title: Alleviating Redox Imbalance Enhances 7-Dehydrocholesterol Production in Engineered Saccharomyces cerevisiae.
abstract: Maintaining redox balance is critical for the production of heterologous secondary metabolites, whereas on various occasions the native cofactor balance does not match the needs in engineered microorganisms. In this study, 7-dehydrocholesterol (7-DHC, a crucial precursor of vitamin D3) biosynthesis pathway was constructed in Saccharomyces cerevisiae BY4742 with endogenous ergosterol synthesis pathway blocked by knocking out the erg5 gene (encoding C-22 desaturase). The deletion of erg5 led to redox imbalance with higher ratio of cytosolic free NADH/NAD+ and more glycerol and ethanol accumulation. To alleviate the redox imbalance, a water-forming NADH oxidase (NOX) and an alternative oxidase (AOX1) were employed in our system based on cofactor regeneration strategy. Consequently, the production of 7-dehydrocholesterol was increased by 74.4% in shake flask culture. In the meanwhile, the ratio of free NADH/NAD+ and the concentration of glycerol and ethanol were reduced by 78.0%, 50.7% and 7.9% respectively. In a 5-L bioreactor, the optimal production of 7-DHC reached 44.49(±9.63) mg/L. This study provides a reference to increase the production of some desired compounds that are restricted by redox imbalance.
authors.affiliation: Key Laboratory of Systems Bioengineering (Ministry of Education), Tianjin University, Tianjin, China; SynBio Research Platform, Collaborative Innovation Center of Chemical Science and Engineering (Tianjin), School of Chemical Engineering and Technology, Tianjin University, Tianjin, China.
authors.forename: Wan
authors.initials: W
authors.lastname: Su
authors.affiliation: Key Laboratory of Systems Bioengineering (Ministry of Education), Tianjin University, Tianjin, China; SynBio Research Platform, Collaborative Innovation Center of Chemical Science and Engineering (Tianjin), School of Chemical Engineering and Technology, Tianjin University, Tianjin, China.
authors.forename: Wen-Hai
authors.initials: WH
authors.lastname: Xiao
authors.affiliation: Key Laboratory of Systems Bioengineering (Ministry of Education), Tianjin University, Tianjin, China; SynBio Research Platform, Collaborative Innovation Center of Chemical Science and Engineering (Tianjin), School of Chemical Engineering and Technology, Tianjin University, Tianjin, China.
authors.forename: Ying
authors.initials: Y
authors.lastname: Wang
authors.affiliation: Key Laboratory of Systems Bioengineering (Ministry of Education), Tianjin University, Tianjin, China; SynBio Research Platform, Collaborative Innovation Center of Chemical Science and Engineering (Tianjin), School of Chemical Engineering and Technology, Tianjin University, Tianjin, China.
authors.forename: Duo
authors.initials: D
authors.lastname: Liu
authors.affiliation: Key Laboratory of Systems Bioengineering (Ministry of Education), Tianjin University, Tianjin, China; SynBio Research Platform, Collaborative Innovation Center of Chemical Science and Engineering (Tianjin), School of Chemical Engineering and Technology, Tianjin University, Tianjin, China.
authors.forename: Xiao
authors.initials: X
authors.lastname: Zhou
authors.affiliation: Key Laboratory of Systems Bioengineering (Ministry of Education), Tianjin University, Tianjin, China; SynBio Research Platform, Collaborative Innovation Center of Chemical Science and Engineering (Tianjin), School of Chemical Engineering and Technology, Tianjin University, Tianjin, China.
authors.forename: Ying-Jin
authors.initials: YJ
authors.lastname: Yuan
mesh_terms: D053898:Biosynthetic Pathways / Q000502:physiology*; D003577:Cytochrome P-450 Enzyme System / Q000235:genetics; D017931:DNA Primers / Q000235:genetics; D003684:Dehydrocholesterols / Q000378:metabolism*; D004875:Ergosterol / Q000096:biosynthesis; D005285:Fermentation; D055786:Gene Knockout Techniques; D005818:Genetic Engineering / Q000379:methods*; D007218:Industrial Microbiology / Q000379:methods*; D030781:Organisms, Genetically Modified / Q000378:metabolism*; D010084:Oxidation-Reduction; D010957:Plasmids / Q000235:genetics; D016133:Polymerase Chain Reaction; D012441:Saccharomyces cerevisiae / Q000378:metabolism*; D029701:Saccharomyces cerevisiae Proteins / Q000235:genetics
chemical_list: D017931:DNA Primers; D003684:Dehydrocholesterols; D029701:Saccharomyces cerevisiae Proteins; D003577:Cytochrome P-450 Enzyme System; C016705:7-dehydrocholesterol; C066577:ERG5 protein, S cerevisiae; D004875:Ergosterol