Microbial metabolism is highly dependent on the environmental conditions. Especially, the substrate concentration, as well as oxygen availability, determine the metabolic rates. In large-scale bioreactors, microorganisms encounter dynamic conditions in substrate and oxygen availability (mixing limitations), which influence their metabolism and subsequently their physiology. Earlier, single substrate pulse experiments were not able to explain the observed physiological changes generated under large-scale industrial fermentation conditions. In this study we applied a repetitive feast–famine regime in an aerobic Escherichia coli culture in a time-scale of seconds. The regime was applied for several generations, allowing cells to adapt to the (repetitive) dynamic environment. The observed response was highly reproducible over the cycles, indicating that cells were indeed fully adapted to the regime. We observed an increase of the specific substrate and oxygen consumption (average) rates during the feast–famine regime, compared to a steady-state (chemostat) reference environment. The increased rates at same (average) growth rate led to a reduced biomass yield (30% lower). Interestingly, this drop was not followed by increased by-product formation, pointing to the existence of energy-spilling reactions. During the feast–famine cycle, the cells rapidly increased their uptake rate. Within 10 s after the beginning of the feeding, the substrate uptake rate was higher (4.68 μmol/gCDW/s) than reported during batch growth (3.3 μmol/gCDW/s). The high uptake led to an accumulation of several intracellular metabolites, during the feast phase, accounting for up to 34% of the carbon supplied. Although the metabolite concentrations changed rapidly, the cellular energy charge remained unaffected, suggesting well-controlled balance between ATP producing and ATP consuming reactions. The adaptation of the physiology and metabolism of E. coli under substrate dynamics, representative for large-scale fermenters, revealed the existence of several cellular mechanisms coping with stress. Changes in the substrate uptake system, storage potential and energy-spilling processes resulted to be of great importance. These metabolic strategies consist a meaningful step to further tackle reduced microbial performance, observed under large-scale cultivations.

中文翻译：

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短期重复底物动力学下的大肠杆菌代谢：适应和权衡。

微生物的新陈代谢高度依赖于环境条件。特别地，底物浓度以及氧气的利用率决定了代谢速率。在大型生物反应器中，微生物会遇到底物和氧气供应（混合限制）的动态条件，这会影响其代谢，进而影响其生理。早期，单底物脉冲实验无法解释在大规模工业发酵条件下产生的观察到的生理变化。在这项研究中，我们在有氧大肠杆菌培养物中以秒为单位应用了重复性的饥荒制度。该方案已应用了几代，使细胞能够适应（重复的）动态环境。在整个周期中，观察到的响应具有高度可重复性，表明细胞确实完全适应了该方案。与稳态（化学稳定剂）参考环境相比，我们在餐饥荒期间观察到了特定底物和氧气消耗（平均）速率的增加。在相同（平均）增长率下增加的速率导致生物量产量降低（降低30％）。有趣的是，这种下降并未伴随着副产物形成的增加，这表明存在能量溢出反应。在大饥荒周期中，细胞会迅速增加其摄取率。在开始进料后的10 s内，底物吸收速率（4.68μmol/ gCDW / s）比分批生长期间的报告值（3.3μmol/ gCDW / s）高。在盛宴阶段，高摄入量导致几种细胞内代谢产物的积累，占所供应碳的34％。尽管代谢物的浓度变化很快，但细胞能量电荷仍然不受影响，表明ATP产生和ATP消耗反应之间的平衡得到了很好的控制。在底物动力学下对大肠杆菌的生理和代谢的适应，代表了大型发酵罐，揭示了应对压力的几种细胞机制的存在。底物吸收系统，存储潜力和能量收集过程的变化非常重要。这些代谢策略是进一步解决微生物繁殖能力下降的有意义的一步，这在大规模种植中观察到。在底物动力学下对大肠杆菌的生理和代谢的适应，代表了大型发酵罐，揭示了应对压力的几种细胞机制的存在。底物吸收系统，存储潜力和能量收集过程的变化非常重要。这些代谢策略是进一步解决微生物繁殖能力下降的有意义的一步，这在大规模种植中观察到。在底物动力学下对大肠杆菌的生理和代谢的适应，代表了大型发酵罐，揭示了应对压力的几种细胞机制的存在。底物吸收系统，存储潜力和能量收集过程的变化非常重要。这些代谢策略是进一步解决微生物繁殖能力下降的有意义的一步，这在大规模种植中观察到。