Metabolic Engineering of Saccharomyces cerevisiae for Isoprenoid Production

Stefan Tippmann1, Sakda Khoomrung1, Verena Siewers1 and Jens Nielsen1

1Systems & Synthetic Biology, Department of Chemical and Biological Engineering, Chalmers University of Technology, Gothenburg/Sweden

The diminishing oil resources and an increasing environmental burden urge the need for alternative transportation fuels. Despite high quality demands, nature already provides compounds, which can be used directly as or be converted into fuels. Among them, isoprenoids were attributed a key function and have therefore received great attention within the last few years. This large group of secondary metabolites is ubiquitous in plant oils, resins and other volatile mixtures and has a broad spectrum of applications including the production of fragrances, flavors and pharmaceuticals. Since plant extraction as well as chemical synthesis suffer from several drawbacks and fail to meet industrial scale, there is a strong demand for an efficient and sustainable production process.
This project is dedicated to isoprenoids, which can be used as biofuel precursors and attempts to establish a yeast cell factory for their production. For this purpose, Saccharomyces cerevisiae was chosen as a host organism, whereas the main focus is set on sesquiterpenes such as farnesene, which can be used as diesel alternative in its hydrogenated form farnesane. In order to enable for efficient production of farnesene, two central aspects are being addressed, i.e. metabolic engineering for enhanced synthesis and analytical method development for accurate quantification of intra- and extracellular metabolites from two-liquid phase fermentations. In the first part, an existing platform optimized for sesquiterpene production was recently used for the integration of farnesene synthase genes from different plant sources to enable the one-step conversion from farnesyl pyrophosphate to farnesene. As a result, maximal titers of ~1 g/L were attained in a comparative evaluation in fed-batch cultivations with exponential feeding. Enhanced synthesis, however, will not only involve heterologous expression of these enzymes, but it will also include further engineering of the endogenous mevalonate pathway as well as the integration of different â??omicsâ?? analysis to support the cycle of metabolic engineering.

References:

Tippmann, S., Chen, Y., Siewers, V. and Nielsen, J. (2013). From flavors and pharmaceuticals to advanced biofuels: Production of isoprenoids in Saccharomyces cerevisiae. Biotechnology Journal, Vol. 8: 1435-1444

Scalcinati, G., Knuf, C., Partow, S., Nielsen, J., Siewers, V. et al. (2013). Dynamic control of gene expression in Saccharomyces cerevisiae engineered for the production of plant sesquiterpene α- santalene in fed-batch mode. Metabolic Engineering, Vol. 14: 91-103

Scalcinati, G., Partow, S., Siewers, S., Schalk, M., Daviet, L. and Nielsen, J., Combined metabolic engineering of precursor and co-factor supply to increase α-santalene production by Saccharomyces cerevisiae. Microbial Cell Factories, Vol. 11, 2012