects
chemistry.chemical_compound ,Sucrose ,Dextran ,chemistry ,Biochemistry ,Strain (chemistry) ,Yield (chemistry) ,Yeast extract ,Fermentation ,Response surface methodology ,Food science ,Enrichment culture - Abstract
A bacterium producing non- or partially digestible dextran was isolated from kimchi broth by enrichment culture technique. The bacterium was identified tentatively as Leuconostoc sp. strain SKY. We established the response surface methodology (Box-Behnken design) to optimize the principle parameters such as culture pH, temperature, and yeast extract concentration for maximizing production of dextran. The ranges of parameters were determined based on prior screening works done at our laboratory and accordingly chosen as 5.5, 6.5, and 7.5 for pH, 25, 30, and for temperature, and 1, 5, and 9 g/l yeast extract. Initial concentration of sucrose was 100 g/l. The mineral medium consisted of 3.0 g , 0.01 g , 0.01 g , 0.2 g , 0.01 g NaCl, and 0.05 g per 1 liter deionized water. The optimum values of pH and temperature, and yeast extract concentration were obtained at pH (around 7.0), temperature (27 to ), and yeast extract (6 to 7 g/l). The best dextran yield was 60% (dextran/g sucrose). The best dextran productivity was 0.8 g/h-l.
- Published
- 2008
15. Magnolol elicits activation of the extracellular signal-regulated kinase pathway by inducing p27KIP1-mediated G2/M-phase cell cycle arrest in human urinary bladder cancer 5637 cells
- Author
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Sung Soo Park, Keerang Park, Kyung-Hwan Jung, Sung-Kwon Moon, Wun-Jae Kim, Se-Jung Lee, Eun-Jung Kim, and Young-Hwa Cho
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G2 Phase ,MAPK/ERK pathway ,medicine.medical_specialty ,Cell Survival ,Apoptosis ,Biochemistry ,Lignans ,chemistry.chemical_compound ,Cyclin-dependent kinase ,Cell Line, Tumor ,Internal medicine ,medicine ,Humans ,Enzyme Inhibitors ,Phosphorylation ,Extracellular Signal-Regulated MAP Kinases ,Cell Proliferation ,Pharmacology ,Cyclin-dependent kinase 1 ,Dose-Response Relationship, Drug ,biology ,Cell growth ,Biphenyl Compounds ,Cell Cycle ,Cell cycle ,Antineoplastic Agents, Phytogenic ,Magnolol ,Cell biology ,Endocrinology ,Urinary Bladder Neoplasms ,chemistry ,Mitogen-activated protein kinase ,biology.protein ,G1 phase ,Cell Division ,Cyclin-Dependent Kinase Inhibitor p27 ,Signal Transduction - Abstract
Magnolol has been reported to play a role in antitumor activity. However, the relevant pathway integrating cell cycle regulation and signaling pathways involved in growth inhibition in cancer cells remains to be identified. In the present study, magnolol treatment of these cells resulted in significant dose-dependent growth inhibition together with apoptosis, G1- and G2/M-phase cell cycle arrest at a 60 microM (IC50) dose in 5637 bladder cancer cells. In addition, magnolol treatment strongly induced p27KIP1 expression, and down-regulated expression of cyclin-dependent kinases (CDKs) and cyclins. Moreover, treatment with magnolol-induced phosphorylation of ERK, p38 MAP kinase, and JNK. Among the pathway inhibitors examined, only PD98059, an ERK-specific inhibitor, blocked magnolol-dependent p27KIP1 expression. Blockade of ERK function consistently reversed magnolol-mediated inhibition of cell proliferation and decreased G2/M cell cycle proteins, but not G1 cell cycle proteins. Furthermore, magnolol treatment increased both Ras and Raf activation. Transfection of cells with dominant negative Ras (RasN17) and Raf (RafS621A) mutant genes suppressed magnolol-induced ERK activity and p27KIP1 expression. Finally, the magnolol-induced reduction in cell proliferation and G2/M cell cycle proteins was also abolished in the presence of RasN17 and RafS621A mutant genes. These data demonstrate that the Ras/Raf/ERK pathway participates in p27KIP1 induction, leading to a decrease in the levels of cyclin B1/Cdc2 complexes and magnolol-dependent inhibition of cell growth. Overall, these novel findings concerning the molecular mechanisms of magnolol in 5637 bladder cancer cells provide a theoretical basis for therapeutic treatment of malignancies.
