Calcium-Phosphate Combination Enhances Spinosad Production in Saccharopolyspora spinosa via Regulation of Fatty Acid...

Phosphate concentration above 10 mM reduces the production of many secondary metabolites; however, the phenomenon is not mechanistically understood yet. Specifically, the problem of phosphorus limitation in antibiotic production remains unresolved. This study investigates the phosphorus inhibition effect on spinosad production and alleviates it by calcium and phosphate supplementation to fermentation media. Furthermore, we examined the mechanism of fatty acids-induced increase in polyketides production. Four phosphates that were supplemented into the fermentation media include NaH₂PO₄, Na₂HPO₄, KH₂PO₄, and K₂HPO₄ and NaH₂PO₄ was found to be the most effective phosphate. Under the optimal phosphate condition of supplementing 20 mM NaH₂PO₄ on the fourth day and 5 g/L CaCO₃, the maximal spinosad production reached 520 mg/L, showing a 1.65-fold increase over the control treatment. In the NaH₂PO₄-CaCO₃ system, the de novo fatty acid biosynthesis was significantly downregulated while spinosad biosynthesis and β-oxidation were upregulated. The coordination of de novo fatty acid biosynthesis and β-oxidation promoted intracellular acetyl-CoA concentration. The results demonstrate that NaH₂PO₄-CaCO₃ combined addition is a simple and effective strategy to alleviate phosphorus inhibition effect through the regulation of fatty acid metabolism and accumulation of immediate precursors. This information improves our understanding of phosphates' influence on the large-scale production of polyketides.