• Microbiology and Biotechnology Letters
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• v.17 no.4
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• pp.379-384
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• 1989

Glutamine production from glutamate was carried out using glutamine synthetase from E. coli K-12 pgln 6 and baker's yeast, which supplies ATP into the reaction system through alcohol fermentation, simultaneously. With whole cells of E. coli K-12 pgln 6 as an enzyme source of glutamine synthetase, 11.8 g/ι of glutamine produced after 18-h incubation (60％ yield based on a substrate, glutamate). Using the partially purified glutamine synthetase, 19.8 git of glutamine was produced after 5-h incubation. This amount of glutamine was correspond to 90％ yield, based on substrate, glutamate.

• Journal of Microbiology and Biotechnology
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• v.22 no.4
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• pp.470-478
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• 2012

Recent genome comparisons of E. coli B and K-12 strains have indicated that the makeup of the cell envelopes in these two strains is quite different. Therefore, we analyzed and compared the envelope proteomes of E. coli BL21(DE3) and MG1655. A total of 165 protein spots, including 62 nonredundant proteins, were unambiguously identified by two-dimensional gel electrophoresis and mass spectrometry. Of these, 43 proteins were conserved between the two strains, whereas 4 and 16 strain-specific proteins were identified only in E. coli BL21(DE3) and MG1655, respectively. Additionally, 24 proteins showed more than 2-fold differences in intensities between the B and K-12 strains. The reference envelope proteome maps showed that E. coli envelope mainly contained channel proteins and lipoproteins. Interesting proteomic observations between the two strains were as follows: (i) B produced more OmpF porin with a larger pore size than K-12, indicating an increase in the membrane permeability; (ii) B produced higher amounts of lipoproteins, which facilitates the assembly of outer membrane ${\beta}$-barrel proteins; and (iii) motility- (FliC) and chemotaxis-related proteins (CheA and CheW) were detected only in K-12, which showed that E. coli B is restricted with regard to migration under unfavorable conditions. These differences may influence the permeability and integrity of the cell envelope, showing that E. coli B may be more susceptible than K-12 to certain stress conditions. Thus, these findings suggest that E. coli K-12 and its derivatives will be more favorable strains in certain biotechnological applications, such as cell surface display or membrane engineering studies.

• Korean Journal of Microbiology
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• v.32 no.1
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• pp.40-46
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• 1994

The pcb genes of Pseudomonas sp. DJ-12 coded for the catabolism of polychlorinated biphenyl (PCBs) and biphenyl. The products of the pcbCD genes were 2,3-dihydroxy-4'-chlorobiphenyl dioxygenase and meta-cleavage product (MCP) hydrolase, which acted on degradation of 2,3-dihydroxy-4'-chlorobiphenyl to 4-chlorobenzoate. The pcbCD genes were cloned in E. coli XLl-Blue, and then the pcbD gene was further subcloned. As a metabolite transformed from 2,3-dihydroxybiphenyl by the cloned cell of E coli CU103, benzoate was detected by the resting cell assay. The enzyme activities of 2,3-dihydroxybiphenyl dioxygease and MCP hydrolase produced in the cloned cells E. coli CU103 and CU105 were about 17 and 3 times higher than those of Pseudomonas sp. DJ-12, respectively.

• Biomedical Science Letters
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• v.17 no.1
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• pp.7-12
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• 2011

Free-living amoebae ingest several kinds of bacteria. In other words, the bacteria can survive within free-living amoeba. To determine how Escherichia coli K1 isolate causing neonatal encephalitis and non-pathogenic K12 interact with free-living amoebae, e.g., Acanthamoeba castellanii (T1), A. astronyxis (T7), Naegleria fowleri, association, invasion and survival assays were performed. To understand pathogenicity of free-living amoebae, in vitro cytotoxicity assay were performed using murine macrophages. T1 destroyed macrophages about 64% but T7 did very few target cells. On the other hand, N. fowleri which needed other growth conditions rather than Acanthamoeba destroyed more than T1 as shown by lactate dehydrogenase (LDH) release assay. In association assays for E. coli binding to amoebae, the T7 exhibited significantly higher association with E. coli, compared with the T1 isolates (P<0.01). Interestingly, N. fowleri exhibited similar percentages of association as T1. Once E. coli bacteria attach or associate with free-living amoeba, they can penetrate into the amoebae. In invasion assays, the K1 (0.67%) within T1 was observed compared with K12 (0%). E. coli K1 and K12 exhibited high association with N. fowleri and bacterial CFU. To determine the fate of E. coli in long-term survival within free-living amoebae, intracellular survival assays were performed by incubating E. coli with free-living amoebae in PBS for 24 h. Intracellular E. coli K1 within T1 (2.5%) and T7 (1.8%) were recovered and grown, while K12 were not found. N. fowleri was not invaded and here it was not recovered.

