• BMB Reports
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• v.30 no.1
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• pp.80-84
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• 1997

The synthesis and secretion of human angiogenin in E. coli by the natural leader sequence has been studied. We constructed a recombinant plasmid containing human angiogenin cDNA which encompassed all the coding region including leader sequence required for secretion. The recombinant plasmid was introduced into a suitable E. coli host. The angiogenin was detected in the culture medium and periplasm upon the induction of gene expression. The molecular weight of the secreted angiogenin was identical to that of authentic angiogenin purfied from human plasma when estimated by SDS-PAGE and immunoblotting. showing that the natural leader sequence was recognized and processed by the secretion machinery of E. coli. The angiogenin concentration in the culture medium reached a maximum within 2 h when expressed at $37^{\circ}C$ with 0.02~2 mM IPTG. In contrast, the expression level increased gradually over time up to 11 h at $23^{\circ}C$ with 0.002~2 mM IPTG and at $37^{\circ}C$ with 0.002 mM IPTG.

• Journal of Industrial Technology
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• v.28 no.B
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• pp.59-63
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• 2008

Rapid production of therapeutic proteins such as angiostatin and endostatin angiogenic inhibititors has been highly demanded for cancer treatment. In this regard, recombinant human angiostatin and endostatin were successfully expressed as soluble forms by maltose binding protein (MBP)-mediated fusion expression in Escherichia coli. PCR amplified, angiostatin and endostatin genes from human placenta cDNA library were inserted into an expression vector pMAL-c2e to construct prokaryotic expression vectors, pMAL-c2e/AS and pMAL-c2e/ES, respectively. Recombinant angiostatin and endostatin were efficiently expressed in E. coli origami (DE3) after IPTG induction and protein expression were confirmed by SDS-PAGE analyses. The expressed recombinant proteins were purified near homogenity using an amylose affinty column chromatography. In contrast that previous E. coli expressions were all insoluble, our results first time demonstrated that MBP fused human angiostatin and endostatin were soluble in E. coli.

• Food Science and Biotechnology
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• v.15 no.5
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• pp.817-819
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• 2006

8-Quinolinol and other quinolinol derivatives were evaluated with regard to their growth-inhibitory effects against intestinal bacteria, using the paper disk-agar diffusion method. The observed growth responses varied according to the chemicals and dosages used, as well as the bacterial species tested. 8-Quinolinol showed a significant inhibitory effect against Clostridium difficile, C. perfringens, and Escherichia coli, at 5, 2, 1, and 0.5 mg/disk, and also exhibited a very strong inhibitory effect at 0.25 mg/disk. At low concentrations, 8-quinolinol had strong inhibitory effects against C. perfringens at 0.1 and 0.05 mg/disk; 8-quinolinol also manifested a moderate inhibitory effect against C. perfringens at 0.025 mg/disk. Furthermore, 8-quinolinol revealed moderate and weak growth inhibition against C. difficile and E. coli at concentrations of 0.1 and 0.05 mg/disk, respectively, but 2-quinolinol, 4-quinolinol, and 6-quinolinol evidenced no growth inhibition against B. bifidum, B. longum, C. difficile, C. perfringens, E. coli, or L. casei. The inhibitory effects of 8-quinolinol against C. difficile, C. perfringens, and E. coli lead to its consideration as a possible therapeutic modality for the treatment of diseases associated with harmful intestinal bacteria.

• Journal of Food Hygiene and Safety
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• v.13 no.1
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• pp.1-5
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• 1998

This study was carried out to measure the antibacterial effect of lactoperoxidase/ thiocyanate/hydrogen peroxide system (LP system) against E. coli 0157:H7. When the initial inoculum levels (($10^{2},\;10^{4},\;10^{7}cfu/ml$), concentration of LP (10 ppm, 20 ppm, 30 ppm), culture media (TSB-YE, UHT milk, raw milk) and storage temperatures ($5^{\circ}C,\;10^{\circ}C,$15$^{\circ}C$) were set up differently for the experiment and the antibacterial effect was compared, the highest antibacterial effect of LP system was shown at $10^{2}cfu/ml$ of initial inoculum level, 10 ppm of LP concentration and $5^{\circ}C$ of incubation temperature.

