Evolutionary Methods for Improving the Production of Biorenewable Fuels and Chemicals

Evolutionary methods to improve strain performance are a powerful complement to rational, predictive strain engineering. These evolutionary methods can consist of serial transfers or engineered methods, such as global transcription machinery engineering and the SCALEs expression libraries. The type of improvements achieved in strain performance include increasing the tolerance to inhibitory products, such as alcohols and carboxylic acids, and increasing the tolerance and utilization of biomass-derived sugars, such as pentose sugars and biomass hydrolysate. In some cases, the selective pressure applied during the evolutionary process has enabled co-selection of growth and strain performance. This approach has proven successful for the production of ethanol, lactic acid, succinate and alanine. In other cases, strains are evolved for tolerance to exogenously supplied inhibitors, with the intention of increasing production by increasing tolerance. This has been successful in some cases, particularly in regards to ethanol production. Identification of the mutations that enable the evolved phenotype and understanding the molecular basis of their contribution to strain fitness and performance can reveal novel strategies for increasing tolerance. In this manner, our knowledge of possible design schemes and our understanding of our workhorse biocatalysts are increased. Reverse engineering has provided rational design strategies for increasing tolerance to isobutanol, furfural, and ethanol as well as increasing the production of succinate.