- Published
- 2008
16. Methyl α-d-glucopyranoside enhances the enzymatic activity of recombinant β-galactosidase inclusion bodies in the araBAD promoter system of Escherichia coli
- Author
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Kyung-Hwan Jung, Sung-Kwon Moon, Ji-Hyeon Yeon, and Joon Ho Choi
- Subjects
Catabolite repression ,Down-Regulation ,Enzyme Activators ,Gene Expression ,Repressor ,Bioengineering ,Biology ,medicine.disease_cause ,Applied Microbiology and Biotechnology ,Inclusion bodies ,law.invention ,law ,Escherichia coli ,medicine ,Promoter Regions, Genetic ,Inclusion Bodies ,Escherichia coli Proteins ,Structural gene ,Methylglucosides ,beta-Galactosidase ,biology.organism_classification ,Molecular biology ,Enterobacteriaceae ,Recombinant Proteins ,Biochemistry ,Recombinant DNA ,Specific activity ,Biotechnology - Abstract
In this study, we utilized a catabolite repressor to improve the enzymatic activity of recombinant beta-galactosidase inclusion bodies (IBs) produced in Escherichia coli under the araBAD promoter system. Specifically, we employed methyl alpha-D: -glucopyranoside (alpha-MG) to lower the transcription rate of the beta-galactosidase structural gene. In deepwell microtiter plate and lab-scale fermentor culture systems, we demonstrated that the addition of alpha-MG after induction improved the specific beta-galactosidase production, even though beta-galactosidase was still produced as an IB. Particularly, the addition of 0.0025% alpha-MG led to the most significant increase in the specific activity of the beta-galactosidase. Interestingly, the beta-galactosidase IBs obtained in the presence of 0.0025% alpha-MG were more loosely packed, as determined by IB solubilization in guanidine hydrochloride solution. We propose that the reduced gene transcription rate was responsible for the increased specific beta-galactosidase activity and the loose packing that characterized the IBs produced in the presence of alpha-MG. This principle could be applied throughout the enzyme bioprocessing industry in order to enhance the activity of aggregate-prone enzymes within IBs.
- Published
- 2008
17. Continuous production of recombinant interferon-α in Escherichia coli via the derepression of trp promoter using casamino acid
- Author
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Kyung-Hwan Jung
- Subjects
Tryptophan ,Casamino acid ,Bioengineering ,Biological activity ,Biology ,medicine.disease_cause ,Applied Microbiology and Biotechnology ,Biochemistry ,Continuous production ,Dilution ,chemistry.chemical_compound ,chemistry ,Cytoplasm ,medicine ,Escherichia coli ,Derepression - Abstract
In this study, the continuous production of trp promoter-directed interferon-α, using recombinant Escherichia coli, was achieved. The trp promoter was derepressed via the depletion of l -tryptophan in the medium, using casamino acid as an organic nitrogen source. The interferon-α production occurring in this condition was confirmed in a flask culture. A medium for continuous operation was also designed and, using that medium, demonstrated that cell concentration and interferon-α production increased proportionally according to glucose concentrations in the reservoir during the continuous culture (dilution rate, D = 0.2 1/h). To examine the optimal condition for continuous production of interferon-α, the continuous operations were carried out at various dilution rates (0.125, 0.254, 0.32, and 0.41 1/h), in which the glucose concentration in the reservoir was 25 g/L. Stable continuous operation was maintained at D = 0.254 1/h, but only for 51 h. The specific interferon-α production (Yp/x) and specific interferon-α production rates (Qp) were estimated at 4.03 ± 1.23 mg/g and 1.02 ± 0.31 mg/g/h, respectively. As these values were far lower than those commensurate with high-level expression or high-cell-density cultures, it was inferred that interferon-α was being expressed as a soluble form. In other words, the interferon-α was expressed as a biologically active form within the cytoplasm. Finally, it was concluded that the recombinant interferon-α could be produced stably on a continuous basis in this study, in which the trp promoter was derepressed simply by including casamino acid in the culture medium.
- Published
- 2006
18. Crystallization of Antiangiogenic Kringle V Derived from Human Apolipoprotein A: Crystallization Applied to Purification and Formulation
- Author
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Jeong Woon Hong, Kwan Yub Kang, In Hwan Lim, Kyung Hwan Jung, See Hyoung Park, Sung-Geun Kim, Jung Hwan Park, Chan Wha Kim, Won Kyung Kim, and Hyun Kyung You
- Subjects
Apolipoprotein B ,Stereochemistry ,Angiogenesis Inhibitors ,Applied Microbiology and Biotechnology ,Biochemistry ,Mass Spectrometry ,Umbilical vein ,Umbilical Cord ,Analytical Chemistry ,law.invention ,Pichia pastoris ,Kringles ,Cell Movement ,law ,Humans ,Molecule ,Crystallization ,Molecular Biology ,Apolipoproteins A ,Cells, Cultured ,Hplc analysis ,Chromatography ,biology ,Human apolipoprotein ,Chemistry ,Organic Chemistry ,Endothelial Cells ,General Medicine ,Hydrogen-Ion Concentration ,biology.organism_classification ,Molecular Weight ,Solubility ,Yield (chemistry) ,biology.protein ,Chromatography, Liquid ,Biotechnology - Abstract
In this study, the Kringle V domain (Glu4225-Ser4310) of human apolipoprotein A, an antiangiogenic polypeptide, was expressed as a secreted form in Pichia pastoris, and was purified via a process consisting of three chromatographic steps. The chromatographically purified kringle V domain contained a C-terminal serine-deleted form and several high-molecular-weight forms, which were suspected to represent glycosylated derivatives. In order to remove these derivatives, we employed a crystallization process. The crystallization of kringle V resulted in an 85% recovery yield, and also resulted in the complete removal of the aforementioned high-molecular-weight forms. However, we were still able to detect a trace of the C-terminal serine-deleted form. The prepared Kringle V crystals were stable within a pH range of 7.0 to 8.0, and were completely dissolved by dilution, which is a crucial factor in the preparation of a highly concentrated formulation. The chromatogram of the crystallized kringle V on reversed-phase HPLC analysis was identical to that observed without crystallization. Also, we noted that the original anti-wound migration activities of the molecule toward human umbilical vein endothelial cells were completely retained.