• Journal of Microbiology and Biotechnology
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• v.14 no.1
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• pp.216-219
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• 2004

The effect of culture temperature on the production of soluble form of B. macerans cyclodextrin glucanotransferase (CGTase) in recombinant E. coli was investigated. E. coli cell was cotransformed with two plasmids (pTCGT1 and pGroll) in which the cgt and groEL/ES genes are under the control of T7 promoter and pzt-1 promoter, respectively. When tetracycline (10 ng/ml) and IPTG (l mM) were added as inducers at the early-exponential phase (2 h) and mid-exponential phase (3h), respectively, the solubilization of the inclusion body CGTase was greatly dependent on the temperature of the culture. At low culture temperature of $25^\circ{C}$, 2- or 3-fold higher activity and specific activity were obtained over $37^\circ{C}$. SDS-PAGE analysis revealed that about 62% of CGTase in the total CGTase protein was found in the soluble fraction by applying overexpression of GroEL/ES chaperone and by cultivation of E. coli at $25^\circ{C}$, whereas 33% of CGTase was detected in the soluble fraction at $37^\circ{C}$. Therefore, the expression of GroEL/ES and cultivation at $25^\circ{C}$ greatly enhanced the soluble production of CGTase in E. coli.

• KSBB Journal
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• v.29 no.4
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• pp.244-249
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• 2014

Biosynthesis pathway of medium-chain-length (MCL) polyhydroxyalkanoates (PHA) from fatty acid ${\beta}$-oxidation pathway was constructed in recombinant Escherichia coli by introducing the Pseudomonas sp. 61-3 PHA synthase gene (phaC2) and the maoC genes from Pseudomonas putida, Sinorhizobium meliloti, and Ralstonia eutropha. The metabolic link between fatty acid ${\beta}$-oxidation pathway and PHA biosynthesis pathway was constructed by MaoC, which is homologous to P. aeruginosa (R)-specific enoyl-CoA hydratase (PhaJ1). When the E. coli W3110 strains expressing the phaC2 gene and one of the maoC genes from P. putida, Sinorhizobium meliloti, and Ralstonia eutropha were cultured in LB medium containing 2 g/L of sodium decanoate as a carbon source, MCL-PHA that mainly consists of 3-hydroxyhexanoate (3HHx), 3-hydroxyoctanoate (3HO) and 3-hydroxydecanoate (3HD), was produced. The monomer composition of PHA and PHA contents varied depending on MaoC employed for the production of PHA. The highest PHA content of 18.7 wt% was achieved in recombinant E. coli W3110 expressing the phaC2 gene and the P. putida maoC gene. These results suggest that MCL-PHA biosynthesis pathway can be constructed in recombinant E. coli strains from the b-oxidation pathway by employing MaoC able to supply (R)-3-hydroxyacyl-CoA, the substrate of PHA synthase.

• BMB Reports
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• v.40 no.1
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• pp.107-112
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• 2007

To study the functioning of HSP70 in Escherichia coli, we selected NtHSP70-2 (AY372070) from among three genomic clones isolated in Nicotiana tabacum. Recombinant NtHSP70-2, containing a hexahistidine tag at the amino-terminus, was constructed, expressed in E. coli, and purified by $Ni^{2+}$ affinity chromatography and Q Sepharose Fast Flow anion exchange chromatography. The expressed fusion protein, $H_6NtHSP70$-2 (hexahistidine-tagged Nicotiana tabacum heat shock protein 70-2), maintained the stability of E. coli proteins up to 90$^{\circ}C$. Measuring the light scattering of luciferase (luc) revealed that NtHSP70-2 prevents the aggregation of luc without ATP during high-temperature stress. In a functional bioassay (1 h at 50$^{\circ}C$) for recombinant $H_6NtHSP70$-2, E. coli cells overexpressing $H_6NtHSP70$-2 survived about seven times longer than those lacking $H_6NtHSP70$-2. After 2 h at 50$^{\circ}C$, only the E. coli overexpressing $H_6NtHSP70$-2 survived under such conditions. Our NtHSP70-2 bioassays, as well as in vitro studies, strongly suggest that HSP70 confers thermo-tolerance to E. coli.

