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Functional Genomic Study of Exogenous n-Butanol Stress in Escherichia coli

ABSTRACT: n-Butanol has been proposed as an alternative biofuel to ethanol, and both Escherichia coli and Saccharomyces cerevisiae have been engineered to produce it. Unfortunately, n-butanol is more toxic than ethanol to these organisms. To understand the basis for its toxicity, cell wide studies were conducted at the transcript, protein and metabolite levels to obtain a global view of the n-butanol stress response. Analysis of the data indicate that n-butanol stress has components common to other stress responses and includes perturbation in respiratory functions (nuo, cyo operons), oxidative stress (sodC, katG, yqhD), heat shock and cell envelope stress (rpoE, clpB, htpG, degP, cpxPR), metabolite transport (malE, opp operon) and biosynthesis. Inducible expression of the yqhD gene was found to improve the host’s tolerance to exogenous n-butanol and confirms the role of this gene in coping with butanol stress. To survey for other potential candidates that may serve to improve host tolerance, mutant strains in several candidates which show changes at the transcript and protein levels were examined for sensitivity during butanol exposure. Chassis engineering based on these cues may be required in a high production titer, butanol-producing host.

ORGANISM(S): Escherichia coli

PROVIDER: GSE16973 | GEO | 2009/12/31

SECONDARY ACCESSION(S): PRJNA117267

REPOSITORIES: GEO

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Project description:The presence of anti-microbial phenolic compounds, such as the model compound ferulic acid, in biomass hydrolysates poses significant challenges to the widespread use of biomass in conjunction with whole cell biocatalysis or fermentation. Biofuel toxicity must also be overcome to allow for efficient production of next generation biofuels such as butanol, isopropanol, and others for widespread usage. Currently, these inhibitory compounds must be removed through additional downstream processing or sufficiently diluted to create environments suitable for most industrially important microbial strains. This study explores the high ferulic acid and n-butanol tolerance in Lactobacillus brevis (L. brevis), a lactic acid bacteria often found in fermentation processes, by global transcriptional response analysis. The transcriptional profile of L. brevis under ferulic acid and butanol stress reveals that the presence of ferulic acid primarily triggers the expression of membrane proteins to counteract ferulic acid induced changes in membrane fluidity and ion leakage. In contrast to the ferulic acid stress response, butanol addition to growing cultures uniquely induced the entire fatty acid synthesis pathway in the midst of a generalized stress response. Overexpression of the rate-limiting acetyl-CoA carboxylase subunits (AccABCD) in E. coli to increase lipid synthesis had no effect on butanol tolerance, suggesting that additional engineering is necessary to produce sufficient levels of appropriate fatty acids to confer butanol tolerance. Several promising routes for understanding both phenolic acid and butanol tolerance have been identified based upon these findings. These insights may be used to guide further engineering of model industrial organisms to better tolerate both classes of inhibitors in processed biomass used for biofuel production. Cultures were grown to OD ~ 0.2 in MRS media (baffled flasks), T = 30 C, 100 rpm. Butanol was then added to the cultures. Samples were harvested 15, 75, and 135 min after butanol addition. Each time point has 3 biological replicates, and dye swaps were incorporated into the microarray experiments.

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Functional Genomic Study of Exogenous n-Butanol Stress in Escherichia coli

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