Enhanced Metabolite Productivity of Escherichia coli Adapted to Glucose M9 Minimal Medium

Peter Rugbjerg,Adam M Feist,Morten Otto Alexander Sommer

doi:10.3389/fbioe.2018.00166

Abstract

High productivity of biotechnological strains is important to industrial fermentation processes and can be constrained by precursor availability and substrate uptake rate. Adaptive laboratory evolution (ALE) of Escherichia coli MG1655 to glucose minimal M9 medium has been shown to increase strain fitness, mainly through a key mutation in the transcriptional regulator rpoB, which increases flux through central carbon metabolism and the glucose uptake rate. We wanted to test the hypothesis that a substrate uptake enhancing rpoB mutation can translate to increased productivity in a strain possessing a heterologous metabolite pathway. When engineered for heterologous mevalonate production, we found that E. coli rpoB E672K strains displayed 114–167% higher glucose uptake rates and 48–77% higher mevalonate productivities in glucose minimal M9 medium. This improvement in heterologous mevalonate productivity of the rpoB E672K strain is likely mediated by the elevated glucose uptake rate of such strains, which favors overflow metabolism toward acetate production and availability of acetyl-CoA as precursor. These results demonstrate the utility of adaptive laboratory evolution (ALE) to generate a platform strain for an increased production rate for a heterologous product.

Highlights

Metabolic engineering of microbial production strains often benefits from evolutionary optimization strategies when a desired phenotype is selectable (Nielsen and Keasling, 2016)
We utilized the improved glucose uptake rates and higher glycolytic flux of medium-adapted rpoB E672K mutant strains to test if the higher flux translated to the biosynthesis of heterologous mevalonate, a glycolysis-derived metabolite
We focus on the rpoB E672K mutation, which enables higher growth rates in M9 glucose minimal medium, globally affects the strain proteome and dispenses unneeded functionality (Utrilla et al, 2016)

Summary

Introduction

Metabolic engineering of microbial production strains often benefits from evolutionary optimization strategies when a desired phenotype is selectable (Nielsen and Keasling, 2016). Adaptive laboratory evolution (ALE) has been shown to increase strain tolerance to toxic substrates and products and enable improved production yields and titers (Dragosits and Mattanovich, 2013). Since biosynthesis of only a few native products is growth-coupled with optimal energy production (Feist et al, 2010), direct ALE to improve production phenotypes is currently limited, and diverse mutation types instead readily disrupt heterologous product genes during long-term cultivation (Rugbjerg et al, 2018a). The specific productivity (mmol/L/gDW) further accounts for biomass differences, which influences productivity

Methods

Results

Conclusion