Identification and metabolomic analysis of chemical elicitors for tacrolimus accumulation in Streptomyces tsukubaensis

Cheng Wang, Di Huang, Shaoxiong Liang

Tacrolimus is a widely used immunosuppressive agent in the treatment of various clinical diseases. However, the low fermentation yield seriously limits its further application. To stimulate tacrolimus synthesis, nine chemical elicitors of five groups were evaluated for their effects on tacrolimus accumulation in S. tsukubaensis. The results showed that sodium butyrate (SB), dimethylsulfoxide (DMSO), and LaCl3 could increase tacrolimus accumulation by more than 30%. Cumulative effects of different chemical elicitors exhibited that the highest tacrolimus yield was improved by 64.7% (303.60 mg/L) in DMSO and La treatment, compared to the control. To decipher possible response mechanism, a weighted correlation network analysis (WGCNA) based on metabolomics was employed and datasets showed 13 distinct metabolic modules and 16 hub metabolites were possibly related to the stimulatory roles of DMSO, La, SB, and their combination treatments. The pathway analysis further exhibited that central carbon metabolism, amino acid metabolism, and fatty acid metabolism showed significant differences in the above chemical elicitor treatments. Thereinto, the carboxylation of propionyl-CoA from isoleucine and methionine degradation was first confirmed to be the main source of methylmalonyl-CoA by RT-PCR analysis in DMSO and La treatment. By further strengthening of the supply of methylmalonyl-CoA precursor in DMSO and La treatment, the final tacrolimus yield could reach to 372.12 mg/L, 2.02-fold higher than the control. To our knowledge, this is the first study to unveil the potential mechanism of different chemical elicitor stresses in S. tsukubaensis based on metabolomics, and the established information provide valuable guidance for further improving tacrolimus production.