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

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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.