Combination of Entner-Doudoroff Pathway with MEP Increases Isoprene Production in Engineered Escherichia coli

Huaiwei Liu,Kris Niño G Valdehuesa,Grace M Nisola,Kristine Rose M Ramos,Wook-Jin Chung,Won–Keun Lee,Si Jae Park,Yuanzhang Sun,Tanya Parish

doi:10.1371/journal.pone.0083290

Abstract

Embden-Meyerhof pathway (EMP) in tandem with 2-C-methyl-D-erythritol 4-phosphate pathway (MEP) is commonly used for isoprenoid biosynthesis in E. coli. However, this combination has limitations as EMP generates an imbalanced distribution of pyruvate and glyceraldehyde-3-phosphate (G3P). Herein, four glycolytic pathways—EMP, Entner-Doudoroff Pathway (EDP), Pentose Phosphate Pathway (PPP) and Dahms pathway were tested as MEP feeding modules for isoprene production. Results revealed the highest isoprene production from EDP containing modules, wherein pyruvate and G3P were generated simultaneously; isoprene titer and yield were more than three and six times higher than those of the EMP module, respectively. Additionally, the PPP module that generates G3P prior to pyruvate was significantly more effective than the Dahms pathway, in which pyruvate production precedes G3P. In terms of precursor generation and energy/reducing-equivalent supply, EDP+PPP was found to be the ideal feeding module for MEP. These findings may launch a new direction for the optimization of MEP-dependent isoprenoid biosynthesis pathways.

Highlights

Isoprenoids, known as terpenes or terpenoids, belong to a large and highly diverse group of compounds which have been utilized or at least have the potential use for biofuels, biopharmaceuticals, nutraceuticals, flavors, fragrances cosmetics, and agrichemicals production [1,2,3,4,5]
All of the four known glycolytic pathways involved in glucose and/or D-xylose metabolism in E. coli, which have been or can be used as methyl-D-erythritol 4-phosphate pathway (MEP) feeding modules for isoprene production were investigated (Figure 2)
In native E. coli, glucose is mainly metabolized by the Embden-Meyerhof pathway (EMP) pathway, which has been widely used in isoprenoid biosynthesis

Summary

Introduction

Isoprenoids, known as terpenes or terpenoids, belong to a large and highly diverse group of compounds which have been utilized or at least have the potential use for biofuels, biopharmaceuticals, nutraceuticals, flavors, fragrances cosmetics, and agrichemicals production [1,2,3,4,5]. The mevalonate pathway (MVA) and 2-C-methyl-D-erythritol 4-phosphate pathway (MEP), have been intensively studied and applied in the microbial production of isoprene. The MVA is mainly present in eukaryotes and Archaea, which commences with the co-condensation of acetyl-CoA to form acetoacetyl-CoA. The MEP which starts with the condensation of pyruvate and glyceraldehyde-3-phosphate (G3P) to form 1-deoxy-D-xylulose-5-phosphate (DXP), is typically found in most bacteria. These two pathways begin with different precursors, both culminate with the production of two universal 5carbon isoprenoid precursors, isopentenyl pyrophosphate (IPP) and dimethylallyl pyrophosphate (DMAPP) [10,11]. The DMAPP can be converted to isoprene by isoprene synthases (IspS), which are cloned from plants and heterologously expressed in microbial hosts [8,9]

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