- Published
- 2006
19. High-yield purification and characterization of recombinant human leukotactin-1 inPichia pastoris
- Author
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Kyung-Hwan Jung, Mu Rim Choi, Yeup Yoon, Kong Ju Lee, Doo-Hong Park, In Hwan Lim, Eun Kyoung Lee, and Gue-Wha Lee
- Subjects
chemistry.chemical_classification ,Chromatography ,Biomedical Engineering ,Bioengineering ,Biological activity ,Reversed-phase chromatography ,Biology ,biology.organism_classification ,Applied Microbiology and Biotechnology ,Yeast ,Pichia pastoris ,Amino acid ,law.invention ,Biochemistry ,chemistry ,law ,Yield (chemistry) ,Recombinant DNA ,Biotechnology ,Chemotaxis assay - Abstract
The human chemokine, the short version of leukotactin-1 (shLkn-1; molecular weight =7.2 kD and 66 amino acids), was expressed and secreted into a culture medium using the methylotrophic yeast,Pichia pastoris. The recombinant shLkn-1 was purified from the culture supernatant using a simple two-step procedure consisting of cation exchange and reverse phase chromatography (RPC), in which shLkn-1 was highly purified (99.5%) with a high recovery yield of 82.7%. The C-terminal truncated derivative of shLkn-1 was found in the supernatant and was separated by RPC. The physicochemical properties of the purified shLkn-1 were verified to be the same as expected. The biological activity of the purified recombinant shLkn-1 was also quantified using a chemotaxis assay. It was observed that the recombinant shLkn-1 had the maximum migration activity at a concentration of 10 nM, as potent as MIP-1α.
- Published
- 2004
20. High dissolved oxygen tension enhances heterologous protein expression by recombinant Pichia pastoris
- Author
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Kyung Hwan Jung, Eun Kyu Lee, Shigeo Katoh, Chun Yeon Lee, and Seok Jea Lee
- Subjects
Oxygenase ,biology ,Elastase ,chemistry.chemical_element ,Heterologous ,Bioengineering ,biology.organism_classification ,Applied Microbiology and Biotechnology ,Biochemistry ,Oxygen ,Pichia pastoris ,Elastase inhibitor ,chemistry ,Derepression ,Pichia - Abstract
The effect of dissolved oxygen (DO) tension on heterologous protein expression by the Pichia system was investigated by applying DO control bands of different set points and bandwidths in the DO-stat fed-batch fermentation. The model protein was elastase inhibiting peptide (EIP) that showed strong inhibitory activity against human neutrophil elastase with very high specificity. In the alcohol oxygenase 1 derepression phase, more frequent glycerol feeding by keeping the DO control band narrower resulted in a higher cell growth rate (0.10 h−1). In the methanol induction phase, maintaining a higher DO set point and allowing more frequent feeding significantly enhanced EIP expression. The EIP expression titre was increased about threefold to 846 mg l−1 on average by raising the lower set point from 10 to 30% air-saturation. Both specific expression titre and methanol consumption rates were increased about threefold, which indicated that higher DO tension stimulated the methanol utilization pathway and improved the expression efficiency. To operate the DO-stats with higher DO set points, an oxygen enriching membrane system that could generate up to 38% oxygen partial pressure from the regular air was used as an alternative oxygen supplier to the traditional gas-mixing system consisting of an air compressor and a pure oxygen cylinder.
- Published
- 2003
21. Ethanol Production from Glycerol by the Yeast Pachysolen tannophilus Immobilized on Celite during Repeated-Batch Flask Culture
- Author
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Kyung-Hwan Jung, Do-Hyung Kang, Hye-Geun Cha, Woon Yong Choi, Yi-Ok Kim, and Hyeon-Yong Lee
- Subjects
0106 biological sciences ,Glycerol ,020209 energy ,02 engineering and technology ,01 natural sciences ,Microbiology ,Pachysolen tannophilus ,chemistry.chemical_compound ,Immobilization ,Celite ,010608 biotechnology ,0202 electrical engineering, electronic engineering, information engineering ,Ethanol fuel ,Chromatography ,Ethanol ,biology ,biology.organism_classification ,Yeast ,Research Note ,Infectious Diseases ,chemistry ,Biochemistry ,Ethanol production ,Fermentation - Abstract
We investigated a novel process for production of ethanol from glycerol using the yeast Pachysolen tannophilus. After optimization of the fermentation medium, repeated-batch flask culture was performed over a period of 378 hr using yeast cells immobilized on Celite. Our results indicated that the use of Celite for immobilization of P. tannophilus was a practical approach for ethanol production from glycerol, and should be suitable for industrial ethanol production.
