Advanced search
1 file | 1.35 MB Add to list

Effect of iclR and arcA knockouts on biomass formation and metabolic fluxes in Escherichia coli K12 and its implications on understanding the metabolism of Escherichia coli BL21 (DE3)

Author
Organization
Project
Biotechnology for a sustainable economy (Bio-Economy)
Abstract
ABSTRACT: BACKGROUND: Gene expression is regulated through a complex interplay of different transcription factors (TFs) which can enhance or inhibit gene transcription. ArcA is a global regulator that regulates genes involved in different metabolic pathways, while IclR as a local regulator, controls the transcription of the glyoxylate pathway genes of the aceBAK operon. This study investigates the physiological and metabolic consequences of arcA and iclR deletions on E. coli K12 MG1655 under glucose abundant and limited conditions and compares the results with the metabolic characteristics of E. coli BL21 (DE3). RESULTS: The deletion of arcA and iclR results in an increase in the biomass yield both under glucose abundant and limited conditions, approaching the maximum theoretical yield of 0.65 c-mole/c-mole glucose under glucose abundant conditions. This can be explained by the lower flux through several CO2 producing pathways in the E. coli K12 arcAiclR double knockout strain. Due to iclR gene deletion, the glyoxylate pathway is activated resulting in a redirection of 30% of the isocitrate molecules directly to succinate and malate without CO2 production. Furthermore, a higher flux at the entrance of the TCA was noticed due to arcA gene deletion, resulting in a reduced production of acetate and less carbon loss. Under glucose limiting conditions the flux through the glyoxylate pathway is further increased in the iclR knockout strain, but this effect was not observed in the double knockout strain. Also a striking correlation between the glyoxylate flux data and the isocitrate lyase activity was observed for almost all strains under both growth conditions. Finally, similar central metabolic fluxes were observed in E. coli K12 arcAiclR compared to the industrially relevant E. coli BL21, especially with respect to the pentose pathway, the glyoxylate pathway, and the TCA fluxes. In addition, a comparison of the genome sequences of the two strains showed that BL21 possesses two mutations in the promoter region of iclR and rare codons are present in arcA, implying a lower tRNA acceptance. Both phenomena presumably result in a reduced ArcA and IclR synthesis in BL21, which contributes to the similar physiology as observed in E. coli K12 arcAiclR. CONCLUSIONS: The deletion of arcA results in a decrease of repression on transcription of TCA cycle genes under glucose abundant conditions, without significantly affecting the glyoxylate pathway activity. IclR clearly represses transcription of glyoxylate pathway genes under glucose abundance, a condition in which Crp activation is absent. Under glucose limitation, Crp is responsible for the high glyoxylate flux, but IclR still represses transcription. Finally, in E. coli BL21 ArcA and IclR are poorly expressed, explaining the similar fluxes observed compared to the arcAiclR strain
Keywords
CRA GENE KNOCKOUT, B STRAINS REL606, CONTINUOUS-CULTURE, GROWTH-RATE, TREHALOSE SYNTHESIS, MAINTENANCE ENERGY, TRANSCRIPTIONAL REGULATION, STOICHIOMETRIC MODEL, REGULATORY NETWORKS, GLUCOSE-UTILIZATION

Downloads

  • 1471-2180-11-70.pdf
    • full text
    • |
    • open access
    • |
    • PDF
    • |
    • 1.35 MB

Citation

Please use this url to cite or link to this publication:

