Presentation Information
[3SBT-04]Optogenetic metabolic engineering for manipulating metabolic flux and driving force
○Yoshihiro Toya1 (1. The University of Osaka (Japan))
Keywords:
Metabolic flux,Optogenetics,Rhodopsin,ATP generation,Escherichia coli
To achieve a sustainable society, we need to replace certain material production processes that rely on fossil resources with microbial fermentation processes that utilize renewable biomass resources. Such bio-manufacturing requires the engineering of microorganisms to improve the conversion efficiency from substrates to target compounds, as well as the production rate.
In metabolic engineering, many efforts have been dedicated to manipulating metabolic flow in pathways to achieve the ideal metabolic state for producing our desirable target by employing metabolic switch at the pathway branch. To facilitate chemical production during the non-growing stationary phase, switches are needed to change metabolism between growth and production modes. We are aiming to use a reversible switch that utilizes light as a signal, to maintain cellular activity during the stationary phase. We developed optogenetic metabolic switches using green/red light-responsive CcaS/R or blue light-responsive EL222 and demonstrated the manipulation of metabolic flux ratios in the glycolytic pathways of Escherichia coli (1). In this talk, I present a new optogenetic switch using antisense RNA and orthogonal protease to establish versatile control of flux distribution. Using the switches, we demonstrated enhanced mevalonate production by repeating the production and growth modes during the stationary phase.
ATP is an important energy cofactor that drives various metabolic reactions. Improving ATP generation is a promising strategy for producing target compounds that consume ATP in their synthesis. We focused on a proton-pump rhodopsin, membrane protein found in organisms like haloarchaea, as a tool to increase ATP generation. Because rhodopsins generate a proton gradient across the membrane, rhodopsin-expressing E. coli cells can produce ATP when exposed to light even in the absence of energy substrates like glucose. Using the light-driven ATP generation technique, an ATP-consuming conversion of mevalonate to isoprenol in E. coli was successfully demonstrated (2). In this talk, I will introduce an acetone production using acetate as a sole carbon source. Two acetate molecules are converted into one acetone molecule consuming one ATP. The carbon yield of acetone production, corrected for volatilization effects, was 0.72 C-mol/C-mol under light conditions, that was higher than the theoretical yield without considering light-driven ATP generation. The use of light in bioprocesses is an effective way to expand possibilities even for heterotrophic microorganisms.
(1) Biotechnol Bioeng. 2024; 121: 1016-1025
(2) ACS Synth Biol. 2022; 11: 3966-3972.
In metabolic engineering, many efforts have been dedicated to manipulating metabolic flow in pathways to achieve the ideal metabolic state for producing our desirable target by employing metabolic switch at the pathway branch. To facilitate chemical production during the non-growing stationary phase, switches are needed to change metabolism between growth and production modes. We are aiming to use a reversible switch that utilizes light as a signal, to maintain cellular activity during the stationary phase. We developed optogenetic metabolic switches using green/red light-responsive CcaS/R or blue light-responsive EL222 and demonstrated the manipulation of metabolic flux ratios in the glycolytic pathways of Escherichia coli (1). In this talk, I present a new optogenetic switch using antisense RNA and orthogonal protease to establish versatile control of flux distribution. Using the switches, we demonstrated enhanced mevalonate production by repeating the production and growth modes during the stationary phase.
ATP is an important energy cofactor that drives various metabolic reactions. Improving ATP generation is a promising strategy for producing target compounds that consume ATP in their synthesis. We focused on a proton-pump rhodopsin, membrane protein found in organisms like haloarchaea, as a tool to increase ATP generation. Because rhodopsins generate a proton gradient across the membrane, rhodopsin-expressing E. coli cells can produce ATP when exposed to light even in the absence of energy substrates like glucose. Using the light-driven ATP generation technique, an ATP-consuming conversion of mevalonate to isoprenol in E. coli was successfully demonstrated (2). In this talk, I will introduce an acetone production using acetate as a sole carbon source. Two acetate molecules are converted into one acetone molecule consuming one ATP. The carbon yield of acetone production, corrected for volatilization effects, was 0.72 C-mol/C-mol under light conditions, that was higher than the theoretical yield without considering light-driven ATP generation. The use of light in bioprocesses is an effective way to expand possibilities even for heterotrophic microorganisms.
(1) Biotechnol Bioeng. 2024; 121: 1016-1025
(2) ACS Synth Biol. 2022; 11: 3966-3972.
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