- Published
- 2014
22. Two-step process using immobilized Saccharomyces cerevisiae and Pichia stipitis for ethanol production from Ulva pertusa Kjellman hydrolysate
- Author
-
Do-Hyung Kang, Hyeon-Yong Lee, Woo Yong Choi, Sang-Eun Lee, Kyung-Hwan Jung, and Yi-Ok Kim
- Subjects
chemistry.chemical_classification ,Ethanol ,Xylose ,biology ,Biomass ,General Medicine ,Saccharomyces cerevisiae ,biology.organism_classification ,Applied Microbiology and Biotechnology ,Hydrolysate ,Yeast ,Pichia ,Reducing sugar ,chemistry.chemical_compound ,Ulva ,Bioreactors ,Glucose ,Biochemistry ,chemistry ,Ethanol fuel ,Food science ,Pichia stipitis ,Biotechnology - Abstract
We established a two-step production process using immobilized S. cerevisiae and P. stipitis yeast to produce ethanol from seaweed (U. pertusa Kjellman) hydrolysate. The process was designed to completely consume both glucose and xylose. In particular, the yeasts were immobilized using DEAE-corncob and DEAE-cotton, respectively. The first step of the process included a continuous column reactor using immobilized S. cerevisiae, and the second step included a repeated-batch reactor using immobilized P. stipitis. It was verified that the glucose and xylose in 20 L of medium containing the U. pertusa Kjellman hydrolysate was converted completely to about 5.0 g/l ethanol through the two-step process, in which the overall ethanol yield from total reducing sugar was 0.37 and the volumetric ethanol productivity was 0.126 g/ l/h. The volumetric ethanol productivity of the two-step process was about 2.7 times greater than that when P. stipitis was used alone for ethanol production from U. pertusa Kjellman hydrolysate. In addition, the overall ethanol yield from glucose and xylose was superior to that when P. stipitis was used alone for ethanol production. This two-step process will not only contribute to the development of an integrated process for ethanol production from glucose and xylose-containing biomass hydrolysates, but could also be used as an alternative method for ethanol production.
- Published
- 2013
23. Production of chlorphenesin galactoside by whole cells of β-galactosidase-containing Escherichia coli
- Author
-
Kyung-Hwan Jung, Sang-Eun Lee, and Hyang-Yeol Lee
- Subjects
Lactose ,medicine.disease_cause ,Applied Microbiology and Biotechnology ,Mass Spectrometry ,chemistry.chemical_compound ,Chlorphenesin ,Enzyme Stability ,medicine ,Escherichia coli ,Sugar moiety ,Beta-galactosidase ,Derivatization ,Biotransformation ,Chromatography ,biology ,Chemistry ,Temperature ,Galactose ,General Medicine ,Hydrogen-Ion Concentration ,beta-Galactosidase ,Galactoside ,Biochemistry ,biology.protein ,Chromatography, Thin Layer ,Metabolic Networks and Pathways ,Biotechnology ,Chromatography, Liquid - Abstract
We investigated the transgalactosylation reaction of chlorphenesin (CPN) using β-galactosidase (β-gal)-containing Escherichia coli (E. coli) cells, in which galactose from lactose was transferred to CPN. The optimal CPN concentration for CPN galactoside (CPN-G) synthesis was observed at 40 mM under the conditions that lactose and β-gal (as E. coli cells) were 400 g/l and 4.8 U/ml, respectively, and the pH and temperature were 7.0 and 40oC, respectively. The time-course profile of CPN-G synthesis under these optimal conditions showed that CPN-G synthesis from 40 mM CPN reached a maximum of about 27 mM at 12 h. This value corresponded to an about 67% conversion of CPN to CPN-G, which was 4.47-5.36-fold higher than values in previous reports. In addition, we demonstrated by thin-layer chromatography to detect the sugar moiety that galactose was mainly transferred from lactose to CPN. Liquid chromatography-mass spectrometry revealed that CPN-G and CPN-GG (CPN galactoside, which accepted two galactose molecules) were definitively identified as the synthesized products using β-gal-containing E. coli cells. In particular, because we did not use purified β-gal, our β-gal-containing E. coli cells might be practical and cost-effective for enzymatically synthesizing CPN-G. It is expected that the use of β-gal-containing E. coli will be extended to galactose derivatization of other drugs to improve their functionality.
- Published
- 2013
24. Ultraviolet A regulates adipogenic differentiation of human adipose tissue-derived mesenchymal stem cells via up-regulation of Kruppel-like factor 2
- Author
-
Eunsun Jung, Youngsoo Kim, Deokhoon Park, Kyung-Hwan Jung, Kyung-Baeg Roh, Jienny Lee, and Jongsung Lee
- Subjects
MAPK/ERK pathway ,medicine.medical_specialty ,Ultraviolet Rays ,Cellular differentiation ,Kruppel-Like Transcription Factors ,Adipose tissue ,Biology ,AMP-Activated Protein Kinases ,Biochemistry ,chemistry.chemical_compound ,Mice ,Internal medicine ,Adipocyte ,3T3-L1 Cells ,medicine ,Animals ,Humans ,RNA, Small Interfering ,Protein kinase A ,Molecular Biology ,Adipogenesis ,Cell Differentiation ,Mesenchymal Stem Cells ,Cell Biology ,Cell biology ,Up-Regulation ,PPAR gamma ,AP-1 transcription factor ,Endocrinology ,chemistry ,Adipose Tissue ,KLF2 ,Cytokines ,sense organs ,Transcription Factors - Abstract
Adipocyte dysfunction is strongly associated with the development of obesity, which is a major risk factor for many disorders, including diabetes, hypertension, and heart disease. This study shows that ultraviolet A (UVA) inhibits adipogenic differentiation of human adipose tissue-derived mesenchymal stem cells and its action mechanisms. The mRNA levels of peroxidase proliferator-activated receptor (PPAR) γ and CCAAT/enhancer-binding protein α (C/EBPα), but not CCAAT/enhancer-binding protein ((C/EBP) β and δ, were reduced by UVA. Moreover, the mRNA levels of PPAR γ target genes (lipoprotein lipase (LPL), CD36, adipocyte protein (aP2), and liver X receptor α (LXR)) were down-regulated by UVA. Additionally, attempts to elucidate a possible mechanism underlying the UVA-mediated effects revealed that UVA induced migration inhibitory factor (MIF) gene expression, and this was mediated through activation of AP-1 (especially JNK and p42/44 MAPK) and nuclear factor-κB. In addition, reduced adipogenesis by UVA was recovered upon the treatment with anti-MIF antibodies. AMP-activated protein kinase phosphorylation and up-regulation of Kruppel-like factor 2 (KLF2) were induced by UVA. Taken together, these findings suggest that the inhibition of adipogenic differentiation of human adipose tissue-derived mesenchymal stem cells by UVA occurs primarily through the reduced expression of PPAR γ, which is mediated by up-regulation of KLF2 via the activation of MIF-AMP-activated protein kinase signaling.
