Collaborative Research: Mechanisms for Cell Membrane Damage during Production of Biorenewable Fuels

通过将生物质衍生的糖发酵成酒精将植物生物质转化为生物燃料是可持续生产可再生燃料的途径。然而，在发酵过程中，产生的生物燃料分子往往在高浓度下对发酵生物产生毒性，这降低了生物燃料的整体生产能力。解决这个问题的一种方法是重新设计发酵细胞周围的膜，这样生物体就可以更耐受高浓度的生物燃料溶解在它周围的液态水中。这一合作项目将对乙醇等生物燃料分子如何与酵母(一种发酵微生物)的细胞膜相互作用产生基本了解。关键的创新是使用复杂的分子动力学模拟工具在计算机上对这些相互作用进行建模。这些研究将提出针对酵母细胞的基因工程的策略，以表达细胞膜，从而提高细胞对高浓度生物燃料的整体耐受性，从而提高生物燃料的产量。与该项目相关的教育活动包括一个初高中推广项目，旨在通过爱荷华州立大学和马里兰大学的项目协调，强调实验和理论如何合作解决重要的科学问题。这项合作研究的总体目标是从根本上了解微生物膜与已知的影响膜破坏的模型生物燃料分子的细胞和生物分子相互作用。这项研究将提出如何设计细胞膜，使细胞对这些生物燃料分子更具耐受性。该研究计划将重点放在酿酒酵母膜如何与模型生物燃料分子或中间体相互作用，包括乙醇、辛酸和正丁醇。这些努力将为设计细胞膜的基因工程方法提供信息，以提高酵母细胞对这些生物燃料分子的耐受性。膜的关键指标包括孔隙率、流动性、疏水性和刚性。该研究计划有三个目标。第一个目标是建立理论和实验模型膜系统，使用乙醇作为表征良好的模型缓蚀剂。理论方法将使用分子动力学模拟来揭示膜的分子特征和脂质组成如何影响乙醇介导的膜破坏。第二个目标是探索辛酸和正丁醇与第一个目标下开发的模型膜系统的相互作用。将考虑更复杂的膜系统，以确定麦角固醇、链不饱和和脂头基团对膜干扰物毒性的影响。将体外组装的膜囊泡与完整的全细胞囊泡进行比较，建立脂质混合物与生物系统的模型膜。第三个目标是使用分子动力学模拟来预测改变脂头基团浓度、链饱和度、链支化和麦角甾醇浓度来改善膜的耐受性。耐受性提高的膜将在体外进行测试，然后在酿酒酵母工程细胞中表达和测试。

The conversion of plant biomass to biofuels by fermentation of biomass-derived sugars to alcohols is a sustainable route for renewable fuels production. However, during the fermentation process, the biofuel molecules produced are often toxic to the fermenting organism at high concentrations, which lowers the overall biofuel production capacity. One way to address this problem is redesign the membrane surrounding the fermenting cell, so that the organism can be more tolerant to high concentrations of biofuel dissolved in the liquid water surrounding it. This collaborative project will develop a fundamental understanding of how biofuel molecules such as ethanol interact with the cell membrane of yeast, a fermenting microorganism. The key innovation is use of sophisticated molecular dynamic simulation tools to model these interactions on computer. These studies will suggest strategies to target genetic engineering of the yeast cell to express cell membranes that improve the overall tolerance of the cell to high concentrations of biofuel, so that biofuel production is improved. The educational activities associated with this project include a middle- and high school outreach program designed to highlight how experiments and theory work together to solve important scientific problems, coordinated through programs at Iowa State University and the University of Maryland.The overall goal of this collaborative research is to gain a fundamental understanding of the cellular and biomolecular interactions of the microbial membranes with model biofuel molecules known to influence membrane disruption. The research will suggest how cell membranes can be engineered so that the cell is more tolerant to these biofuel molecules. The research plan will focus on how Saccharomyces cerevisiae membranes interact with model biofuel molecules or intermediates, including ethanol, octanoic acid, and n-butanol. These efforts will inform genetic engineering approaches to design cell membranes that improve the tolerance of the yeast cell to these biofuel molecules. Key membrane metrics include porosity, fluidity, hydrophobicity and rigidity. The research plan has three objectives. The first objective is to establish theoretical and experimental model membrane systems, using ethanol as a well-characterized model inhibitor. The theoretical approach will use molecular dynamics simulations to reveal how the molecular character and lipid composition of the membrane influence ethanol-mediated membrane disruption. The second objective is to probe the interaction of octanoic acid and n-butanol with model membrane systems developed under the first objective. More complex membrane systems will be considered to determine the effect of ergosterol, chain unsaturation, and lipid head groups on membrane disrupter toxicity. Membrane vesicles assembled in vitro will be compared vesicles made from whole cells as well as intact whole cells to establish model membrane of lipid mixtures to living systems. The third objective is to use molecular dynamics simulations to predict the improved tolerance of membranes with altered lipid head group concentrations, chain saturation, chain branching, and ergosterol concentration. Membranes with improved tolerance will be tested in vitro and then expressed and tested in engineered S. cerevisiae cells.