Effect of <it>iclR </it>and <it>arcA </it>knockouts on biomass formation and metabolic fluxes in <it>Escherichia </it><it>coli </it>K12 and its implications on understanding the metabolism of <it>Escherichia coli </it>BL21 (DE3)
<p>Abstract</p> <p>Background</p> <p>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 pathwa...
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| Format: | Article |
| Language: | English |
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BMC
2011-04-01
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| Series: | BMC Microbiology |
| Online Access: | http://www.biomedcentral.com/1471-2180/11/70 |
| _version_ | 1811282435982229504 |
|---|---|
| author | Charlier Daniel Heijnen Joseph J Foulquié-Moreno Maria R De Mey Marjan Maertens Jo Moens Helena Beauprez Joeri Waegeman Hendrik Soetaert Wim |
| author_facet | Charlier Daniel Heijnen Joseph J Foulquié-Moreno Maria R De Mey Marjan Maertens Jo Moens Helena Beauprez Joeri Waegeman Hendrik Soetaert Wim |
| author_sort | Charlier Daniel |
| collection | DOAJ |
| description | <p>Abstract</p> <p>Background</p> <p>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 <it>aceBAK </it>operon. This study investigates the physiological and metabolic consequences of <it>arcA </it>and <it>iclR </it>deletions on <it>E. coli </it>K12 MG1655 under glucose abundant and limiting conditions and compares the results with the metabolic characteristics of <it>E. coli </it>BL21 (DE3).</p> <p>Results</p> <p>The deletion of <it>arcA </it>and <it>iclR </it>results in an increase in the biomass yield both under glucose abundant and limiting 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 CO<sub>2 </sub>producing pathways in the <it>E. coli </it>K12 Δ<it>arcA</it>Δ<it>iclR </it>double knockout strain. Due to <it>iclR </it>gene deletion, the glyoxylate pathway is activated resulting in a redirection of 30% of the isocitrate molecules directly to succinate and malate without CO<sub>2 </sub>production. Furthermore, a higher flux at the entrance of the TCA was noticed due to <it>arcA </it>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 Δ<it>iclR </it>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 and under both growth conditions, illustrating the transcriptional control of this pathway. Finally, similar central metabolic fluxes were observed in <it>E. coli </it>K12 Δ<it>arcA </it>Δ<it>iclR </it>compared to the industrially relevant <it>E. coli </it>BL21 (DE3), 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 <it>iclR </it>and rare codons are present in <it>arcA </it>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 <it>E. coli </it>K12 Δ<it>arcA</it>Δ<it>iclR</it>.</p> <p>Conclusions</p> <p>The deletion of <it>arcA </it>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 <it>E. coli </it>BL21 (DE3), ArcA and IclR are poorly expressed, explaining the similar fluxes observed compared to the Δ<it>arcA</it>Δ<it>iclR </it>strain.</p> |
| first_indexed | 2024-04-13T01:52:38Z |
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| institution | Directory Open Access Journal |
| issn | 1471-2180 |
| language | English |
| last_indexed | 2024-04-13T01:52:38Z |
| publishDate | 2011-04-01 |
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| series | BMC Microbiology |
