IMPROVING AND EXPANDING ISOPENTENOL-UTILIZATION PATHWAY FOR PRODUCING ISOPRENOIDS

Abstract

Isoprenoids are a large family of natural products with diverse structures. They have important commercial uses as pharmaceuticals, flavouring agents and fragrances. Recently, metabolic engineering has been intensively investigated for the production of isoprenoids through microbial fermentation. The Isopentenol Utilization pathway (IUP) was recently reported to efficiently provide the building blocks of isoprenoid biosynthesis from affordable C5 substrates. In this thesis, IUP was engineered in Escherichia coli to convert isopentenols into geranate, a valuable isoprenoid compound. The best strain produced 764 mg/L geranate in 24 h from 2 g/L isopentenols, among the highest geranate titers reported in the literature. A key limitation of IUP is that the supply of isopentenols depends on cracking of non-renewable petroleum. Thus, E. coli native methylerythritol 4-phosphate (MEP) pathway was then engineered to produce geranate. The geranate titer reached to 48.6 mg/L in 72 h by knocking out the formation of E. coli fermentation metabolites. However, the titer achieved was not comparable to IUP. Thus, a novel pathway was proposed to produce 4-hydroxy-prenol (4-HP) from renewable compounds and 4-HP was subsequently utilized as the substrate for IUP. This novel pathway would allow production of isoprenoids from simple and renewable resources.