• Journal of Microbiology and Biotechnology
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• v.9 no.3
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• pp.303-310
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• 1999

A cost-effective mass production method for a strong antimicrobial peptide, buforin II, which was isolated from the stomach of Bufo bufo gargarizans, has been developed. This method is based on the neutralization of the positive charge of buforin II by fusion with a cysteine-rich acidic peptide (CAP) to avoid any lethal effect on the host. The neutralized fusion peptide was multimerized and expressed in Escherichia coli as tandem repeats to increase the production yield. Multimers of the CAP-buforin II fusion peptide were successfully expressed at high levels in E. coli as inclusion bodies. More than 100mg of pure buforin II was obtained per 11 of E. coli culture after cleaving the multimeric polypeptide with CNBr. The buforin II obtained from the recombinant E. coli had antimicrobial activity identical to that of natural buforin II. The proposed expression system can provide a cost-effective mass production method for both antimicrobial peptides and other host-lethal basic proteins.

• Journal of Microbiology and Biotechnology
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• v.32 no.6
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• pp.768-775
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• 2022

In this study we aimed to derive the response surface models for Escherichia coli reduction in wheat flour using atmospheric cold plasma (ACP) with three types of gas. The jet-type atmospheric cold plasma wand system was used with a 30 W power supply, and three gases (argon, air, and nitrogen) were applied as the treatment gas. The operating parameters for process optimization considered were wheat flour mass (g), treatment time (min), and gas flow rate (L/min). The wheat flour samples were artificially contaminated with E. coli at a concentration of 9.25 ± 0.74 log CFU/g. ACP treatments with argon, air, and nitrogen resulted in 2.66, 4.21, and 5.55 log CFU/g reduction of E. coli, respectively, in wheat flour under optimized conditions. The optimized conditions to reduce E. coli were 0.5 g of the flour mass, 15 min of treatment time, and 0.20 L/min of nitrogen gas flow rate, and the predicted highest reduction level from modeling was 5.63 log CFU/g.

• Journal of Microbiology and Biotechnology
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• v.17 no.7
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• pp.1162-1168
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• 2007

Although widely used as a host for recombinant protein production, Escherichia coli is unsuitable for massive screening of recombinant clones, owing to its poor secretion of proteins. A vector system containing T4 holin and T7 lysozyme genes under the control of the ptsG promoter derivative that is inducible in the absence of glucose was developed for programmed cell lysis of E. coli. Because E. coli harboring the vector grows well in the presence of glucose, but is lysed upon glucose exhaustion, the activity of the foreign gene expressed in E. coli can be monitored easily without an additional step for cell disruption after the foreign gene is expressed sufficiently with an appropriate concentration of glucose. The effectiveness of the vector was demonstrated by efficient screening of the amylase gene from a Bacillus subtilis genomic library. This vector system is expected to provide a more efficient and economic screening of bioactive products from DNA libraries in large quantities.

• Proceedings of the Korean Biophysical Society Conference
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• 1998.06a
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• pp.12-12
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• 1998

The interaction between the N-terminal domain of enzyme I (EIN) and the histidine-containing phosphocarrier protein HPr of the Escherichia coli phosphoenolpyruvate：sugar phosphotransferase system has been investigated by Isothermal Titration Calorimetry and heteronuclear magnetic resonance spectroscopy.(omitted)

• Biomolecules & Therapeutics
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• v.13 no.2
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• pp.101-106
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• 2005

