Increase of organic solvent tolerance of Escherichia coli by the deletion of two regulator genes, fadR and marR.
The improvement of bacterial tolerance to organic solvents is a main prerequisite for the microbial production of biofuels which are toxic to cells. For targeted genetic engineering of Escherichia coli to increase organic solvent tolerances (OSTs), we selected and investigated a total of 12 genes that participate in relevant mechanisms to tolerance. In a spot assay of 12 knockout mutants with n-hexane and cyclohexane, the genes fadR and marR were finally selected as the two key genes for engineering. Fatty acid degradation regulon (FadR) regulates the biosynthesis and degradation of fatty acids coordinately, and the multiple antibiotic resistance repressor (MarR) is the repressor of the global regulator MarA for multidrug resistance. In the competitive growth assay, the ΔmarR mutant became dominant when the pooled culture of 11 knockout mutants was cultivated successively in the presence of organic solvent. The increased OSTs in the ΔmarR and ΔfadR mutants were confirmed by a growth experiment and a viability test. The even more highly enhanced OSTs in the ΔfadR ΔmarR double mutant were shown compared with the two single mutants. Cellular fatty acid analysis showed that the high ratio of saturated fatty acids to unsaturated fatty acids plays a crucial role in OSTs. Furthermore, the intracellular accumulation of OST strains was significantly decreased compared with the wild-type strain.
Department of Life Science, Ewha Womans University, 120-750, Seoul, South Korea.
Hye Yun
HY
Oh
Jae Ok
JO
Lee
Ok Bin
OB
Kim
D001426:Bacterial Proteins / Q000235:genetics* / Q000378:metabolism; D024881:Drug Resistance, Bacterial; D004926:Escherichia coli / Q000187:drug effects* / Q000235:genetics* / Q000378:metabolism; D029968:Escherichia coli Proteins / Q000235:genetics* / Q000378:metabolism; D017353:Gene Deletion; D015964:Gene Expression Regulation, Bacterial / Q000187:drug effects; D005809:Genes, Regulator; D009930:Organic Chemicals / Q000494:pharmacology*; D012097:Repressor Proteins / Q000235:genetics* / Q000378:metabolism; D015854:Up-Regulation
D001426:Bacterial Proteins; D029968:Escherichia coli Proteins; C057119:FadR protein, Bacteria; C080004:MarR protein, E coli; D009930:Organic Chemicals; D012097:Repressor Proteins