System Biology Approaches to Determine the Factors for Acetic Acid Resistance by Comparing S. cerevisiae 424A (LNH-ST) and 424A (LNH-ST) - AAR during Glucose / Xylsoe Co-Fermentation

Chia-Ling Wu, Purdue University

Abstract

Cellulosic ethanol provides a great alternative as a renewable and green low-carbon fuel. Inhibitors, especially acetic acid, found in the biomass hydrolyzate present a major challenge converting the feedstock to ethanol. Acetic acid inhibits microbial fermentation on both cell growth and ethanol production. Acetic acid is particularly inhibitory to the fermentation of xylose, which accounts for about 30% of the total sugars in the feedstock. Therefore, developing a microorganism with sufficient tolerance to acetic acid during fermentation is key to the success of the industry. The focus of this study was on the mechanisms of acetic acid resistance during xylose fermentation. Fermentation analysis and two systems biology approaches, metabolic and transcriptomic analysis, were used to compare two S. cerevisiae strains, 424A (LNH-ST) and 424A (LNH-ST) - AAR, during glucose/xylose co-fermentation in the presence of 10 g L-1 acetic acid, which represented the condition that could be expected in the industry. 424A (LNH-ST) - AAR was an acetic acid-resistant strain developed from the parent strain 424A (LNH-ST) through adaptation. Fermentation analysis showed that AAR strain could ferment twice as much xylose into ethanol with a specific xylose consumption rate that was 642% faster compared to the original strain. It also showed that AAR strain was capable of producing half of the amount of acetic acid per gram of sugar consumed and maintaining a higher medium pH during fermentation. Both metabolic and transcriptomic analysis indicated that maintaining and conserving cellular energy was paramount for xylose fermentation in the presence of acetic acid, which could be achieved from four aspects suggested by the expression profiles of the genes from transcriptomic analysis: (1) increasing the production of cellular energy by increasing the number of transporters with high affinity for xylose, (2) increasing the tolerance to acetic acid and ethanol simultaneously by changing the fluidity of the plasma membrane such as elevating the amount of ergosterol, (3) shutting down the expression of some less important energy-consuming proteins, and (4) utilizing the more energy-efficient vacuolar proton pumps instead of those in the plasma membrane to recover the internal pH that was perturbed upon acetic acid treatment. This dissertation presented the first study in the literature identifying the key factors for xylose fermentation by yeast in the presence of acetic acid. The findings from this study provide valuable information in order to better design microorganisms suitable for cellulosic ethanol production and can serve as a starting point for overcoming a major obstacle in the industry.

Degree

Ph.D.

Advisors

Sedlak, Purdue University.

Subject Area

Alternative Energy|Microbiology|Bioinformatics

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