The sole introduction of two single-point mutations establishes glycerol utilization in Saccharomyces cerevisiae CEN.PK derivatives

Abstract

BackgroundGlycerol is an abundant by-product of biodiesel production and has several advantages as a substrate in biotechnological applications. Unfortunately, the popular production host Saccharomyces cerevisiae can barely metabolize glycerol by nature.ResultsIn this study, two evolved derivatives of the strain CEN.PK113-1A were created that were able to grow in synthetic glycerol medium (strains PW-1 and PW-2). Their growth performances on glycerol were compared with that of the previously published evolved CEN.PK113-7D derivative JL1. As JL1 showed a higher maximum specific growth rate on glycerol (0.164 h−1 compared to 0.119 h−1 for PW-1 and 0.127 h−1 for PW-2), its genomic DNA was subjected to whole-genome resequencing. Two point mutations in the coding sequences of the genes UBR2 and GUT1 were identified to be crucial for growth in synthetic glycerol medium and subsequently verified by reverse engineering of the wild-type strain CEN.PK113-7D. The growth rate of the resulting reverse-engineered strain was 0.130 h−1. Sanger sequencing of the GUT1 and UBR2 alleles of the above-mentioned evolved strains PW-1 and PW-2 also revealed one single-point mutation in these two genes, and both mutations were demonstrated to be also crucial and sufficient for obtaining a maximum specific growth rate on glycerol of ~0.120 h−1.ConclusionsThe current work confirmed the importance of UBR2 and GUT1 as targets for establishing glycerol utilization in strains of the CEN.PK family. In addition, it shows that a growth rate on glycerol of 0.130 h−1 can be established in reverse-engineered CEN.PK strains by solely replacing a single amino acid in the coding sequences of both Ubr2 and Gut1.Electronic supplementary materialThe online version of this article (doi:10.1186/s13068-016-0696-6) contains supplementary material, which is available to authorized users.

Metabolic Engineering of Saccharomyces cerevisiae for Enhanced Dihydroartemisinic Acid Production

“…During the fermentation process, a significant accumulation of glycerol was also found at about 5 g/L in glucose consumption stage. Ho and colleagues reported both UBR2 and GUT1 with single point mutation could be regarded as targets for establishing glycerol utilization in strains of the CEN.PK family (Ho et al, 2017). It was also found that regulatory and metabolic trade-offs of glycerol utilization in S. cerevisiae were revealed by laboratory evolution, which could guide us to enhance glycerol consumption rationally in our study (Strucko et al, 2018).…”

Engineering Saccharomyces cerevisiae for co-utilization of d-galacturonic acid and d-glucose from citrus peel waste

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Metabolic Remodeling during Long-Lasting Cultivation of the Endomyces magnusii Yeast on Oxidative and Fermentative Substrates

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