Work

Coupling genome-wide continuous perturbation with biosensor screening reveals the potential targets in yeast isopentanol synthesis network

Year: 2024

Type: article

Abstract: The increasing consumption of fossil fuels is contributing to global resource depletion and environmental pollution. Branched-chain higher alcohols, such as isopentanol and isobutanol, have attracted significant attention as next-generation biofuels. Biofuel production through microbial fermentation offers a green, sustainable, and renewable alternative to chemical synthesis. While enhanced production of isopentanol has been achieved in a variety of chassis, the fermentation yield has not yet reached levels suitable for industrial-scale production. In this study, we employed a continuous perturbation tool to construct a genome-scale perturbation library, combined with an isopentanol biosensor to screen for high-yielding mutants. We identified five high-yielding mutants, each exhibiting an increased glucose conversion rate and isopentanol titer. The F2 strain, in particular, achieved an isopentanol titer of 1.57 ± 0.014 g/L and a yield of 14.04 ± 0.251 mg/g glucose (10% glucose), surpassing the highest values reported to date in engineered Saccharomyces cerevisiae . Systematic transcriptome analysis of the isopentanol synthesis, glycolysis, glycerol metabolism, and ethanol synthesis pathways revealed that MPC , OAC1 , BAT2 , GUT2 , PDC6 , and ALD4 are linked to efficient isopentanol production. Further analysis of differentially expressed genes (DEGs) identified 17 and 12 co-expressed DEGs (co-DEGs) in all mutants and the two second-round mutants, respectively. In addition, we validated the knockout or overexpression of key co-DEGs. Our results confirmed the critical roles of HOM3 and DIP5 in isopentanol production, along with genes associated with the aerobic respiratory chain ( SDH3 , CYT1 , COX7 , ROX1 , and ATG41 ) and cofactor balance ( BNA2 and NDE1 ). Additionally, functional analysis of the co-DEGs revealed that MAL33 is associated with the synthesis of branched-chain higher alcohols, expanding the intracellular metabolic network and offering new possibilities for green, cost-effective biofuel production.

Source: Synthetic and Systems Biotechnology

DOI: 10.1016/j.synbio.2024.12.010

Authors: Qi Xiao, Jingjing Shi, Lixian Wang, Guoping Zhao, Yanfei Zhang

Institutions: Chinese Academy of Sciences, Tianjin Institute of Industrial Biotechnology, University of Chinese Academy of Sciences, Center for Excellence in Molecular Plant Sciences

Language: English

Topic: Microbial Metabolic Engineering and Bioproduction

Subfield: Molecular Biology

Field: Biochemistry, Genetics and Molecular Biology

Domain: Life Sciences

SDG: Affordable and clean energy

Open Access status: gold

Estimated APC paid: $1,500