Boosting Ascomycin Production through n-Butanol Addition in the Fermentation Process of Streptomyces hygroscopicus var. ascomyceticus ATCC 14891

Ascomycin (FK520) is a macrolide antibiotic known for its immunosuppressive activities. In this study, we screened several short-chain alcohols to enhance titer of FK520 in Streptomyces hygroscopicus var. ascomyceticus ATCC 14891, with a particular focus on n-butanol addition. After optimizing the n-butanol addition process, we achieved a FK520 yield of 569.37 mg/L with 0.8% n-butanol addition at 27 h, representing a 1.72-fold increase compared with the control group. We subsequently found that ROS levels of n-butanol addition group reached 1.49x105 RFU/g biomass at 29 h, which was 3.02 times higher than the control group. The 0.8% n-butanol addition also promote the accumulation of biosynthesis precursors for FK520 production. The highest ethylmalonyl-CoA content surged to 93.1 nmol/g DCW at 48 h, marking a 5.3-fold increase. Likewise, the highest methylmalonyl-CoA and malonyl-CoA levels increased remarkable 4.33-fold and 3.33-fold compared with the control group at 72 h and at 120 h, respectively. Then, we explored the effects of oxygen supply on FK520 production with n-butanol addition, and improving oxygen supply caused a significant increase of FK520 in shake flask fermentation. Our research has revealed that addition of short-chain alcohols can regulate carbon flux toward FK520 biosynthesis by supplementing various CoA-esters, including ethylmalonyl-CoA, methylmalonyl-CoA, and malonyl-CoA.