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Abstract
The fermentation inhibitors from the pretreatment of lignocellulosic materials, e.g., acetic acid and furfural, are notorious due to their negative effects on the cell growth and chemical production. However, the metabolic reprogramming of the cells under these stress conditions, especially metabolic response for resistance to mixed inhibitors, has not been systematically investigated and remains mysterious. Therefore, in this study, 13C metabolic flux analysis (13C-MFA), a powerful tool to elucidate the intracellular carbon flux distributions, has been applied to two Saccharomyces cerevisiae strains with different tolerances to the inhibitors under acetic acid, furfural, and mixed (i.e., acetic acid and furfural) stress conditions to unravel the key metabolic responses. By analyzing the intracellular carbon fluxes as well as the energy and cofactor utilization under different conditions, we uncovered varied metabolic responses to different inhibitors. Under acetate stress, ATP and NADH production was slightly impaired, while NADPH tended towards overproduction. Under furfural stress, ATP and cofactors (including both NADH and NADPH) tended to be overproduced. However, under dual-stress condition, production of ATP and cofactors was severely impaired due to synergistic stress caused by the simultaneous addition of two fermentation inhibitors. Such phenomenon indicated the pivotal role of the energy and cofactor utilization in resisting the mixed inhibitors of acetic acid and furfural. Based on the discoveries, valuable insights are provided to improve the tolerance of S. cerevisiae strain and further enhance lignocellulosic fermentation.
Highlights
The production of biofuels and other bio-based chemicals from renewable resources has been widely applied to overcome the limitation of non-renewable fossil fuel energy and the challenge of global warming
When cultivating the parent S. cerevisiae strain, S-C1, under the acetic acid stress condition, the growth rate and glucose uptake rate were repressed by 80% and 60%, respectively (Table 2)
The carbon fluxes in the tricarboxylic acid (TCA) cycle dramatically decreased (>85%) under the acetic acid stress condition, which is consistent with previous studies [13, 15]
Summary
The production of biofuels and other bio-based chemicals from renewable resources has been widely applied to overcome the limitation of non-renewable fossil fuel energy and the challenge of global warming. Due to the fast growth and high biofuel productivity, Saccharomyces. Metabolic Response to Mixed Inhibitors Using 13C-MFA cerevisiae is one of the most important workhorses in producing biofuels from various carbohydrates of biomass with high abundance and low cost. Several fermentation inhibitors generated from the pretreatment of lignocellulosic materials, such as acetic acid [1], furfural [2], and furan [3], seriously impair the biofuel production by repressing or even stopping the cell growth of S. cerevisiae [4,5,6,7]. It is important to elucidate the general metabolic responses of S. cerevisiae to different inhibitors, especially to mixed inhibitors, to further improve stress resistance and reduce stress impairment. By identifying a common target, it is more likely to find a way to solve the problem in practice
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