Abstract
BackgroundEconomically viable biobased production of bulk chemicals and biofuels typically requires high product titers. During microbial bioconversion this often leads to product toxicity, and tolerance is therefore a critical element in the engineering of production strains.ResultsHere, a systems biology approach was employed to understand the chemical stress response of Escherichia coli, including a genome-wide screen for mutants with increased fitness during chemical stress. Twelve chemicals with significant production potential were selected, consisting of organic solvent-like chemicals (butanol, hydroxy-γ-butyrolactone, 1,4-butanediol, furfural), organic acids (acetate, itaconic acid, levulinic acid, succinic acid), amino acids (serine, threonine) and membrane-intercalating chemicals (decanoic acid, geraniol). The transcriptional response towards these chemicals revealed large overlaps of transcription changes within and between chemical groups, with functions such as energy metabolism, stress response, membrane modification, transporters and iron metabolism being affected. Regulon enrichment analysis identified key regulators likely mediating the transcriptional response, including CRP, RpoS, OmpR, ArcA, Fur and GadX. These regulators, the genes within their regulons and the above mentioned cellular functions therefore constitute potential targets for increasing E. coli chemical tolerance. Fitness determination of genome-wide transposon mutants (Tn-seq) subjected to the same chemical stress identified 294 enriched and 336 depleted mutants and experimental validation revealed up to 60 % increase in mutant growth rates. Mutants enriched in several conditions contained, among others, insertions in genes of the Mar-Sox-Rob regulon as well as transcription and translation related gene functions.ConclusionsThe combination of the transcriptional response and mutant screening provides general targets that can increase tolerance towards not only single, but multiple chemicals.Electronic supplementary materialThe online version of this article (doi:10.1186/s12934-016-0577-5) contains supplementary material, which is available to authorized users.
Highlights
Viable biobased production of bulk chemicals and biofuels typically requires high product titers
Selection of chemicals and growth conditions The E. coli transcriptome has been investigated under chemical stress conditions with 12 different compounds (Fig. 1a) in order to understand the cellular response and potentially infer the cellular effects of the imposed stress
Five other chemicals were selected based on other criteria including acetate (Ace), a common inhibitor during fermentation; butanol (But), a high-potential biofuel; 1,4-butanediol (Diol), a polyester precursor; geraniol (Ger), a biofuel and fragrance ingredient and decanoic acid (Deca)
Summary
Viable biobased production of bulk chemicals and biofuels typically requires high product titers. During microbial bioconversion this often leads to product toxicity, and tolerance is a critical element in the engineering of production strains. Rational engineering attempts include improving tolerance towards butanol [18, 19], produced FFAs [20], and furfural [21, 22], while screening strategies have been employed towards butanol [23], acetate [24], and furfural stresses [25]. High extracellular titers are needed for an economically viable production
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