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Isoprenoid Pathway Optimization for Taxol Precursor Overproduction in Escherichia coli

P. K. Ajikumar,Wenhai Xiao,Keith E. J. Tyo,Yong Wang,F. Simeon,E. Leonard,Oliver Mucha,Too Heng Phon,B. Pfeifer,G. Stephanopoulos

Published 2010 in Science

ABSTRACT

Toward Microbial Taxol Taxol, a minor chemical constituent of yew tree bark, has provided a potent cancer treatment. Production methods presently rely on plant cell cultures. Ajikumar et al. (p. 70; see the Perspective by Liu and Khosla) engineered Escherichia coli cells to produce a key taxol precursor, in which the polycyclic carbon skeleton is intact. The approach relied on optimizing the relative activity of two pathways, the first of which synthesized isoprenoid building blocks that were then stitched together with the second pathway. Accumulation of indole as a by-product inhibited the isoprenoid pathway—an insight that should facilitate more efficient engineered biosynthesis of a wide range of commercially important isoprenoid derivatives. Engineered bacteria synthesize a precursor to a potent cancer drug, raising prospects for efficient production of analogs. 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 (MEP) 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 MEP pathway for the engineered production of terpenoid natural products.

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