Integration of a CO₂-fixation pathway and PAN2 truncation in Saccharomyces cerevisiae for low-carbon single-cell protein production

Abstract

Anthropogenic carbon dioxide (CO₂) emissions drive global climate change, motivating the development of bioprocesses that improve carbon utilization and enable CO₂ recycling. In this study, we developed a CO₂-fixing Saccharomyces cerevisiae chassis for single-cell protein (SCP) production using xylose derived from cellulosic biomass as a carbon source. A RuBisCO- based CO₂-fixation pathway was previously integrated into a xylose-utilizing strain, enabling the routing of CO₂ into central metabolism. Flux balance analysis combined with ¹³C-based intracellular metabolite analysis verified the assimilation of externally supplied CO₂ into central metabolism, suggesting the potential for assimilation of fermentation-derived CO₂ and improved carbon utilization during SCP production. To enhance SCP production, the PAN2 gene encoding a component of the poly(A)-ribonuclease complex, previously associated with increased global protein production in S. cerevisiae, was truncated in the CO₂-fixing strain. Under anaerobic conditions, the engineered strain exhibited a significant increase in cellular protein content, accompanied by an overall upward trend in amino acid levels relative to the parental strain. Collectively, these results metabolically verify RuBisCO-mediated CO₂-fixation in yeast strain and demonstrate the feasibility of coupling CO₂-fixation to SCP production. This work also provides insights into the potential of integrating CO₂-fixation with renewable carbon metabolism and establishing a proof-of-concept platform for the development of low-carbon yeast bioprocess.

Keywords

Carbon dioxide fixation / Flux balance analysis / PAN2 truncation / Xylose fermentation

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Funding

National Research Foundation of Korea(RS-2024- 00439872)

RIGHTS & PERMISSIONS

Jiangnan University