Effects of gene expression for pyruvate and phosphoketolase bypass on production of terpenoids in the yeast Yarrowia lipolyticaMarkuš, Tadej (Avtor) Petrovič, Uroš (Mentor) Yarrowia lipolyticabeta-carotenelipidscarnosic acidmetabolic bypasspyruvate bypassphosphoketolasephosphoketolase bypasscarnitinecarnitine shuttleOur research focused on developing and engineering a terpenoid-producing Y. lipolytica strain to evaluate the impact of central carbon modifications on the production of cytosolic acetyl-CoA. First, our aim was to establish carnosic acid production in Y. lipolytica; however, our efforts were unsuccessful, and gene expression analysis revealed we were unable to successfully express the ATR1 gene, which was needed for the correct functioning of the cytochrome P450 monooxygenases. On the other hand, we successfully prepared a set of beta-carotene-producing parental strains, each with a different capacity for its production, which we used down the line to evaluate central carbon metabolism modifications. We evaluated two gene combinations for the pyruvate bypass, one for the phosphoketolase bypass, and three for the carnitine shuttle. Upon introducing all bypasses in the beta-carotene-producing parental strains, we discovered that most bypasses negatively impacted beta-carotene production. On the other hand, when we examined lipid production, most of the modifications that negatively impacted beta-carotene production positively impacted lipid production. The highest lipid content we measured was 0.61 ± 0.04 glipids/gDCW, demonstrated by the strains carrying the YlCAT2 gene. At the same time, we observed the highest lipid efficiency improvement for the strains carrying the YlPDC1, YlALD3, and YlACS1 genes of the pyruvate bypass. The inverse correlation between the biosynthesis of lipids and beta-carotene production has not been observed before and hints at possible regulation response at higher acetyl-CoA availability.20252025-01-31 07:15:16Doktorsko delo/naloga166920VisID: 256003COBISS_ID: 224641027sl