- Published
- 2010
25. c-Jun N-terminal kinase 1 is required for cordycepin-mediated induction of G2/M cell-cycle arrest via p21WAF1 expression in human colon cancer cells
- Author
-
Kyung-Hwan Jung, Sung-Kwon Moon, Gi-Seong Moon, Wun-Jae Kim, and Se-Jung Lee
- Subjects
Cyclin-Dependent Kinase Inhibitor p21 ,G2 Phase ,Small interfering RNA ,Cell Survival ,Blotting, Western ,Apoptosis ,Enzyme-Linked Immunosorbent Assay ,Biology ,Toxicology ,p38 Mitogen-Activated Protein Kinases ,chemistry.chemical_compound ,Cell Line, Tumor ,Humans ,Immunoprecipitation ,Mitogen-Activated Protein Kinase 9 ,Mitogen-Activated Protein Kinase 8 ,Enzyme Inhibitors ,RNA, Small Interfering ,Extracellular Signal-Regulated MAP Kinases ,Anthracenes ,Cyclin-dependent kinase 1 ,Cordycepin ,Deoxyadenosines ,Cell growth ,Kinase ,c-jun ,Cell Cycle ,General Medicine ,Cell cycle ,Cell biology ,Biochemistry ,chemistry ,Cell culture ,Colonic Neoplasms ,Cell Division ,Food Science - Abstract
Cordycepin (3'-deoxyadenosine) has many anti-cancer properties. However, neither its molecular mechanism nor its molecular targets are well understood. In the present study, we investigated novel molecular mechanisms for the anti-tumor effects of cordycepin in human colon cancer HCT116 cells. After treatment of cells with cordycepin, dose-dependent cell growth inhibition was observed at an IC(50) value of 200muM. Cordycepin treatment resulted in G2/M-phase cell-cycle arrest, which was associated with increased p21WAF1 levels and reduced amounts of cyclin B1, Cdc2, and Cdc25c in a p53-independent pathway. Moreover, cordycepin treatment induced activation of JNK (c-Jun N-terminal kinases). Pretreatment with SP600125, a JNK-specific inhibitor, abrogated cordycepin-mediated p21WAF1 expression, cell growth inhibition, and reduced cell-cycle proteins. Furthermore, JNK1 inhibition by small interfering RNA (siRNA) produced similar results: suppression of cordycepin-induced p21WAF1 expression, decreased cell growth, and reduced cell-cycle proteins. Together, these results suggest a critical role for JNK1 activation in cordycepin-induced inhibition of cell growth and G2/M-phase arrest in human colon cancer cells.
- Published
- 2009
26. Supplement of nutrients for effective cultivation of hepatitis B surface antigen-producing recombinant yeast
- Author
-
Joon Shick Rhee, Kyung Hwan Jung, Hong Mo Moon, and Mahn Hoon Park
- Subjects
chemistry.chemical_classification ,Hepatitis ,HBsAg ,biology ,Saccharomyces cerevisiae ,Bioengineering ,General Medicine ,biology.organism_classification ,medicine.disease ,Applied Microbiology and Biotechnology ,Yeast ,Amino acid ,Plasmid ,chemistry ,Biochemistry ,Cell culture ,medicine ,Bioorganic chemistry ,Biotechnology - Abstract
In the hepatitis B surface antigen (HBsAg)-producing recombinant yeast culture medium, the supply of Bacto-yeast nitrogen base without amino acids was found to be inadequate due to the lack of the several kinds of vitamins and trace elements. When the culture medium for this recombinant yeast was supplemented with sufficient vitamins and trace elements, its growth, HBsAg production and the stability of plasmid were improved.
