Isoprenoid Pathway Optimization for Taxol Precursor Overproduction in Escherichia coli

• Published in: Science (American Association for the Advancement of Science) Vol. 330; no. 6000; pp. 70 - 74
• Main Authors: Ajikumar, Parayil Kumaran, Xiao, Wen-Hai, Tyo, Keith E. J., Wang, Yong, Simeon, Fritz, Leonard, Effendi, Mucha, Oliver, Phon, Too Heng, Pfeifer, Blaine, Stephanopoulos, Gregory
• Format: Journal Article
• Language: English
• Published: United States American Association for the Advancement of Science 01.10.2010; The American Association for the Advancement of Science
• Subjects: Alkenes - metabolism; Biochemistry; Bioreactors; Biosynthesis; Cellular metabolism; Chemical engineering; Cytochrome P-450 Enzyme System - genetics; Cytochrome P-450 Enzyme System - metabolism; Diterpenes - metabolism; Drugs; E coli; Enzymes; Erythritol - analogs & derivatives; Erythritol - metabolism; Escherichia coli; Escherichia coli K12 - enzymology; Escherichia coli K12 - genetics; Escherichia coli K12 - metabolism; Farnesyltranstransferase - genetics; Farnesyltranstransferase - metabolism; Fermentation; Genetic Engineering; Hemiterpenes - metabolism; Indoles; Indoles - metabolism; Isomerases - genetics; Isomerases - metabolism; Metabolic Networks and Pathways - genetics; Metabolomics; Modular; Modules; NADPH-Ferrihemoprotein Reductase - genetics; NADPH-Ferrihemoprotein Reductase - metabolism; Operons; Optimization; Organophosphorus Compounds - metabolism; Oxidation-Reduction; Pacific yew; Paclitaxel - biosynthesis; Pathways; Pharmacology; Plasmids; Recombinant Fusion Proteins - metabolism; Searching; Streams; Sugar Phosphates - metabolism; Taxoids - metabolism; Taxol; Taxus - enzymology; Taxus brevifolia; Terpenes - metabolism; Terpenoids
• ISSN: 0036-8075; 1095-9203
• DOI: 10.1126/science.1191652

Taxol (paclitaxel) is a potent anticancer drug first isolated from the Taxus brevifolia Pacific yew tree. Currently, cost-efficient production of Taxol and its analogs remains limited. Here, we report a multivariate-modular approach to metabolic-pathway engineering that succeeded in increasing titers of taxadiene—the first committed Taxol intermediate—approximately 1 gram per liter (~15,000-fold) in an engineered Escherichia coli strain. Our approach partitioned the taxadiene metabolic pathway into two modules: a native upstream methylerythritol-phosphate pathway forming isopentenyl pyrophosphate and a heterologous downstream terpenoid-forming pathway. Systematic multivariate search identified conditions that optimally balance the two pathway modules so as to maximize the taxadiene production with minimal accumulation of indole, which is an inhibitory compound found here. We also engineered the next step in Taxol biosynthesis, a P450-mediated 5α-oxidation of taxadiene to taxadien-5α-ol. More broadly, the modular pathway engineering approach helped to unlock the potential of the pathway for the engineered production of terpenoid natural products.