MLA
Waegeman, Hendrik et al. “Effect of iclR and arcA Knockouts on Biomass Formation and Metabolic Fluxes in Escherichia Coli K12 and Its Implications on Understanding the Metabolism of Escherichia Coli BL21 (DE3).” BMC MICROBIOLOGY 11 (2011): n. pag. Print.
APA
Waegeman, H., Beauprez, J., Moens, H., Maertens, J., De Mey, M., Foulque-Moreno, M. R., Heijnen, J., et al. (2011). Effect of iclR and arcA knockouts on biomass formation and metabolic fluxes in Escherichia coli K12 and its implications on understanding the metabolism of Escherichia coli BL21 (DE3). BMC MICROBIOLOGY, 11.
Chicago author-date
Waegeman, Hendrik, Joeri Beauprez, Helena Moens, Jo Maertens, Marjan De Mey, Maria R Foulque-Moreno, Joseph Heijnen, Daniel Charlier, and Wim Soetaert. 2011. “Effect of iclR and arcA Knockouts on Biomass Formation and Metabolic Fluxes in Escherichia Coli K12 and Its Implications on Understanding the Metabolism of Escherichia Coli BL21 (DE3).” Bmc Microbiology 11.
Chicago author-date (all authors)
Waegeman, Hendrik, Joeri Beauprez, Helena Moens, Jo Maertens, Marjan De Mey, Maria R Foulque-Moreno, Joseph Heijnen, Daniel Charlier, and Wim Soetaert. 2011. “Effect of iclR and arcA Knockouts on Biomass Formation and Metabolic Fluxes in Escherichia Coli K12 and Its Implications on Understanding the Metabolism of Escherichia Coli BL21 (DE3).” Bmc Microbiology 11.
Vancouver
1.
Waegeman H, Beauprez J, Moens H, Maertens J, De Mey M, Foulque-Moreno MR, et al. Effect of iclR and arcA knockouts on biomass formation and metabolic fluxes in Escherichia coli K12 and its implications on understanding the metabolism of Escherichia coli BL21 (DE3). BMC MICROBIOLOGY. 2011;11.
IEEE
[1]
H. Waegeman et al., “Effect of iclR and arcA knockouts on biomass formation and metabolic fluxes in Escherichia coli K12 and its implications on understanding the metabolism of Escherichia coli BL21 (DE3),” BMC MICROBIOLOGY, vol. 11, 2011.
@article{1232510,
  abstract     = {ABSTRACT: BACKGROUND: Gene expression is regulated through a complex interplay of different transcription factors (TFs) which can enhance or inhibit gene transcription. ArcA is a global regulator that regulates genes involved in different metabolic pathways, while IclR as a local regulator, controls the transcription of the glyoxylate pathway genes of the aceBAK operon. This study investigates the physiological and metabolic consequences of arcA and iclR deletions on E. coli K12 MG1655 under glucose abundant and limited conditions and compares the results with the metabolic characteristics of E. coli BL21 (DE3). RESULTS: The deletion of arcA and iclR results in an increase in the biomass yield both under glucose abundant and limited conditions, approaching the maximum theoretical yield of 0.65 c-mole/c-mole glucose under glucose abundant conditions. This can be explained by the lower flux through several CO2 producing pathways in the E. coli K12 arcAiclR double knockout strain. Due to iclR gene deletion, the glyoxylate pathway is activated resulting in a redirection of 30% of the isocitrate molecules directly to succinate and malate without CO2 production. Furthermore, a higher flux at the entrance of the TCA was noticed due to arcA gene deletion, resulting in a reduced production of acetate and less carbon loss. Under glucose limiting conditions the flux through the glyoxylate pathway is further increased in the iclR knockout strain, but this effect was not observed in the double knockout strain. Also a striking correlation between the glyoxylate flux data and the isocitrate lyase activity was observed for almost all strains under both growth conditions. Finally, similar central metabolic fluxes were observed in E. coli K12 arcAiclR compared to the industrially relevant E. coli BL21, especially with respect to the pentose pathway, the glyoxylate pathway, and the TCA fluxes. In addition, a comparison of the genome sequences of the two strains showed that BL21 possesses two mutations in the promoter region of iclR and rare codons are present in arcA, implying a lower tRNA acceptance. Both phenomena presumably result in a reduced ArcA and IclR synthesis in BL21, which contributes to the similar physiology as observed in E. coli K12 arcAiclR. CONCLUSIONS: The deletion of arcA results in a decrease of repression on transcription of TCA cycle genes under glucose abundant conditions, without significantly affecting the glyoxylate pathway activity. IclR clearly represses transcription of glyoxylate pathway genes under glucose abundance, a condition in which Crp activation is absent. Under glucose limitation, Crp is responsible for the high glyoxylate flux, but IclR still represses transcription. Finally, in E. coli BL21 ArcA and IclR are poorly expressed, explaining the similar fluxes observed compared to the arcAiclR strain},
  articleno    = {70},
  author       = {Waegeman, Hendrik and Beauprez, Joeri and Moens, Helena and Maertens, Jo and De Mey, Marjan and Foulque-Moreno, Maria R  and Heijnen, Joseph and Charlier, Daniel and Soetaert, Wim},
  issn         = {1471-2180},
  journal      = {BMC MICROBIOLOGY},
  keywords     = {CRA GENE KNOCKOUT,B STRAINS REL606,CONTINUOUS-CULTURE,GROWTH-RATE,TREHALOSE SYNTHESIS,MAINTENANCE ENERGY,TRANSCRIPTIONAL REGULATION,STOICHIOMETRIC MODEL,REGULATORY NETWORKS,GLUCOSE-UTILIZATION},
  language     = {eng},
  pages        = {17},
  title        = {Effect of iclR and arcA knockouts on biomass formation and metabolic fluxes in Escherichia coli K12 and its implications on understanding the metabolism of Escherichia coli BL21 (DE3)},
  url          = {http://dx.doi.org/10.1186/1471-2180-11-70},
  volume       = {11},
  year         = {2011},
}

Altmetric
View in Altmetric
Web of Science
Times cited: