Application of an Optogenetic Switch for Controlling the Central Carbon Metabolism in Escherichia coli

Optimization of metabolic flux distribution on pathways is an important approach for efficient bio-production using microorganisms. In particular, dynamic controlling the metabolic flux is a promising way for enhancing the productivity. To adjust the flux distribution, fine-tuning the expression level of metabolic enzymes is highly required, and inducible systems using chemicals have been frequently used. However, such chemically-inducible systems cannot be applied as a dynamic on/off control of metabolic pathways, because it is difficult to immediately remove added chemicals from the culture. Optogenetic switch is an attractive technique for controlling gene expression levels and for realizing a dynamic bio-process control, because light can readily be applied and removed from the culture multiple times.

In the present study, we applied a CcaS/CcaR optogenetic system to control a flux ratio of glycolytic pathways such as Embden “Meyerhof“Parnas (EMP) and oxidative pentose phosphate (OPP) pathways in Escherichia coli. The expression of phosphoglucose isomerase (Pgi) that pulls glucose-6-phosphate at the branch point into the EMP pathway was regulated. The output EMP flux from the switch was optimized by adjusting plasmid copy number and by tagging Pgi with ssRA protein degradation signal. We evaluated the light-induced redirection of glycolytic flux with ¹³C-metabolic flux analysis. The EMP:OPP flux ratio was controlled to 50:49 and 0.5:99 by exposure to green and red light, respectively [1]. Furthermore, we also demonstrated the CcaS/CcaR optogenetic switch for controlling the carbon flow between the EMP and methylglyoxal pathway in E. coli [2]. These switches would be useful tools for metabolic engineering.

[1] Tandar et al. Optogenetic switch for controlling the central metabolic flux of Escherichia coli. Metab Eng. 2019; 55:68-75.

[2] Senoo et al. Light-inducible flux control of triosephosphate isomerase on glycolysis in Escherichia coli. Biotechnol Bioeng. In press.