The generation of secretory IgA antibodies (Abs) for specific immune protection of mucosal surfaces depends on stimulation of the mucosal immune system, but this is not effectively achieved by parenteral or even oral administration of most soluble antigens. Thus, to produce a possible vaccine antigen against urinary tract infections, the uropathogenic E. coli (UPEC) adhesin was genetically coupled to the heat-labile Escherichia coli enterotoxin A2B (ltxa2b) gene and cloned into a pMAL-p2E expression vector. The chimeric construction of pMALfimH/ltxa2b was then transformed into E. coli K-12 TB1 and its nucleotide sequence was verified. The chimeric protein was then purified by applying the affinity chromatography. The purified chimeric protein was confirmed by SDS-PAGE and westem blotting using antibodies to the maltose binding protein (MBP) or the heat labile E. coli subunit B (LTXB), plus the N-terminal amino acid sequence was analyzedd. The orderly-assembled chimeric protein was confirmed by a modified $G_{M1}$-ganglioside ELISA using antibodies to adhesin. The results indicate that the purified chimeric protein was an Adhesin/LTXA2B protein containing UPEC adhesin and the $G_{M1}$-ganglioside binding activity of LTXB. thisstudy also demonstrate that peroral administration of this chimeric immunogen in mice elicited high level of secretory IgA (sIgA) and serum IgG Abs to the UPEC adhesin. The results suggest that the genetically linked LTXA2B acts as a useful mucosal adjuvant, and that adhesin/LTXA2A chimeric protein might be a potential antigen for oral immunization against UPEC.

• International Journal of Industrial Entomology and Biomaterials
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• v.8 no.2
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• pp.145-149
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• 2004

The expression of a fusion protein comprised of the B. thuringiensis crystal protein, Cry1Ac, and enhanced green fluorescent protein (EGFP) in Escherichia coli XLl-blue was examined. Three recombinant plasmids were transformed into E. coli XL1-blue and named as ProAc/Ec, MuEGFP/Ec and ProMu-EGFP/Ec, respectively. All transformants were observed by light and fluorescence microscopy at mid-log phase. The expression in E. coli transformants, ProMu-EGFP/Ec and MuEGFP/Ec, exhibited bright enough fluorescence to be observed. Furthermore, ProMu-EGFP/Ec produced fluorescent inclusions, which may have been recombinant crystals between EGFP and Cry1Ac while MuEGFP/Ec expressed soluble EGFP in cell. In SDS-PAGE, ProAc/Ec had 130 kDa crystal protein band and MuEGFP/Ec had thick 27 kDa EGFP band. However, ProMu-EGFP/Ec had about 150 kDa fusion protein band. Accordingly, these results indicated that a fusion protein between the B. thuringiensis crystal protein and a foreign protein under the lacZ promoter was successfully expressed as granular structure in E. coli. It is suggested that the E. coli expression system by N-terminal fusion of B. thuringiensis crystal protein may be useful as excellent means for fusion expression and characterization of B. thuringiensis fusion crystal protein.

• Reproductive and Developmental Biology
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• v.37 no.4
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• pp.213-217
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• 2013

The objective of this study was to investigate the effect of bacterial contamination on elapsed time after preservation on boar semen. Known numbers of Escherichia coli (E. coli) were inoculated to freshly ejaculated semen and sperm parameters such as viability, motility, agglutination, acrosome integrity and hypo-osmotic swelling test were performed during 7 days of liquid preservation. Semen samples were prepared using antibiotic free BTS extender and 4 different levels of E. coli were treated to semen with following concentrations; 3,000, 5,000, 7,000, 10,000 CFU/ml of sperms. Semen samples were preserved at $17^{\circ}C$ for 7 days in semen storage until analyzed. Aliquots were subjected to measure the sperm viability, motility and agglutination using computer assisted sperm analysis (CASA) system, acrosome integrity was performed using chlortetracycline (CTC) staining method and hypo-osmotic swelling test was performed using hypotonic solution from day 1 (day of semen collection) to 7. Detrimental effects on sperm motility and viability were observed 3 days after preservation at the level of 5,000 CFU/ml (p<0.05). Percentage of sperm abnormality was higher (p<0.05) in over 5,000 CFU/ml groups. Sperm agglutination rate was also significantly higher (p<0.05) in groups of 5,000 and 7,000 CFU/ml. The rate of acrosome reacted sperm was higher as preservation time goes in all the samples but the pattern was clearly higher among E. coli contaminated groups (p<0.05). The sperm membrane integrity in terms of hypo-osmotic test, E. coli affects little compared to other sperm parameters. The deleterious effects observed due to the bacterial contamination in semen suggest that importance of hygiene protocol to minimize the bacterial contamination during semen collection and processing.

