Carboxylic acids are an attractive biorenewable chemical. However, like many other fermentatively produced compounds, they are inhibitory to the biocatalyst. An understanding of the mechanism of toxicity can aid in mitigating this problem. Here, we show that hexanoic and octanoic acids are completely inhibitory to Escherichia coli MG1655 in minimal medium at a concentration of 40 mM, while decanoic acid was inhibitory at 20 mM. This growth inhibition is pH-dependent and is accompanied by a significant change in the fluorescence polarization (fluidity) and integrity. This inhibition and sensitivity to membrane fluidization, but not to damage of membrane integrity, can be at least partially mitigated during short-term adaptation to octanoic acid. This short-term adaptation was accompanied by a change in membrane lipid composition and a decrease in cell surface hydrophobicity. Specifically, the saturated/unsaturated lipid ratio decreased and the average lipid length increased. A fatty acid-producing strain exhibited an increase in membrane leakage as the product titer increased, but no change in membrane fluidity. These results highlight the importance of the cell membrane as a target for future metabolic engineering efforts for enabling resistance and tolerance of desirable biorenewable compounds, such as carboxylic acids. Knowledge of these effects can help in the engineering of robust biocatalysts for biorenewable chemicals production.

Carboxylic acids are an attractive biorenewable chemical in terms of their flexibility and usage as precursors for a variety of industrial chemicals. It has been demonstrated that such carboxylic acids can be fermentatively produced using engineered microbes, such as Escherichia coli and Saccharomyces cerevisiae. However, like many other attractive biorenewable fuels and chemicals, carboxylic acids become inhibitory to these microbes at concentrations below the desired yield and titer. In fact, their potency as microbial inhibitors is highlighted by the fact that many of these carboxylic acids are routinely used as food preservatives. This review highlights the current knowledge regarding the impact that saturated, straight-chain carboxylic acids, such as hexanoic, octanoic, decanoic and lauric acids can have on E. coli and S. cerevisiae, with the goal of identifying metabolic engineering strategies to increase robustness. Key effects of these carboxylic acids include damage to the cell membrane and a decrease of the microbial internal pH. Certain changes in cell membrane properties, such as composition, fluidity, integrity and hydrophobicity, and intracellular pH are often associated with increased tolerance. The availability of appropriate exporters, such as Pdr12, can also increase tolerance. The effect on metabolic processes, such as maintaining appropriate respiratory function, regulation of Lrp activity and inhibition of production of key metabolites such as methionine, are also considered. Understanding the mechanisms of biocatalyst inhibition by these desirable products can aid in the engineering of robust strains with improved industrial performance.