| spelling | doaj.art-336cc4d704e845439c831242ac91e3562022-12-22T03:07:51ZengBMCBMC Microbiology1471-21802011-04-011117010.1186/1471-2180-11-70Effect of <it>iclR </it>and <it>arcA </it>knockouts on biomass formation and metabolic fluxes in <it>Escherichia </it><it>coli </it>K12 and its implications on understanding the metabolism of <it>Escherichia coli </it>BL21 (DE3)Charlier DanielHeijnen Joseph JFoulquié-Moreno Maria RDe Mey MarjanMaertens JoMoens HelenaBeauprez JoeriWaegeman HendrikSoetaert Wim<p>Abstract</p> <p>Background</p> <p>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 <it>aceBAK </it>operon. This study investigates the physiological and metabolic consequences of <it>arcA </it>and <it>iclR </it>deletions on <it>E. coli </it>K12 MG1655 under glucose abundant and limiting conditions and compares the results with the metabolic characteristics of <it>E. coli </it>BL21 (DE3).</p> <p>Results</p> <p>The deletion of <it>arcA </it>and <it>iclR </it>results in an increase in the biomass yield both under glucose abundant and limiting 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 CO<sub>2 </sub>producing pathways in the <it>E. coli </it>K12 Δ<it>arcA</it>Δ<it>iclR </it>double knockout strain. Due to <it>iclR </it>gene deletion, the glyoxylate pathway is activated resulting in a redirection of 30% of the isocitrate molecules directly to succinate and malate without CO<sub>2 </sub>production. Furthermore, a higher flux at the entrance of the TCA was noticed due to <it>arcA </it>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 Δ<it>iclR </it>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 and under both growth conditions, illustrating the transcriptional control of this pathway. Finally, similar central metabolic fluxes were observed in <it>E. coli </it>K12 Δ<it>arcA </it>Δ<it>iclR </it>compared to the industrially relevant <it>E. coli </it>BL21 (DE3), 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 <it>iclR </it>and rare codons are present in <it>arcA </it>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 <it>E. coli </it>K12 Δ<it>arcA</it>Δ<it>iclR</it>.</p> <p>Conclusions</p> <p>The deletion of <it>arcA </it>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 <it>E. coli </it>BL21 (DE3), ArcA and IclR are poorly expressed, explaining the similar fluxes observed compared to the Δ<it>arcA</it>Δ<it>iclR </it>strain.</p>http://www.biomedcentral.com/1471-2180/11/70 |
| spellingShingle | Charlier Daniel Heijnen Joseph J Foulquié-Moreno Maria R De Mey Marjan Maertens Jo Moens Helena Beauprez Joeri Waegeman Hendrik Soetaert Wim Effect of <it>iclR </it>and <it>arcA </it>knockouts on biomass formation and metabolic fluxes in <it>Escherichia </it><it>coli </it>K12 and its implications on understanding the metabolism of <it>Escherichia coli </it>BL21 (DE3) BMC Microbiology |
| title | Effect of <it>iclR </it>and <it>arcA </it>knockouts on biomass formation and metabolic fluxes in <it>Escherichia </it><it>coli </it>K12 and its implications on understanding the metabolism of <it>Escherichia coli </it>BL21 (DE3) |
| title_full | Effect of <it>iclR </it>and <it>arcA </it>knockouts on biomass formation and metabolic fluxes in <it>Escherichia </it><it>coli </it>K12 and its implications on understanding the metabolism of <it>Escherichia coli </it>BL21 (DE3) |
| title_fullStr | Effect of <it>iclR </it>and <it>arcA </it>knockouts on biomass formation and metabolic fluxes in <it>Escherichia </it><it>coli </it>K12 and its implications on understanding the metabolism of <it>Escherichia coli </it>BL21 (DE3) |
| title_full_unstemmed | Effect of <it>iclR </it>and <it>arcA </it>knockouts on biomass formation and metabolic fluxes in <it>Escherichia </it><it>coli </it>K12 and its implications on understanding the metabolism of <it>Escherichia coli </it>BL21 (DE3) |
| title_short | Effect of <it>iclR </it>and <it>arcA </it>knockouts on biomass formation and metabolic fluxes in <it>Escherichia </it><it>coli </it>K12 and its implications on understanding the metabolism of <it>Escherichia coli </it>BL21 (DE3) |
| title_sort | effect of it iclr it and it arca it knockouts on biomass formation and metabolic fluxes in it escherichia it it coli it k12 and its implications on understanding the metabolism of it escherichia coli it bl21 de3 |
| url | http://www.biomedcentral.com/1471-2180/11/70 |
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