Distributions of metabolic flux and reducing equivalents/ATP in Saccharomyces cerevisiae during redox potential-controlled very-high-gravity ethanol fermentation

氧化还原电位控制超重力乙醇发酵过程中酿酒酵母代谢通量和还原当量/ATP的分布

• 批准号: RGPIN-2019-07035
• 负责人: Lin, YenHan
• 金额: $ 2.04万
• 依托单位: University of Saskatchewan
• 依托单位国家: 加拿大
• 项目类别: Discovery Grants Program - Individual
• 财政年份: 2022
• 资助国家: 加拿大
• 起止时间: 2022-01-01 至 2023-12-31
• 项目状态: 已结题
• 来源: https://www.nserc-crsng.gc.ca/ase-oro/Details-Detailles_eng.asp?id=746776
• 关键词: Distributions; metabolic; flux; reducing; equivalents

Maintaining a healthy yeast population is required during ethanol fermentation, particularly under very-high-gravity (VHG) conditions. My long-term objective is to integrate bioinformatics, genomics, proteomics, metabolic flux analysis, and continuous cultivation with feedback control algorithms to develop purpose-oriented fermentation processes to investigate cellular metabolism and regulation subject to extraneous perturbations. Herein, I propose to investigate the effect of fermentation redox potential on the distribution of metabolic flux, reducing equivalents, and ATP within Saccharomyces cerevisiae during very-high-gravity (VHG) ethanol production, as well as the underlying yeast physiology. My short-term objectives are to: 1.develop a continuous redox potential-controlled VHG ethanol fermentation process; 2.investigate gene expression profiles during VHG ethanol fermentation under various redox potential levels; 3.reconstruct an integrated genomic and metabolic network specific to ethanol fermentation under the influence of redox potential; and 4.construct a protein-protein interaction network specific to VHG ethanol fermentation under the influence of redox potential. Fermentation redox potential reflects the momentary balance between oxidizing and reducing powers in a fermenter. By manipulating the extracellular fermentation redox potential, the intracellular reducing equivalent pairs and ATP can be indirectly altered, ultimately resulting in the redistribution of metabolic flux inside the yeast cells. keeping fermenter aerobic during VHG ethanol production is essential. However, too much oxygen will re-route metabolic flux towards biomass formation, thereby lowering ethanol yield. We propose to develop a redox potential-controlled micro-aerobic fermentation process to study the distribution of metabolic flux, reducing equivalents, and ATP within Ethanol Redr, an industrial S. cerevisiae strain. This strain has recently been sequenced, and its genome information is available. This information will be used to reconstruct a genome-scale metabolic model (GSMM). Combining experimental data collected from the above-mentioned fermentation process and the GSMM, the genes being modulated under specific conditions can be identified and their metabolic functions relating to ethanol production under VHG conditions can be elucidated. This will consequently result in an in-depth understanding of gene expression of yeast cells, which can then be used to design an effective fermentation strategy to increase ethanol productivity.  The proposed research targets the use of S. cerevisiae during ethanol production, but the fermentation process so developed can also be adapted to produce important intermediate monomers, such as succinic acid or 1,3-propanediol, and other biofuels such as biobutanol. This research program will help in the development of highly qualified personnel and the economic growth of the biofuel and chemical industry.

在乙醇发酵过程中，需要维持健康的酵母菌种群，尤其是在非常高重力（VHG）条件下。我的长期目标是整合生物信息学，基因组学，蛋白质组学，代谢通量分析，并连续培养与反馈控制算法，以开发出目的导向的发酵过程，以研究细胞代谢和受到外部扰动的调节。在本文中，我建议研究发酵氧化还原电位对酿酒酵母在非常高的（VHG）乙醇产生期间酿酒酵母中的代谢通量，减少等效物的分布，以及基本酵母菌生理学的影响。我的短期目标是：1。开发连续的氧化还原电位控制的VHG乙醇发酵过程； 2.在各种氧化还原电位水平下VHG乙醇发酵过程中的基因表达谱； 3.在氧化还原电位的影响下，对乙醇发酵特有的集成基因组和代谢网络进行了认可； 4.在氧化还原电位的影响下构建特定于VHG乙醇发酵的蛋白质 - 蛋白质相互作用网络。发酵氧化还原电位反映了发酵罐中氧化和还原力之间的瞬时平衡。通过操纵细胞外发酵氧化还原电位，可以间接改变细胞内还原等效对和ATP，最终导致酵母细胞内代谢通量的重新分布。在VHG乙醇生产过程中保持发酵罐有氧运动至关重要。然而，过多的氧气会重新释放代谢通量向生物量形成，从而降低乙醇的产量。我们建议开发一种氧化还原电势控制的微氧化盐发酵过程，以研究代谢通量，减少等效物的分布，以及在乙醇REDR中的ATP，这是一种工业S. cerevisiae菌株。最近被测序，其基因组信息可获得。该信息将用于重建基因组规模的代谢模型（GSMM）。可以鉴定从上述发酵过程和GSMM中收集的实验数据，可以鉴定在特定条件下调节的基因，并且可以阐明其与乙醇产生有关的代谢功能。因此，这将导致对酵母细胞基因表达的深入了解，然后可以使用该基因来设计有效的发酵策略来增加乙醇的产生。拟议的研究针对乙醇生产过程中酿酒酵母的使用，但是如此开发的发酵过程也可以适应生产重要的中间单体，例如琥珀酸或1,3-丙二醇，以及其他生物织物，例如生物丁醇。该研究计划将有助于发展高素质的人员以及生物燃料和化学工业的经济增长。