Ethanol production in syngas-fermenting Clostridium ljungdahlii is controlled by thermodynamics rather than by enzyme expression
Ethanol production in syngas-fermenting Clostridium ljungdahlii is controlled by thermodynamics rather than by enzyme expression
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DOI：
10.1039/c6ee01108j
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发表时间：
2016-01-01
影响因子：
32.5
通讯作者：
Angenent, L. T.
中科院分区：
文献类型：
--
作者：
Richter, H.;Molitor, B.;Angenent, L. T.
产乙酸细菌作为生物催化剂在生物技术工业中引起了人们的兴趣，因为它们能够发酵富含一氧化碳（CO）的气体。野生型菌株主要生产乙酸和乙醇，但基因改造已经扩大了产品组合。为了提高内在或异源生物化学品的生产，微生物生理学的知识是必要的。这种生理学包括两个不同的阶段：产酸（生长/乙酸盐生产）和溶剂生成（饥饿/乙醇生产）。我们操作两个连续的，连续的生物反应器与纯培养的杨氏梭菌达到稳态条件下，在乙酸和乙醇生产阶段，空间分离产酸和溶剂。在这里，近2000种蛋白质和它们的差异丰度产酸和solventogenesis被确定。此外，我们还测量了重要的代谢物。结果表明，营养有限的条件下引发的过渡到solventogenesis没有改变差异丰富的酶在中央能量代谢。我们的蛋白质组学结果表明，乙醇生产酶（AOR/ADH）是一贯丰富的，即使在产酸。在此基础上，我们开发了一个溢流模型，热力学，而不是遗传调控。该模型确定乙酸和还原辅因子作为饱和反应物。当胞内未解离的乙酸浓度达到一个热力学阈值时，C. ljungdahlii将能够立即将剩余的还原当量转向乙醇。这在生长迟缓期间是重要的，此时还原当量不再能够转向生物质生产，而富含CO的气体的供应仍然很高。可以操纵营养物质的可用性和pH值，以实现生物加工过程中所需的溶剂生成水平。
Acetogenic bacteria are attracting interest as biocatalysts in the biotechnology industry, since they are able to ferment carbon monoxide (CO)-rich gases. Wild-type strains produce mainly acetate and ethanol, but genetic modifications have already broadened the product portfolio. To enhance the production of intrinsic or heterologous biochemicals, knowledge of the microbial physiology is necessary. This physiology includes two different phases: acidogenesis (growth/acetate production) and solventogenesis (starvation/ethanol production). We operated two sequential, continuous bioreactors with a pure culture of Clostridium ljungdahlii to achieve steady-state conditions in an acetate-and an ethanol-producing stage to spatially separate acidogenesis and solventogenesis. Here, nearly 2000 proteins and their differential abundance between acidogenesis and solventogenesis were identified. In addition, we measured important metabolites. The results showed that nutrient-limited conditions triggered a transition to solventogenesis without altering the differential abundance of enzymes in the central energy metabolism. Our proteomics results revealed that the enzymes for ethanol production (AOR/ADH) were consistently abundant, even during acidogenesis. Based on this work, we developed an overflow model with thermodynamic rather than genetic regulation. The model identifies acetic acid and reduced cofactors as saturation reactants. When the intracellular concentration of undissociated acetic acid reaches a thermodynamic threshold, C. ljungdahlii will be able to shunt surplus reducing equivalents toward ethanol immediately. This is important during retarded growth, when reducing equivalents can no longer be shunted toward biomass production, while the supply of CO-rich gas is still high. Nutrient availability and pH can be manipulated to achieve the desirable level of solventogenesis during bioprocessing.