Engineering Saccharomyces cerevisiae for isoprenol production

Isoprenol (3-methyl-3-butene-1-ol) is a valuable drop-in biofuel and an important precursor of several commodity chemicals. Synthetic microbial systems using the heterologous mevalonate pathway have recently been developed for the production of isoprenol in Escherichia coli, and a significant yield and titer improvement has been achieved through a decade of research. Saccharomyces cerevisiae has been widely used in the biotechnology industry for isoprenoid production, but there has been no good example of isoprenol production reported in this host. In this study, we engineered the budding yeast S. cerevisiae for improved biosynthesis of isoprenol. The strain engineered with the mevalonate pathway achieved isoprenol production at the titer of 36.02 ± 0.92 mg/L in the flask. The IPP (isopentenyl diphosphate)-bypass pathway, which has shown more efficient isoprenol production by avoiding the accumulation of the toxic intermediate in E. coli, was also constructed in S. cerevisiae and improved the isoprenol titer by 2-fold. We further engineered the strains by deleting a promiscuous endogenous kinase that could divert the pathway flux away from the isoprenol production and improved the titer to 130.52 ± 8.01 mg/L. Finally, we identified a pathway bottleneck using metabolomics analysis and overexpressed a promiscuous alkaline phosphatase to relieve this bottleneck. The combined efforts resulted in the titer improvement to 383.1 ± 31.62 mg/L in the flask. This is the highest isoprenol titer up to date in S. cerevisiae and this work provides the key strategies to engineer yeast as an industrial platform for isoprenol production.