Light-driven eosin Y-Ralstonia eutropha biohybrid for CO2 conversion to acetoin via specific photo-induced electron transfer and metabolic engineering

Abstract

Whole-cell photosynthetic biohybrid systems offer a promising route for CO₂ conversion into value-added chemicals. However, many existing systems suffer from inefficient electron transfer to intracellular catalytic CO₂ center, which arises from non-specific charge transfer between photosensitizer and microbe. Herein, we developed an efficient eosin Y-Ralstonia eutropha biohybrid system that achieved targeted electron transfer for powering CO₂ reduction into acetoin, a valuable platform chemical with broad applications. By spontaneously attaching eosin Y to the membrane-bound hydrogenase (MBH) of R. eutropha, we enabled direct and specific electron flow. The biohybrid system demonstrated sustainable and efficient light-energized CO₂ conversion to acetoin, even surpassing the yields from H₂-supplied autotrophic fermentation, which is considered the optimal source of reducing equivalents and energy for CO₂ fixation by R. eutropha. Mechanistic investigations indicated that the photo-induced electrons from eosin Y were transferred to MBH, resulting in the formation of H₂ intermediate in periplasm, which was subsequently oxidized by cytosolic soluble hydrogenase to generate NADH for energizing CO₂ fixation. To further enhance efficiency, metabolic engineering was applied to boost ATP synthesis by introducing a non-oxygen-dependent proton pump (Gloeobacter rhodopsin), while blocking L-lactate and acetate biosynthesis pathways to divert carbon flux toward acetoin production. The engineered eosin Y-R. eutropha biohybrid system achieved an acetoin yield of 1.41 ± 0.06 mM, representing 2.07 times greater than that of H₂-supplied autotrophic fermentation. This system exemplifies a sustainable, light-driven CO₂ conversion process, contributing significantly to the advancement of sustainable biocatalytic processes in the realms of green chemistry and CO₂ management.