Reconstituting Non-Mevalonate Pathway in Yeast for Overproduction of Isoprenoids

Isoprenoids are a large group of natural products, which include numerous molecules that are valuable to human beings, such as pharmaceuticals (e.g. paclitaxel and artemisinin), next-generation biofuels (e.g. isopentenol and farnesene), and household chemicals (e.g. nootkatone and squalene). All isoprenoids are derived from two building blocks, isoprenyl pyrophosphate (IPP) and dimethylallyl diphosphate, which can be synthesized via mevalonate (MVA) pathway or methylerythritol phosphate (MEP) pathway in nature. Theoretically, the MEP pathway is more efficient in terms of pathway yield, largely due to the fact that using this pathway loses less carbon as CO₂ as compared to the MVA pathway (MEP pathway: 1 CO₂/IPP; MVA pathway: 4 CO₂/IPP; sugar as carbon source). Therefore, it would be highly attractive to reconstitute the MEP pathway in Saccharomyces cerevisiae, one of the most widely used industrial microbes that, however, naturally possesses the MVA pathway. To date, only part of the MEP pathway has been successfully reconstructed in the yeast; this is because the last two enzymes of the pathway (ISPG and ISPH) are both metalloproteins which require reduced iron-sulfur clusters to function. Here, we report new strategies for the functional expression of ISPG/ISPH in yeast and demonstrate new approaches for expressing iron-sulfur cluster containing enzymes in S. cerevisiae, thus laying the foundation for reconstituting the MEP pathway in the yeast, which would undoubtedly advance current microbial isoprenoid production processes.