• Journal of Microbiology and Biotechnology
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• v.17 no.2
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• pp.244-252
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• 2007

Escherichia coli has a PhoR-PhoB two-component regulatory system to detect and respond to the changes of environmental phosphate concentration. For the E. coli W3110 strain growing under phosphate-limiting condition, the changes of global gene expression levels were investigated by using DNA microarray analysis. The expression levels of some genes that are involved in phosphate metabolism were increased as phosphate became limited, whereas those of the genes involved in ribosomal protein or amino acid metabolism were decreased, owing to the stationary phase response. The upregulated genes could be divided into temporarily and permanently inducible genes by phosphate starvation. At the peak point showing the highest expression levels of the phoB and phoR genes under phosphate-limiting condition, the phoB- and/or phoR-dependent regulatory mechanisms were investigated in detail by comparing the gene expression levels among the wild-type and phoB and/or phoR mutant strains. Overall, the phoB mutation was epistatic over the phoR mutation. It was found that PhoBR and PhoB were responsible for the upregulation of the phosphonate or glycerol phosphate metabolism and high-affinity phosphate transport system, respectively. These results show the complex regulation by the PhoR-PhoB two-component regulatory system in E. coli.

• Journal of Microbiology and Biotechnology
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• v.29 no.3
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• pp.401-409
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• 2019

Heat-resistant microbial hosts are required for bioprocess development using high cell density cultivations at the industrial scale. We report that the thermotolerance of Escherichia coli can be enhanced by overexpressing ybeD, which was known to encode a hypothetical protein of unknown function. In the wild-type E. coli BL21(DE3), ybeD transcription level increased over five-fold when temperature was increased from $37^{\circ}C$ to either $42^{\circ}C$ or $46^{\circ}C$. To study the function of ybeD, a deletion strain and an overexpression strain were constructed. At $46^{\circ}C$, in comparison to the wild type, the ybeD-deletion reduced cell growth half-fold, and the ybeD-overexpression promoted cell growth over two-fold. The growth enhancement by ybeD-overexpression was much more pronounced at $46^{\circ}C$ than $37^{\circ}C$. The ybeD-overexpression was also effective in other E. coli strains of MG1655, W3110, DH10B, and BW25113. These findings reveal that ybeD gene plays an important role in enduring high-temperature stress, and that ybeD-overexpression can be a prospective strategy to develop thermotolerant microbial hosts.

• Journal of Microbiology and Biotechnology
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• v.19 no.10
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• pp.1127-1134
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• 2009

Escherichia coli excretes acetate during aerobic growth on glycolytic carbon sources, which has been explained as an overflow metabolism when the carbon flux into the cell exceeds the capacity of central metabolic pathways. Nonacetogenic growth of E. coli on gluconeogenic carbon sources like succinate or in carbon-limited slow growth conditions is believed an evidence for the explanation. However, we found that a strain defected in the acs (acetyl Co-A synthetase) gene, the product of which is involved in scavenging acetate, accumulated acetate even in succinate medium and in carbon-limited low growth rate condition, where as its isogenic parental strain did not. The acs promoter was inducible in noncatabolite repression condition, whereas the expression of the ackA-pta operon encoding acetate kinase and phosphotransacetylase for acetate synthesis was constitutive. Results in this study suggest that E. coli excretes and scavenges acetate simultaneously in the carbon-limited low growth condition and in nonacetogenic carbon source, and the activity of the acetate consumption pathway directly affects the accumulation level of acetate in the culture broth.

• Biotechnology and Bioprocess Engineering:BBE
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• v.9 no.5
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• pp.389-392
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• 2004

In order to produce a harmful protein more efficiently, this expression cassette, dubbed pCol-MICT, is directed by the colicin promoter, and was constructed by the insertion of a $rrnBT_1T_2$ fragment of pEXP7, and a MxelnteinCBD fragment of pTXB3, into pSH375. To test whether harmful proteins, including proteolytic enzymes, could be effectively produced by this cassette, the carboxypeptidase (CPase) Taq gene was inserted into the pCol-MICT cassette to yield pCol-CPase Taq-MICT. E coli W3l 10 tells harboring pCol-CPase Taq-MICT produced a large quantity of this enzyme, as much as 47.2 mg of purified from per liter of culture, when cultured in the presence of mitomycin C ($0.4{\mu}g/mL$). This indicates that the colicin promoter-controlled E, coli expression cassette was able to produce almost 8 times of protein than the conventional tar promoter-based system, and that this cassette may be useful in the Synthesis of other harmful proteins.

