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，特别是在极高重力（VHG）条件下，需要保持健康的酵母菌群。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，并与反馈控制算法的连续培养，以开发以目的为导向的发酵过程，以研究细胞代谢和调节受到外来干扰。在此，我建议调查的影响发酵氧化还原电位的代谢通量的分布，还原当量，ATP在酿酒酵母在极高重力（VHG）乙醇生产，以及潜在的酵母生理。本论文的近期目标是：1.建立一个氧化还原电位控制的VHG乙醇发酵过程; 2.研究不同氧化还原电位下VHG乙醇发酵过程中基因表达谱; 3.构建一个完整的氧化还原电位影响下的乙醇发酵基因组和代谢网络; 4.构建氧化还原电位影响下的VHG乙醇发酵特异性蛋白质-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能力之间的瞬时平衡。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当量对和ATP，最终导致酵母细胞内代谢通量的重新分配。在VHG乙醇生产过程中保持发酵罐有氧是必要的。然而，太多的氧气将使代谢通量重新导向生物质形成，从而降低乙醇产率。 本研究拟开发一种氧化还原电位控制的微氧发酵工艺，以研究乙醇Redr发酵过程中代谢通量、还原当量和ATP的分布。酿酒酵母菌株该菌株最近已被测序，其基因组信息可用。这些信息将用于重建基因组规模的代谢模型（GSMM）。结合从上述发酵过程和GSMM收集的实验数据，可以鉴定在特定条件下被调节的基因，并且可以阐明它们在VHG条件下与乙醇生产相关的代谢功能。因此，这将导致深入了解酵母细胞的基因表达，然后可以用来设计一个有效的发酵策略，以提高乙醇生产率。在乙醇生产过程中，发酵方法可以用于生产酿酒酵母，但如此开发的发酵方法也可以用于生产重要的中间体单体，如琥珀酸或1，3-丙二醇，以及其他生物燃料，如生物丁醇。该研究计划将有助于高素质人才的发展和生物燃料和化学工业的经济增长。