Abstract
Monoterpenes have an established use in the food and cosmetic industries and have recently also found application as advanced biofuels. Although metabolic engineering efforts have so far achieved significant yields of larger terpenes, monoterpene productivity is lagging behind. Here, we set out to establish a monoterpene-specific production platform in Saccharomyces cerevisiae and identified the sequential reaction mechanism of the yeast farnesyl diphosphate synthase Erg20p to be an important factor limiting monoterpene yield. To overcome this hurdle, we engineered Erg20p into a geranyl diphosphate synthase and achieved a significant increase in monoterpene titers. To further improve production, we converted the engineered geranyl diphosphate synthase into a dominant negative form, so as to decrease the ability of the endogenous Erg20p to function as a farnesyl diphosphate synthase, without entirely abolishing sterol biosynthesis. Fusion of the synthetic dominant negative Erg20p variant with the terpene synthase, combined with yeast strain engineering, further improved monoterpene yields and achieved an overall 340-fold increase in sabinene yield over the starting strain. The design described here can be readily incorporated to any dedicated yeast strain, while the developed plasmid vectors and heterozygous ERG20 deletion yeast strain can also be used as a plug-and-play system for enzyme characterization and monoterpene pathway elucidation.
| Originalsprog | Engelsk |
|---|---|
| Tidsskrift | A C S Synthetic Biology |
| Vol/bind | 3 |
| Udgave nummer | 5 |
| Sider (fra-til) | 298-306 |
| Antal sider | 9 |
| ISSN | 2161-5063 |
| DOI | |
| Status | Udgivet - 16 maj 2014 |
| Udgivet eksternt | Ja |