• Journal of Food Hygiene and Safety
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• v.32 no.1
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• pp.64-69
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• 2017

This study was conducted to investigate the effect of cold plasma combined with UV-C irradiation against Escherichia coli O157:H7, Salmonella enterica serovar Typhimurium, and Listeria monocytogenes on lettuce. E. coli O157:H7, S. Typhimurium, and L. monocytogenes, corresponding to approximately 5.82, 5.09, 5.65 log CFU/g, were inoculated on lettuce, respectively. Then, the lettuce was treated with cold plasma, UV-C and combination (cold plasma + UV-C), respectively. The treated lettuce was stored for 9 days at $4^{\circ}C$ for microbiological analysis and sensory evaluation. Cold plasma reduced the populations of E. coli O157:H7, S. Typhimurium, and L. monocytogenes by 0.26, 0.65, and 0.93 log CFU/g, respectively. Each microorganism were reduced by 0.87, 0.88, and 1.14 log CFU/g after UV-C treatment. And, the combined treatment that was treated by cold plasma after UV-C treatment reduced the populations of inoculated microorganisms by 1.44, 2.70, 1.62 log CFU/g, respectively. The all treatment significantly (p < 0.05) reduced the populations of all inoculated bacteria compared to untreated lettuce. UV-C combined with cold plasma was the most effective for reducing the pathogenic bacteria on lettuce, by showing log-reductions of ${\geq}2.0\;log\;CFU/g$. All treatment was not significantly different until 6 day storage compared to control group in terms of appearance, texture and overall acceptability. Therefore, the combined treatment will be an effective intervention method to control the bacteria on lettuce.

• Korean Journal of Food Science and Technology
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• v.34 no.6
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• pp.1134-1139
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• 2002

Campylobacter, Arcobacter, and Helicobacter, classified into the same rRNA superfamily VI by taxonomy, cause food-borne diseases, stomach ulcer, and gastric cancer. To detect each strain selectively from contaminated foods, PCR, multiplex-PCR, and restricion fragment length polymorphism (RFLP) were applied on Campylobacter, Arcobacter, and Helicobacter. The same PCR products could be detected using CHA primer targeted for 16S rRNA of Campylobacter, Arcobacter, and Helicobacter. To detect C. jejuni and C. coli from A. butzleri and H. pylori, pg50/pg3 primer targeted for fla A gene was used, and for A. butzleri, Arco2/Butz primer targeted for 23S rRNA was utilized. For H. pylori detection, icd1/icd2 primer targeted for isocitrate dehydrogenase gene was employed, and JEJ1/JEJ2 primer targeted for ceuE gene was effective for C. jejuni detection from the three strains. C. jejuni, C. coli could be separated from A. butzleri and H. pylori through PCR-RFLP using restriction enzyme Dde I. Such primers would be effective for detecting each strain selectively through PCR when C. jejuni, C. coli, A butzleri and H. pylori are contaminated together.

• BMB Reports
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• v.38 no.1
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• pp.97-103
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• 2005

Under the condition of expression of $\lambda$ P protein at lethal level, the oriC DNA-binding activity is significantly affected in wild-type E. coli but not in the rpl mutant. In purified system, the $\lambda$ P protein inhibits the binding of both oriC DNA and ATP to the wild-type DnaA protein but not to the rpl DnaA protein. We conclude that the $\lambda$ P protein inhibits the binding of oriC DNA and ATP to the wild-type DnaA protein, which causes the inhibition of host DNA synthesis initiation that ultimately leads to bacterial death. A possible beneficial effect of this interaction of $\lambda$ P protein with E. coli DNA initiator protein DnaA for phage DNA replication has been proposed.