- Published
- 1991
27. Improving the yield of soluble 6xHis-tagged interferon-alpha via the addition of repressor of the araBAD promoter system in Escherichia coli
- Author
-
Kyung-Hwan Jung, Ji-Hyeon Yeon, Byeong-Churl Jang, Young-Seo Park, Sung-Hun Kim, and Sun Kyun Yoo
- Subjects
Glycerol ,Blotting, Western ,Alpha interferon ,Repressor ,Bioengineering ,medicine.disease_cause ,Applied Microbiology and Biotechnology ,law.invention ,chemistry.chemical_compound ,law ,Operon ,medicine ,Escherichia coli ,Humans ,Histidine ,Promoter Regions, Genetic ,Polyacrylamide gel electrophoresis ,Electrophoresis, Agar Gel ,Inclusion Bodies ,biology ,Interferon-alpha ,General Medicine ,biology.organism_classification ,Enterobacteriaceae ,Recombinant Proteins ,Repressor Proteins ,Biochemistry ,chemistry ,Solubility ,Recombinant DNA ,Agarose ,Electrophoresis, Polyacrylamide Gel ,Oligopeptides ,Biotechnology - Abstract
The inhibition of inclusion body formation in Escherichia coli by the addition of alpha-D-glucopyranoside or D-fucose after induction improved the purification yield of soluble recombinant interferon-alpha. When D-fucose was added after induction, more soluble 6xHis-tagged interferon-alpha could be purified compared to when methyl alpha-D-glucopyranoside was added. It was shown that, on the basis of 1 mg dry cell weight, 16.6 microg of soluble 6xHis-tagged interferon-alpha was purified when D-fucose was added after induction and 6 ml nickel-chelated agarose gel column was used. This was about 15 times greater than when induction only was performed and 1 ml nickel-chelated agarose gel was used.
- Published
- 2008
28. Paradoxical effects of elastase inhibitor guamerin on the tissue repair of two different wound models: sealed cutaneous and exposed tongue wounds
- Author
-
Jin-Woo Kim, Heung Sik Um, Yeon Sook Kim, Sang Shin Lee, In Sun Song, Soo Il Chung, Kyung-Hwan Jung, Kyeong Yeon Kim, Suk Keun Lee, Jae Yong Joo, Young Wook Park, and Seung-Jin Choi
- Subjects
Pathology ,medicine.medical_specialty ,Stromal cell ,Invertebrate Hormones ,Clinical Biochemistry ,Fibrin Tissue Adhesive ,Biochemistry ,Fibrin ,Mice ,Tongue ,medicine ,Animals ,Enzyme Inhibitors ,Molecular Biology ,Skin ,Wound Healing ,integumentary system ,biology ,business.industry ,Macrophages ,Elastase ,Granulation tissue ,medicine.disease ,Elastase inhibitor ,medicine.anatomical_structure ,Immunology ,biology.protein ,Molecular Medicine ,Female ,Wound healing ,business ,Leukocyte Elastase ,Infiltration (medical) - Abstract
Innate elastase inhibitors are known to be putatively involved in the regulation of tissue inflammation by inhibiting polymorphonuclear leukocyte (PMN) derived proteinases. The aim of this study was to evaluate affects of leukocyte elastase suppression and PMN infiltration on wound healing in mouse by administering the recombinant elastase inhibitor guamerin (rEIG) in two different wound models; 1) impaired pin-punctured dorsal mucosa of anterior tongue wound, 60 mice, treated with saline containing rEIG that were fed ad libitum and 2) stable linear excisional cutaneous wound, 40 mice, covered with fibrin sealant containing rEIG. The progress of healing was analyzed by histological methods. The tongue wounds treated with rEIG became edematous around the pin-punctured tongue wound, and influx of inflammatory cells and PMN into the underlying stromal tissue were seen rapidly after wounding and peaked between 2-4 days. Whereas the control mice showed almost no wheal formation in the pin-punctured wound, a far lesser levels of PMN infiltration, and almost complete wound closure in 4 days. In the other model, the liner excisional cutaneous wound treated with fibrin sealant containing rEIG showed early wound constriction, lesser degree of inflammatory cells influx, and complete reepithelialization in 4-5 days, whereas the wound of control mice with the fibrin sealant alone showed contrary delayed reepithelialization, greater degree of inflammatory cell infiltration, and consequencial formation of greater granulation tissue at wound site. Taken together, these data suggest paradoxical effects of rEIG on the wound healing where in the wound exposed to infiltrating milieu of microorganisms in the oral cavity, the rEIG aggravates the wound healing by interfering with other innate defensive factors and extended greater flux of PMNs to inflamed wound site, while in the wound enclosed by fibrin, the rEIG accelerated wound healing by inhibiting the inflammation-generated proteases and the acute inflammatory reaction.