• Journal of Environmental Health Sciences
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• v.39 no.3
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• pp.279-288
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• 2013

Objectives: Effectiveness and conditions of vapor-phase hydrogen peroxide (VPHP) system on decontamination of Geobacillus stearothermophilus(GS) spores, Escherichia coli (E.coli) and Enterobacteria phage felix01 (felix01) were determined. Methods: The VPHP system was designed to vaporize 35% (w/w) solution of hydrogen peroxide, continuously to inject and withdraw VPHP. The system and VHP 1000ED (Steris) were operated such that dehumidification and conditioning were initiated without samples in the chamber. Then the samples were loaded into and removed. Coupons (glass, anodizing, silicon, viton) with GS spores ($1{\times}10^6$ colony forming unit/mL [CFU/mL]), E.coli ($1{\times}10^7$ CFU/mL) and felix01 ($1{\times}10^7$ plaque forming unit/mL[PFU/mL]), and Biological Indicator (BI) with GS spores ($1{\times}10^6$ CFU/mL) on stainless steel coupons were used. The tested samples were sonicated and vortexed, and then were plated for enumeration, followed by incubation at $55^{\circ}C$, 24 hr for GS spores, and at $37^{\circ}C$, 24 hr for E.coli and felix01. BI analysis in broth culture was only qualitative. Results: The efficacy of the VPHP system on decontamination was almost equivalent to that of VHP 1000ED. The conditions for complete decontamination with the VPHP system was as follows: concentration; 700~450 ppm, relative humidity; approximately 55%, and temperature; $34{\sim}32^{\circ}C$. When comparing the decontamination efficiency among different kinds of coupons, glass was the most effective, however, all kinds of coupons were decontaminated completely after 60 min exposure in both systems. Conclusion: The VPHP system can be recommended as an alternative system for traditional system using ethylene oxide, formaldehyde or chlorine dioxide.

• Journal of the Korean Society of Food Science and Nutrition
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• v.40 no.8
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• pp.1195-1199
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• 2011

Inactivation of foodborne pathogenic bacteria in corn silk tea was evaluated using a microwave plasma sterilization system (MPSS). Corn silk tea was inoculated with Escherichia coli and Listeria monocytogenes, treated with an MPSS treatment, and stored at 25$^{\circ}C$ for 12 days. The one, two, and three cycles of treatment with MPSS reduced the population of E. coli by 1.14, 2.49, and 5.72 log CFU/mL, respectively, compared to that of the control. In the case of L. monocytogenes, one, two, and three cycles of MPSS treatment reduced the population by 1.93, 4.49, and 6.62 log CFU/mL, respectively. Both E. coli and L. monocytogenes were eliminated within four cycles of treatment with MPSS, and even after 12 days of storage, the bacteria were not detected. Total polyphenol content in the corn silk tea did not change much among treatments, and turbidity of the corn silk tea improved following four cycles of MPSS treatment. These results suggest that MPSS treatment can be useful for improving the microbial safety and quality of corn silk tea during storage.

• The Microorganisms and Industry
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• v.19 no.4
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• pp.14-26
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• 1993

Optimizing protein production in recombinant E. coli strains involves manipulation of genetic and environmental factors. In designing a production system, attention must be paid to gene expression efficiency, culture conditions and bioreactor configuration. Although not much emphasis was given to the physiology of host strains in this review, an understanding of the relationship between the physiology of host cell growth and the overproduction of a cloned gene protein is of primary importance to the improvement of the recombinant fermentation processes. Sometimes it is desirable to make use of gene fusion systems, e.g. protein A, polypeptide, gutathione-S-transferase, or pneumococcal murein hydrolase fusion, to facilitate protein purification.