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Significantly improved solvent tolerance of Escherichia coli by global transcription machinery engineering.

Zhang F, Qian X, Si H, Xu G, Han R, Ni Y - Microb. Cell Fact. (2015)

Bottom Line: The rpoD mutant contains three amino-acid substitutes and a stop-codon mutation, resulting a truncated sequence containing regions σ(1.1) and σ(1.2).Our results show that several genes (gapA, sdhB, pepB and dppA) play critical roles in enhanced solvent tolerance of E. coli, mainly involving in maintaining higher intracellular energy level under solvent stress.Global transcription machinery engineering is therefore a feasible and efficient approach for engineering strain with enhanced OST-phenotype.

View Article: PubMed Central - PubMed

Affiliation: The Key Laboratory of Industrial Biotechnology, Ministry of Education, School of Biotechnology, Jiangnan University, 214122, Wuxi, Jiangsu, China. zf23.32@163.com.

ABSTRACT

Background: Escherichia coli has emerged as a promising platform microorganism to produce biofuels and fine chemicals of industrial interests. Certain obstacles however remain to be overcome, among which organic-solvent tolerance is a crucial one.

Results: We used global transcription machinery engineering (gTME) to improve the organic-solvent tolerance (OST) of E. coli JM109. A mutant library of σ(70) encoded by rpoD was screened under cyclohexane pressure. E. coli JM109 strain harboring σ(70) mutant C9 was identified with capability of tolerating 69 % cyclohexane. The rpoD mutant contains three amino-acid substitutes and a stop-codon mutation, resulting a truncated sequence containing regions σ(1.1) and σ(1.2). Total protein difference produced by E. coli JM109 strain harboring C9 was examined with 2D-PAGE, and 204 high-abundant proteins showed over twofold variation under different solvent stress.

Conclusions: Our results show that several genes (gapA, sdhB, pepB and dppA) play critical roles in enhanced solvent tolerance of E. coli, mainly involving in maintaining higher intracellular energy level under solvent stress. Global transcription machinery engineering is therefore a feasible and efficient approach for engineering strain with enhanced OST-phenotype.

No MeSH data available.


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2D-PAGE of total proteins of E. coli JM109/pHACM-rpoDC9 under different solvent treatments. a Without solvent; b with 38 % (v/v) cyclohexane. For each treatment condition, 2-DE experiment was conducted in triplicates
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Fig3: 2D-PAGE of total proteins of E. coli JM109/pHACM-rpoDC9 under different solvent treatments. a Without solvent; b with 38 % (v/v) cyclohexane. For each treatment condition, 2-DE experiment was conducted in triplicates

Mentions: Two-DE, a powerful protein separation technique to illustrate proteins associated with certain phenotype, was used to investigate the proteomics of E. coli strains harboring C9 mutant when grown with or without cyclohexane. 2-DE analysis of WT strain (without cyclohexane) was also conducted as control. Our results show that there was no obvious difference between WT and C9 strain in the absence of cyclohexane (Additional file 1: Figure S1). Compared with control (C9 without solvent), 204 high-abundant proteins in C9 strain showed over twofold difference in the presence of 38 % cyclohexane (Fig. 3).Fig. 3


Significantly improved solvent tolerance of Escherichia coli by global transcription machinery engineering.

Zhang F, Qian X, Si H, Xu G, Han R, Ni Y - Microb. Cell Fact. (2015)

2D-PAGE of total proteins of E. coli JM109/pHACM-rpoDC9 under different solvent treatments. a Without solvent; b with 38 % (v/v) cyclohexane. For each treatment condition, 2-DE experiment was conducted in triplicates
© Copyright Policy - OpenAccess
Related In: Results  -  Collection

License 1 - License 2
Show All Figures
getmorefigures.php?uid=PMC4635540&req=5

Fig3: 2D-PAGE of total proteins of E. coli JM109/pHACM-rpoDC9 under different solvent treatments. a Without solvent; b with 38 % (v/v) cyclohexane. For each treatment condition, 2-DE experiment was conducted in triplicates
Mentions: Two-DE, a powerful protein separation technique to illustrate proteins associated with certain phenotype, was used to investigate the proteomics of E. coli strains harboring C9 mutant when grown with or without cyclohexane. 2-DE analysis of WT strain (without cyclohexane) was also conducted as control. Our results show that there was no obvious difference between WT and C9 strain in the absence of cyclohexane (Additional file 1: Figure S1). Compared with control (C9 without solvent), 204 high-abundant proteins in C9 strain showed over twofold difference in the presence of 38 % cyclohexane (Fig. 3).Fig. 3

Bottom Line: The rpoD mutant contains three amino-acid substitutes and a stop-codon mutation, resulting a truncated sequence containing regions σ(1.1) and σ(1.2).Our results show that several genes (gapA, sdhB, pepB and dppA) play critical roles in enhanced solvent tolerance of E. coli, mainly involving in maintaining higher intracellular energy level under solvent stress.Global transcription machinery engineering is therefore a feasible and efficient approach for engineering strain with enhanced OST-phenotype.

View Article: PubMed Central - PubMed

Affiliation: The Key Laboratory of Industrial Biotechnology, Ministry of Education, School of Biotechnology, Jiangnan University, 214122, Wuxi, Jiangsu, China. zf23.32@163.com.

ABSTRACT

Background: Escherichia coli has emerged as a promising platform microorganism to produce biofuels and fine chemicals of industrial interests. Certain obstacles however remain to be overcome, among which organic-solvent tolerance is a crucial one.

Results: We used global transcription machinery engineering (gTME) to improve the organic-solvent tolerance (OST) of E. coli JM109. A mutant library of σ(70) encoded by rpoD was screened under cyclohexane pressure. E. coli JM109 strain harboring σ(70) mutant C9 was identified with capability of tolerating 69 % cyclohexane. The rpoD mutant contains three amino-acid substitutes and a stop-codon mutation, resulting a truncated sequence containing regions σ(1.1) and σ(1.2). Total protein difference produced by E. coli JM109 strain harboring C9 was examined with 2D-PAGE, and 204 high-abundant proteins showed over twofold variation under different solvent stress.

Conclusions: Our results show that several genes (gapA, sdhB, pepB and dppA) play critical roles in enhanced solvent tolerance of E. coli, mainly involving in maintaining higher intracellular energy level under solvent stress. Global transcription machinery engineering is therefore a feasible and efficient approach for engineering strain with enhanced OST-phenotype.

No MeSH data available.


Related in: MedlinePlus