Abstract:[Background] Acetyl-CoA is an important intermediate for lycopene synthesis in Saccharomyces cerevisiae, and acetyl-CoA in cytosol is mainly derived from acetic acid catalyzed by acetyl-CoA synthase.[Objective] To improve cell growth and lycopene production by increasing the content of intracellular acetyl-CoA through adding acetate combined with regulation of acetic acid stress-responsive gene. [Methods] Lycopene recombinant yeast strains overexpressing acetyl-CoA synthase (acs2) were added with 10 g/L acetate during the fermentation process. The transcriptomic analysis was combined to excavate acetic acid stress-responsive genes for single and combined regulation. [Results] After adding acetate, the lycopene content of the recombinant Y02 strain increased by 19.14%, but cell growth was suppressed. The results of the transcriptional analysis indicated that the expression levels of adk2, fap7, hem13, elo3, pdc5, set5, pmt5, hst4, clb2, and swe1 were increased significantly. Therefore, a single-gene and dual-gene were overexpressed in the Y02 strain. It was found that the growth of Y02-set5-hst4 was significantly improved in the presence of acetate. At the same time, the intracellular acetyl-CoA concentration was increased by 78.21%, and the lycopene content and yield reached 12.62 mg/g-DCW and 108.67 mg/L, respectively, which were increased by 42.76% and 67.13%, respectively, as compared with the control strain Y02. In addition, the expression levels of key genes erg12, erg20, and hmg1 in the mevalonate pathway were increased by 1.70, 1.44, and 1.96 folds, respectively, as compared with the control strain. [Conclusion] On the basis of overexpression of acs2, the overexpression of set5 and hst4 can improve the tolerance of yeast to acetic acid stress and increase the synthesis level of acetyl-CoA and the metabolic flux of mevalonate pathway, thus promoting the synthesis of lycopene. The results of this study provide valuable references for the metabolic engineering of other isoprenoid products.
