Abstract
Higher chain alcohols have gained much attention as next generation transport fuels because of their higher energy density and low moisture absorption capacity compared to ethanol. In the present study, we attempted to engineer Saccharomyces cerevisiae for the synthesis of isoamyl alcohol via de novo leucine biosynthetic pathway coupled with Ehrlich degradation pathway. To achieve high-level production of isoamyl alcohol, two strategies are used in the current study: (1) reconstruction of a chromosome-based leucine biosynthetic pathway under the control of galactose-inducible promoters; (2) overexpression of the mitochondrial 2-isopropylmalate (α-IPM) transporter to boost the transportation of α-IPM from mitochondria to the cytosol. We found engineered yeast cells with a combinatorially assembled leucine biosynthetic pathway coupled with the Ehrlich degradation pathway resulted in high-level production of isoamyl alcohol; however, there was still a significant amount of isobutanol co-formed during the fermentation process. Further introducing an α-IPM transporter not only boosted the isoamyl alcohol biosynthetic pathway activity but also reduced isobutanol to a much lower level. Taken together, our work represents the first study to construct a chromosome-based leucine biosynthetic pathway for isoamyl alcohol production. Furthermore, the utilization of the mitochondrial compartment coupled with the transporter engineering serves as an effective approach to minimize the by-product formation and to improve the isoamyl alcohol production.
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Acknowledgements
This work was funded by the National University of Singapore (Start-up Grant: R279 000 364 133).
Authors’ contributions
JY conceived the study, designed the experiments, performed the experiments, analyzed the data, and drafted the manuscript. XC participated in the experiments, analyzed the data, and helped to revise the manuscript. PM participated in the experiments, analyzed the data, and helped to revise the manuscript. CBC supervised the project and revised the manuscript. All authors read and approved the final manuscript.
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Yuan, J., Chen, X., Mishra, P. et al. Metabolically engineered Saccharomyces cerevisiae for enhanced isoamyl alcohol production. Appl Microbiol Biotechnol 101, 465–474 (2017). https://doi.org/10.1007/s00253-016-7970-1
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DOI: https://doi.org/10.1007/s00253-016-7970-1