(280b) The Isoprenoid Alcohol (IPA) Pathway: A Synthetic Route for Isoprenoid Biosynthesis

Isoprenoids comprise more than 50,000 different structures found in all kingdoms of life, with numerous applications including high-value pharmaceuticals, commodity chemicals and fuels. Despite there being two distinct native pathways, the mevalonate (MVA) and 2-C-methyl-D-erythritol-4-phosphate (MEP) pathways, for production of prenylated pyrophosphate precursors leading to isoprenoid compounds, engineering isoprenoid production at high flux, titer and yield remains a challenge due to the inherent carbon- and energy-inefficiencies and the roles of these pathways in native metabolism. To overcome these limitations, we have developed an alternative, synthetic pathway, termed the Isoprenoid Alcohol (IPA) pathway, that centers around the synthesis and subsequent phosphorylation of isoprenoid alcohols. Opposed to working within the confines of native metabolic pathways, our approach focused on utilizing all available biochemical reactions to identify potentially more efficient route(s) to isoprenoid compounds compared to native isoprenoid biosynthesis pathways. The work presented here will highlight the development of the IPA pathway in which we first established an efficient lower IPA pathway that can operate at high flux for the conversion of isoprenoid alcohols to isoprenoid pyrophosphate intermediates, facilitating the synthesis of diverse products, including geraniol, limonene, farnesol, diaponeurosporene, and lycopene. We then designed upper IPA pathways for the generation of isoprenoid alcohols from central carbon metabolites, providing the IPA pathway with the potential to operate with a large range of feedstocks. Integration of the full IPA pathway consisting of these upper and lower branches enabled the synthesis of geraniol, nerol, and citronellol from glycerol as the sole carbon source, demonstrating the implementation of a 3^rd, non-natural route to isoprenoid biosynthesis. This first-of-its-kind synthetic isoprenoid pathway provides an alternative route to isoprenoids that is more energy efficient than native pathways and can serve as a platform for targeting a repertoire of isoprenoid compounds.