- Published
- 2004
29. Human apolipoprotein(a) kringle V inhibits angiogenesis in vitro and in vivo by interfering with the activation of focal adhesion kinases
- Author
-
Young Ae Joe, Yeup Yoon, Kyung Hwan Jung, Hyun Kyung Yu, Soo-Ik Chang, Si Myung Byun, Ho Jeong Lee, Soon Won Hong, Jin Hyung Ahn, Suk Keun Lee, Jang Seong Kim, Soo Il Chung, and Yong Kil Hong
- Subjects
Umbilical Veins ,Time Factors ,Angiogenesis ,Chick Embryo ,Apoprotein(a) ,Biochemistry ,Umbilical vein ,Neovascularization ,Kringles ,Cell Movement ,Phosphorylation ,Cells, Cultured ,Mitogen-Activated Protein Kinase 1 ,Mitogen-Activated Protein Kinase 3 ,Neovascularization, Pathologic ,Chorion ,Protein-Tyrosine Kinases ,Recombinant Proteins ,Angiogenesis inhibitor ,Drug Combinations ,Electrophoresis, Polyacrylamide Gel ,Proteoglycans ,Collagen ,medicine.symptom ,Mitogen-Activated Protein Kinases ,DNA, Complementary ,Blotting, Western ,Genetic Vectors ,Molecular Sequence Data ,Biophysics ,Biology ,Kringle domain ,Focal adhesion ,In vivo ,medicine ,Animals ,Humans ,Amino Acid Sequence ,Molecular Biology ,Wound Healing ,Dose-Response Relationship, Drug ,Cell Biology ,Molecular biology ,In vitro ,Actins ,Protein Structure, Tertiary ,Apolipoproteins ,Focal Adhesion Kinase 1 ,Focal Adhesion Protein-Tyrosine Kinases ,Tyrosine ,Endothelium, Vascular ,Laminin ,Lipoprotein(a) - Abstract
Apolipoprotein(a) [apo(a)] contains the largest numbers of kringle domains identified to date. Of these, apo(a) kringle V shows significant sequence homology with plasminogen kringle 5, which is reported to be a potent angiogenesis inhibitor. To determine the effects of apo(a) kringle V on angiogenesis, it was expressed as a soluble protein (termed rhLK8) in Pichia pastoris and its in vitro and in vivo anti-angiogenic properties were examined. rhLK8 inhibited the migration of human umbilical vein endothelial cells in vitro in a dose-dependent manner. This function was associated with the down-regulation of the activation of focal adhesion kinase and the inhibition of the consequent formation of actin stress fibers/focal adhesions. rhLK8 also inhibited new capillary formation in vivo, as assessed by the chick chorioallantoic membrane assay and the Matrigel plug assay. These results indicate that rhLK8 may be an effective angiogenesis inhibitor both in vitro and in vivo.
- Published
- 2003
30. Inhibition of angiogenesis and angiogenesis-dependent tumor growth by the cryptic kringle fragments of human apolipoprotein(a)
- Author
-
Yeup Yoon, Jung-Sun Yum, Suk-Keun Lee, Doo-Hong Park, Jin-Hyung Ahn, Si-Myung Byun, Soo-Il Chung, Hyun-Kyung Yu, Jihoon Chang, Kyung-Hwan Jung, and Jangseong Kim
- Subjects
Vascular Endothelial Growth Factor A ,Umbilical Veins ,Lung Neoplasms ,Angiogenesis ,Basic fibroblast growth factor ,Gene Expression ,Endothelial Growth Factors ,Biochemistry ,Neovascularization ,chemistry.chemical_compound ,Mice ,Kringles ,Allantois ,Cell Movement ,Neoplasms ,Tumor Cells, Cultured ,Phosphorylation ,Cells, Cultured ,In Situ Hybridization ,Mitogen-Activated Protein Kinase 1 ,Lymphokines ,Mice, Inbred BALB C ,Mitogen-Activated Protein Kinase 3 ,Neovascularization, Pathologic ,Vascular Endothelial Growth Factors ,Chorion ,Recombinant Proteins ,Vascular endothelial growth factor ,medicine.anatomical_structure ,Colonic Neoplasms ,Intercellular Signaling Peptides and Proteins ,Human umbilical vein endothelial cell ,Female ,Fibroblast Growth Factor 2 ,medicine.symptom ,Mitogen-Activated Protein Kinases ,Cell Division ,Angiogenin ,Mice, Nude ,Neovascularization, Physiologic ,Biology ,Kringle domain ,medicine ,Escherichia coli ,Animals ,Humans ,RNA, Messenger ,Fibroblast ,Molecular Biology ,Apolipoproteins A ,Dose-Response Relationship, Drug ,Cell Biology ,Ribonuclease, Pancreatic ,Molecular biology ,Peptide Fragments ,chemistry ,Endothelium, Vascular ,Chickens ,Neoplasm Transplantation - Abstract
Apolipoprotein(a) (apo(a)) contains tandemly repeated kringle domains that are closely related to plasminogen kringle 4, followed by a single kringle 5-like domain and an inactive protease-like domain. Recently, the anti-angiogenic activities of apo(a) have been demonstrated both in vitro and in vivo. However, its effects on tumor angiogenesis and the underlying mechanisms involved have not been fully elucidated. To evaluate the anti-angiogenic and anti-tumor activities of the apo(a) kringle domains and to elucidate their mechanism of action, we expressed the last three kringle domains of apo(a), KIV-9, KIV-10, and KV, in Escherichia coli. The resultant recombinant protein, termed rhLK68, exhibited a dose-dependent inhibition of basic fibroblast growth factor-stimulated human umbilical vein endothelial cell proliferation and migration in vitro and inhibited the neovascularization in chick chorioallantoic membranes in vivo. The ability of rhLK68 to abrogate the activation of extracellular signal-regulated kinases appears to be responsible for rhLK68-mediated anti-angiogenesis. Furthermore, systemic administration of rhLK68 suppressed human lung (A549) and colon (HCT-15) tumor growth in nude mice. Immunohistochemical examination and in situ hybridization analysis of the tumors showed a significant decrease in the number of blood vessels and the reduced expression of vascular endothelial growth factor, basic fibroblast growth factor, and angiogenin, indicating that suppression of angiogenesis may have played a significant role in the inhibition of tumor growth. Collectively, these results suggest that a truncated apo(a), rhLK68, is a potent anti-angiogenic and anti-tumor molecule.