• Journal of Microbiology and Biotechnology
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• v.16 no.7
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• pp.1125-1131
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• 2006

A kan'::luxCDABE fusion strain that was both highly bioluminescent and responsive to benzoic acid was constructed by transforming E. coli strain W3110 with the plasmid pUCDK, which was constructed by digesting and removing the 7-kb KpnI fragment from the promoterless luxCDABE plasmid pUCD615. Experiments using buffered media showed that this induction was dependent on the pH of the media, which influences the degree of benzoic acid protonation, and the expression levels seen are likely due to acidification of the cytoplasm by uncoupling of benzoic acid. Consequently, the sensitivity of this strain for benzoic acid was increased by nearly 20-fold when the pH was shifted from 8.0 to 6.5. Benzoic acid derivatives and several phenolics also resulted in significantly increased bioluminescent signals. Although these compounds are known to damage membranes and induce the heat-shock response within E. coli, bacterial strains harboring mutations in the fadR and rpoH genes, which are responsible for fatty acid biosynthesis during membrane stress and induction of the heat-shock response, respectively, showed that these mutations had no effect on the responses observed.

• Journal of Life Science
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• v.22 no.3
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• pp.417-427
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• 2012

We compared the transcriptome in response to propanol stress in wild-type and propanol-resistant mutant Escherichia coli using the DNA microarray technique. The correlation value of RNA expression between the propanol-treated wild type and the untreated-one was about 0.949, and 50 genes were differentially expressed by more than twofold in both samples. The correlation value of RNA expression between the propanol-treated mutant and the untreated one was about 0.951, and 71 genes in two samples showed differential expression patterns. However, the values between the wild type and mutant, regardless of propanol addition, were 0.974-0.992 and only 1-2 genes were differentially expressed in the two strains. The representative characteristics among differentially expressed genes in W3110 or P19 treated with propanol compared to untreated samples were up-regulation of hest shock response genes and down-regulation of genes relating to ribosome biosynthesis. In addition, many genes were regulated by transcription regulation factors such as ArcA, CRP, FNR, H-NS, GatR, or PurR and overexpressed by sigma factor RpoH. We confirmed that RpoH mediated an important host defense function in propanol stress in E. coli W3110 and P19 by comparison of cell growth rate among the wild type, rpoH disruptant mutant, and rpoH-complemented strain.

• Biotechnology and Bioprocess Engineering:BBE
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• v.9 no.5
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• pp.374-378
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• 2004

The effects of various environmental factors such as pH (5, 6, 7, 8 and 9), temperature (30, 37 and $40^{\circ}C$) and rotational speed (150, 200 and 250 rpm) on the growth and the hepatitis B core antigen (HBcAg) production of Escherichia coli W3110IQ were examined in the present Study. The highest growth rate is achieved at pH 7, $37^{\circ}C$ and at a rotational Speed of 250 rpm which is 0.927 $h^{-1}$. The effect of pH on cell growth is more substantial compared to other parameters; it recorded a $123\%$ different between the highest growth rate (0.927 $h^{-1}$) at pH 7 and lowest growth at pH 5. The highest protein yield is achieved at pH 9, rotational speed of 250 rpm and $40^{\circ}C$. The yield of protein at pH 7 is $154\%$ higher compared to the lowest yield achieved at pH 5. There is about $28\%$ different of the protein yield for the E. coli cultivated at 250 rpm compared to that at 150 rpm which has the lowest HBcAg yield. The yield of protein at $40^{\circ}C$ is $38\%$ higher compared to the lowest yield achieved at $30^{\circ}C$.

• KSBB Journal
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• v.21 no.2
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• pp.90-95
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• 2006

Escherichia coli harboring pAC-LYCO4 and pDdxs was used for lycopene production. Three wild type strains of E. coli OW1, MG1655, and W3110 were compared with DH5${\alpha}$ used before for lycopene production. Lycopene productivity of E. coli MG1655 was similar to DH5${\alpha}$ and the highest among those wild type strain. Therefore, MG1655 strain was used for random transposon and NTG mutagenesis to increase lycopene productivity. Through transposon mutation, five transposon mutants with increased lycopene productivity were obtained. It was found that genes knocked out by transposon insertion were treB in Tn1 mutant, B2436 in Tn2 mutant, and rfaH in Tn3, 4, and 5 mutants. Lycopene productivity was the highest in Tn4 mutant among the Tn mutants, which was 6-fold and 8-fold higher in lycopene concentration and content, respectively, in comparison with those obtained with wild type strain. NTG4 mutant was acquired with NTG mutation. The highest lycopene productivity of 6 mg/L and 4 mg/g DCW was obtained from the NTG4 mutant when arabinose of 0.013 mM was added for induction of dxs, rate-limiting gene of MEP pathway. The lycopene productivity of NTG4 mutant was increased 18-fold and 12-fold in lycopene concentration and content, respectively when comparing with the wild type strain.