Chemical genomic guided engineering of gamma-valerolactone tolerant yeast

Microbial Cell Factories

卷:17

Alex Hebert¹  Joshua J. Coon¹  Chad L. Myers²  Scott Bottoms³  Li Hinchman³  Robert Landick³  Mick McGee³  Jose Serate³  Dan Xie³  Jeff S. Piotrowski³  Quinn Dickinson³  Yaoping Zhang³  Alan Higbee⁴ 

[1] Department of Biomolecular Chemistry, University of Wisconsin–Madison;

[2] Department of Computer Science and Engineering, University of Minnesota-Twin Cities;

[3] Great Lakes Bioenergy Research Center, University of Wisconsin-Madison;

[4] University of Wisconsin Biotechnology Center;

关键词: Chemical genomics;    Gamma-valerolactone;    Lignocellulosic;    Biofuel;    Biocatalysts;    Saccharomyces cerevisiae;   

DOI  :  10.1186/s12934-017-0848-9

来源: DOAJ

【 摘 要 】

Abstract Background Gamma valerolactone (GVL) treatment of lignocellulosic bomass is a promising technology for degradation of biomass for biofuel production; however, GVL is toxic to fermentative microbes. Using a combination of chemical genomics with the yeast (Saccharomyces cerevisiae) deletion collection to identify sensitive and resistant mutants, and chemical proteomics to monitor protein abundance in the presence of GVL, we sought to understand the mechanism toxicity and resistance to GVL with the goal of engineering a GVL-tolerant, xylose-fermenting yeast. Results Chemical genomic profiling of GVL predicted that this chemical affects membranes and membrane-bound processes. We show that GVL causes rapid, dose-dependent cell permeability, and is synergistic with ethanol. Chemical genomic profiling of GVL revealed that deletion of the functionally related enzymes Pad1p and Fdc1p, which act together to decarboxylate cinnamic acid and its derivatives to vinyl forms, increases yeast tolerance to GVL. Further, overexpression of Pad1p sensitizes cells to GVL toxicity. To improve GVL tolerance, we deleted PAD1 and FDC1 in a xylose-fermenting yeast strain. The modified strain exhibited increased anaerobic growth, sugar utilization, and ethanol production in synthetic hydrolysate with 1.5% GVL, and under other conditions. Chemical proteomic profiling of the engineered strain revealed that enzymes involved in ergosterol biosynthesis were more abundant in the presence of GVL compared to the background strain. The engineered GVL strain contained greater amounts of ergosterol than the background strain. Conclusions We found that GVL exerts toxicity to yeast by compromising cellular membranes, and that this toxicity is synergistic with ethanol. Deletion of PAD1 and FDC1 conferred GVL resistance to a xylose-fermenting yeast strain by increasing ergosterol accumulation in aerobically grown cells. The GVL-tolerant strain fermented sugars in the presence of GVL levels that were inhibitory to the unmodified strain. This strain represents a xylose fermenting yeast specifically tailored to GVL produced hydrolysates.

【 授权许可】

Unknown