- Published
- 2003
31. Production and characterization of recombinant guamerin, an elastase-specific inhibitor, in the methylotrophic yeast Pichia pastoris
- Author
-
Kyung-Hwan Jung, Kong-Ju Lee, Hyung-Kwon Lim, Ke-Won Kang, Soo-Il Chung, Kyeong Yeon Kim, and Doo-Hong Park
- Subjects
Invertebrate Hormones ,Molecular Sequence Data ,Pichia ,law.invention ,Pichia pastoris ,Immune system ,law ,Amino Acid Sequence ,Enzyme Inhibitors ,Pancreatic elastase ,Peptide sequence ,Molecular mass ,biology ,Base Sequence ,Pancreatic Elastase ,Temperature ,Hydrogen-Ion Concentration ,biology.organism_classification ,Molecular biology ,Yeast ,Recombinant Proteins ,ELASTASE-SPECIFIC INHIBITOR ,Biochemistry ,Recombinant DNA ,Biotechnology ,Chromatography, Liquid - Abstract
The elastase-specific inhibitor, guamerin, was expressed and secreted into a culture medium using the methylotrophic yeast Pichia pastoris, and the resulting recombinant guamerin was purified from the culture media using a two-step procedure composed of a hydrophobic interaction and reverse-phase chromatography. Up to 90 g/L of dry cell weight, the guamerin-producing recombinant P. pastoris was cultivated and guamerin was secreted into the culture medium at a level of 0.69 g/L. The recombinant guamerin was highly purified (>98%) with a recovery yield of 68%. Analyses of the purified guamerin revealed the same N-terminal amino acid sequence, amino acid composition, and molecular mass as found in the native leech protein. The recombinant guamerin exhibited the tight binding to porcine pancreatic elastase. Furthermore, the recombinant guamerin did not produce a humoral immune response in mice.
- Published
- 2000
32. Improved Bioethanol Production Using Activated Carbon-treated Acid Hydrolysate from Corn Hull inPachysolen tannophilus
- Author
-
Seung-Seop Kim, Hyeon-Beom Seo, Kyung-Hwan Jung, and Hyeon-Yong Lee
- Subjects
chemistry.chemical_classification ,Arabinose ,Pachysolen tannophilus ,Bioethanol ,Biology ,Xylose ,Microbiology ,Hydrolysate ,chemistry.chemical_compound ,Hydrolysis ,Corn hull ,Infectious Diseases ,chemistry ,Biochemistry ,Biofuel ,medicine ,Acid hydrolysis ,Monosaccharide ,Food science ,Activated-carbon treatment ,Research Article ,Activated carbon ,medicine.drug - Abstract
To optimally convert corn hull, a byproduct from corn processing, into bioethanol using Pachysolen tannophlius, we investigated the optimal conditions for hydrolysis and removal of toxic substances in the hydrolysate via activated carbon treatment as well as the effects of this detoxification process on the kinetic parameters of bioethanol production. Maximum monosaccharide concentrations were obtained in hydrolysates in which 20 g of corn hull was hydrolyzed in 4% (v/v) H2SO4. Activated carbon treatment removed 92.3% of phenolic compounds from the hydrolysate. When untreated hydrolysate was used, the monosaccharides were not completely consumed, even at 480 h of culture. When activated carbon-treated hydrolysate was used, the monosaccharides were mostly consumed at 192 h of culture. In particular, when activated carbon-treated hydrolysate was used, bioethanol productivity (P) and specific bioethanol production rate (Qp) were 2.4 times and 3.4 times greater, respectively, compared to untreated hydrolysate. This was due to sustained bioethanol production during the period of xylose/arabinose utilization, which occurred only when activated carbon-treated hydrolysate was used.
- Published
- 2009
33. Modulation of the distribution of soluble and insoluble recombinant protein expression by α-MG in araBAD promoter system of Escherichia coli
- Author
-
Byeong-Churl Jang, Kyung-Hwan Jung, and Sun Kyun Yoo
- Subjects
Biochemistry ,Chemistry ,Modulation ,medicine ,Distribution (pharmacology) ,Bioengineering ,General Medicine ,medicine.disease_cause ,Applied Microbiology and Biotechnology ,Escherichia coli ,Protein expression ,Biotechnology - Published
- 2007
34. Production of HBsAg by growth rate control with recombinantSaccharomyces cerevisiae in fed-batch
- Author
-
Mahn Hoon Park, Man Bock Gu, Kwang Soon Shin, Kyong Ho Kim, and Kyung Hwan Jung
- Subjects
HBsAg ,biology ,Cell growth ,Glucose uptake ,Saccharomyces cerevisiae ,Bioengineering ,Industrial fermentation ,General Medicine ,biology.organism_classification ,Applied Microbiology and Biotechnology ,Yeast ,law.invention ,Biochemistry ,law ,Recombinant DNA ,Food science ,Growth rate ,Biotechnology - Abstract
To maintain a constant specific growth rate for a recombinantS.cerevisiae in fed-batch, the medium feeding rate has been controlled with respect to the hourly calculated glucose uptake rate. The recombinant yeast producing HBsAg showed the exponential production trend in proportion to the exponential cell growth. Total cell yield in fed-batch was about 0.402 g cells/g glucose, and HBsAg was produced about ten times more than in batch. Decrease of growth rate by HBsAg produced was not shown.
- Published
- 1989
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