• Microbiology and Biotechnology Letters
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• v.50 no.3
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• pp.328-336
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• 2022

Aptamers are short, chemically synthesized, single-stranded DNA or RNA oligonucleotides that fold into unique three-dimensional structures. In this study, we aim to determine the antibiofilm activity and binding specificity of the six polyclonal DNA aptamers (S15K3, S15K4, S15K6, S15K13, S15K15, and S15K20) on Staphylococcus aureus BPA-12 and Escherichia coli EPEC 4. Aptamer S15K6 showed the highest percentage of antibiofilm activity against S. aureus BPA-12 (37.4%) as shown by the lowest OD570 value of 0.313. Aptamer S15K20 showed the highest percentage of antibiofilm activity against E. coli EPEC 4 (15.4%) as shown by the lowest OD570 value of 0.515. Aptamers S15K13 and S15K20 showed antibiofilm activities against both S. aureus BPA-12 and E. coli EPEC4, and thus potentially have broad reactivity. Furthermore, based on the binding capacity and Kd values from our previous study, the binding specificity assay of selected polyclonal DNA aptamers (S15K3 and S15K15) against S. aureus BPA-12, E. coli EPEC 4, S. aureus BPA-6, S. agalactiae, E. coli MHA-6, and Listeria monocytogenes were performed using qPCR. Aptamers S15K3 and S15K15 showed specific binding to S. aureus BPA-12, E. coli EPEC 4, S. aureus BPA-6, and S. agalactiae, but could not bind to E. coli MHA-6 and L. monocytogenes. Therefore, this study showed that the polyclonal DNA aptamers have antibiofilm activity and were able to bind to S. aureus BPA-12 and E. coli EPEC 4 bacteria.

• Environmental Engineering Research
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• v.14 no.1
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• pp.63-67
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• 2009

Escherichia coli K-12 (E. coli K-12) is a representative indicator globally used for distinguishing and monitoring dynamic fates of pathogenic microorganisms in the environment. This study investigated how to most critically quantify lacZ ($\beta$-galactosidase) gene in E. coli K-12 by two different real-time polymerase chain reaction (real-time PCR) in association with three different DNA extraction practices. Three DNA extractions, i.e., sodium dodecyl sulfate (SDS)/proteinase K, magnetic beads and guanidium thiocyanate (GTC)/silica matrix were each compared for extracting total genomic DNA from E. coli K-12. Among them, GTC/silica matrix and magnetic beads beating similarly worked out to have the highest (22-23 ng/${\mu}L$) concentration of DNA extracted, but employing SDS/proteinase K had the lowest (10 ng/${\mu}L$) concentration of DNA retrieved. There were no significant differences in the quantification of the copy numbers of lacZ gene between SYBR Green I qPCR and QProbe-qPCR. However, SYBR Green I qPCR obtained somewhat higher copy number as $1{\times}10^8$ copies. It was decided that GTC/silica matrix extraction or magnetic beads beating in combination with SYBR Green I qPCR can be preferably applied for more effectively quantifying specific gene from a pure culture of microorganism.

• Korean Journal of Veterinary Research
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• v.28 no.1
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• pp.59-65
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• 1988

This paper deals with the 0 groups of citrate utilizing variants of Escherichia coli ($Cit^+$ E. coli) isolated from cattle, the production of colicin, hemolysin, K99 antigen, heat stable enterotoxin, and the isolation of plasmid DNA. Among 42 $Cit^+$ E. coli, 12 strains were 020, 9 strains 08, 5 strains 045, 3 strains 0115, 1 strain 064, 1 strain 0139 and remaining strains(11) were untypable. Thirty-nine(81.3%) out of 48 $Cit^+$ E. coli were produced colicin and 13(27.0%) were produced hemolysin. Of 12 $Cit^+$ E. coli bearing K99 antigen, 6(50.0%) were produced heat stable enterotoxin. In gel electrophoresis for the isolation of plasmid DNA, the number of plasmids varied from 1 to 7 in 10 $Cit^+$ E. coli. It's molecular weight ranged from 2 to 50 Mdalton, and 50 Mdalton plasmid was commonly existed in all strains.

