Monday, August 2, 2010
Pacific Concourse (Hyatt Regency San Francisco)
(E,E)-α-farnesene (3,7,11-trimethyldodeca-1,3E,6E,10-tetraene, abbreviated to EAF) is an acyclic hydrocarbon, which is primly discovered in apple peel and plays a role in plant defense. Recently, farnesene has been developed to a fuel precursor that can be hydrogenated to farnesane. The FPP precursor is synthesized from the universal C5 units, isopentenyl pyrophosphate (IPP) and its allylic isomer dimethylallyl pyrophosphate (DMAPP) by FPP synthase. Two distinct pathways, mevalonate (MVA) pathway dominantly in eukaryotes and some bacteria and 2-methyl-erythrotol-4-phosphate (MEP) pathway in bacteria and plastids, generate these two primary C5 building blocks. It has been reported MVA pathway is a superior route to MEP pathway for biosynthesis of IPP and DMAPP. High EAF production requires the sufficient FPP precursor supply and metabolic flux shift to MVA pathway. Poor expression of plant originated terpene synthase in E. coli has been considered as a limiting factor of low production. Synthesized α-farnesene synthase (FS) gene with codon optimization for expression in E. coli can minimize influence of poor protein expression to EAF production. Augmentation of FPP pool and FS overexpression elevated EAF production. In this study, we proved the feasibility of EAF production in metabolically engineered E. coli. EAF of 21.6 mg/L was produced by introduction of heterologous MVA pathway. Additional overexpression of ispA gave rise to 108.3mg/L of EAF production. This work was supported by the 21C Frontier Microbial Genomics and Applications Center Program, EB-NCRC (Grant No. R15-2003-012-02001-0), and BK21 program of Korea.