S6: Engineering microbial biofuel tolerance and export using efflux pumps

A major challenge when using microorganisms to produce bulk chemicals like biofuels is that the production targets are often toxic to cells. Biofuel-like compounds are known to reduce cell viability through damage to the cell membrane and interference with essential physiological processes. Thus, cells must trade off biofuel production and survival, reducing potential yields. The majority of microbial biofuel research to-date has focused on engineering metabolic pathways for biofuel production. It is essential that we also engineer strains for biofuel tolerance. Here, we focus on engineering tolerance to advanced biofuels, taking a targeted approach toward improving biofuel tolerance and yield. In particular, we use efflux pumps, which are membrane transporters, as a direct mechanism for reducing biofuel toxicity. For several advanced biofuels, we identified pumps that restored growth in the presence of biofuel. We then tested a beneficial pump directly in a production strain and demonstrated that it improved biofuel yields. Our findings introduce new tools for engineering production strains.