• Journal of Microbiology and Biotechnology
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• v.25 no.11
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• pp.1835-1841
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• 2015

A cell surface display system for heterologous expression of the multifunctional cellulase, CelEx-BR12, in Escherichia coli was developed using truncated E. coli outer membrane protein C (OmpC) as an anchor motif. Cell surface expression of CelEx-BR12 cellulase in E. coli harboring OmpC-fused CelEx-BR12, designated MC4100 (pTOCBR12), was confirmed by fluorescence-activated cell sorting and analysis of outer membrane fractions by western blotting, which verified the expected molecular mass of OmpC-fused CelEx-BR12 (~72 kDa). Functional evidence for exocellulase activity was provided by enzymatic assays of whole cells and outer membrane protein fractions from E. coli MC4100 (pTOCBR12). The stability of E. coli MC4100 (pTOCBR12) cellulase activity was tested by carrying out repeated reaction cycles, which demonstrated the reusability of recombinant cells. Finally, we showed that recombinant E. coli cells displaying the CelEx-BR12 enzyme on the cell surface were capable of growth using carboxymethyl cellulose as the sole carbon source.

• Journal of Ginseng Research
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• v.7 no.2
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• pp.102-107
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• 1983

The effects of saponin, one of major components (Panax ginseng C.A. Meyer), on the growth of E. coli K-12 and the formation of siderphore was observed The following results were obtained. 1. When E. coli was grown on medium containing 1${\times}$10-5%-11${\times}$10-1% of the saponin, the rate of growth was stimulated at 10-1% of the saponin significantly compared to that of control. 2. When E. coli K-12 was grown on medium containing 1${\times}$10-1% of the saponin, the amount of siderphore was two times as much as the control. 3. The growth of E. coli was observed to be dependent on the concentration of siderophore when siderophore was added to medium. 4. The effect of saponin on the formation of siderophore in vitro was observed to reach maximum at 1${\times}$10-3% of the saponin. Such results suggest that the growth rate of E. coli K-12 could be enhanced by ginseng saponin fraction through stimulation of siderphore formation. We have described the fast growth of E. coli, K-12 and B. subtilis, rapid uptake of 14C-glucose, and high level of other metabolites such as lipids and proteins of E. coli, and B. subtilis in medium containing saponing fraction compared to that of microorganisms without saponin fraction.1∼3Such differences were claimed to be due to rapid uptake of 14C-glucose by widened periplasmic region throught unknown mechanism in the prescence of saponin fraction in medium3 and have raised a question whether there is another possible factor, siderophore4(Greek for iron bears), since microorganisms must secure a sufficient amount of iron for normal growth. These are known to be synthesized by the cells under iron-deficient condition and in most case, excreted into the medium5, where they can complex and solubilize any iron present there. It is generally believed that these complexes are then taken into the cells presumably by specific transport systems, thus providing iron for cell metabolism. Within the group of enteric bacteria, only three species (E. coli, S. typhimurium, and A. aerogense) have, so far, been studied in a ny detail. The main iron-binding compound produced by these species is enterochelin, and its role in iron transport is now well established. And biosynthesis of enterochelin from 2, 3- dihydroxybenzoate and serine in the prescence of magnesium ions and ATP was reported6. 2, 3-dihydroxybenzoate was also shown to involve isochorismate and 2, 3-dihydro-2, 3-dihydroxybenzoate as intermediate.7∼11 The present paper deals with the effect of ginseng saponin fraction on growth, the level of enterochelin formation in vivo and the conversion of 2, 3-dihydroxybenzoate and serine into entrochelin in vitro, and entrochelin obtained on the growth in relation to possible explanation of ginseng saponin fraction on the rapid growth of E. coli, K-12.

• Journal of Microbiology and Biotechnology
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• v.27 no.11
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• pp.2028-2036
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• 2017

The Escherichia coli K-12 and B strains are among the most frequently used bacterial hosts for scientific research and biotechnological applications. However, omics analyses have revealed that E. coli K-12 and B exhibit notably different genotypic and phenotypic attributes, even though they were derived from the same ancestor. In a previous study, we identified a limited number of proteins from the two strains using two-dimensional gel electrophoresis and tandem mass spectrometry (MS/MS). In this study, an in-depth analysis of the physiological behavior of the E. coli K-12 and B strains at the proteomic level was performed using six-plex isobaric tandem mass tag-based quantitative MS. Additionally, the best lysis buffer for increasing the efficiency of protein extraction was selected from three tested buffers prior to the quantitative proteomic analysis. This study identifies the largest number of proteins in the two E. coli strains reported to date and is the first to show the dynamics of these proteins. Notable differences in proteins associated with key cellular properties, including some metabolic pathways, the biosynthesis and degradation of amino acids, membrane integrity, cellular tolerance, and motility, were found between the two representative strains. Compared with previous studies, these proteomic results provide a more holistic view of the overall state of E. coli cells based on a single proteomic study and reveal significant insights into why the two strains show distinct phenotypes. Additionally, the resulting data provide in-depth information that will help fine-tune processes in the future.