Chao Wang’s research while affiliated with Technical Institute of Physics and Chemistry, Chinese Academy of Sciences and other places

Butenyl-spinosyn discovered from Saccharopolyspora pogona, is a broad-spectrum bioinsecticide. In order to further improve the production, the fermentation medium of a high-production strain Sa. pogona ASAGF30A11 obtained by mutagenesis, was optimized by adding different species and concentrations vegetable oil. In our study, the effect of peanut oil on the growth and production was proved by monitoring the growth curves, key gene transcription level and content of acyl-CoA. After adding 10 g/L of peanut oil, the additional carbon sources redirected the carbon flux toward strain growth, inhibiting the synthesis of butenyl-spinosyn, while increasing biomass by approximately 1.5-fold. However, when adding 1 g/L of peanut oil, it functions as a surfactant, greatly promoting the synthesis of butenyl-spinosyn, resulting in a 1.52-fold increase in production. The research provides a promising strategy to improve butenyl-spinosyn production.

Butenyl-spinosyn, derived from Saccharopolyspora pogona, is a broad-spectrum and effective bioinsecticide. However, the regulatory mechanism affecting butenyl-spinosyn synthesis has not been fully elucidated, which hindered the improvement of production. Here, a high-production strain S. pogona H2 was generated by Cobalt-60 γ-ray mutagenesis, which showed a 2.7-fold increase in production compared to the wild-type strain S. pogona ASAGF58. A comparative transcriptomic analysis between S. pogona ASAGF58 and H2 was performed to elucidate the high-production mechanism that more precursors and energy were used to synthesize of butenyl-spinosyn. Fortunately, a PurR family transcriptional regulator TF00350 was discovered. TF00350 overexpression strain RS00350 induced morphological differentiation and butenyl-spinosyn production, ultimately leading to a 5.5-fold increase in butenyl-spinosyn production (141.5 ± 1.03 mg/L). Through transcriptomics analysis, most genes related to purine metabolism pathway were downregulated, and the butenyl-spinosyn biosynthesis gene was upregulated by increasing the concentration of c-di-GMP and decreasing the concentration of c-di-AMP. These results provide valuable insights for further mining key regulators and improving butenyl-spinosyn production. Key points • A high production strain of S. pogona H2 was obtained by⁶⁰Co γ-ray mutagenesis. • Positive regulator TF00350 identified by transcriptomics, increasing butenyl-spinosyn production by 5.5-fold. • TF00350 regulated of butenyl-spinosyn production by second messengers.

Butenyl-spinosyn is an insecticide produced by Saccharopolyspora pogona, which has both the safety of biological pesticides and the quick-acting properties of chemical pesticides. However, the low synthesis efficiency of butenyl-spinosyn by wild-type strains cannot meet the need of industrial production, obtaining high-yield strains is an urgent problem. At present, there are few related studies on butenyl-spinosyn. The spinosyn produced by Saccharopolyspora spinosa has a similar structural and biosynthetic pathway. This article describes the basic characteristics of them, draws on the research experience of spinosyn, summarizes the available strategies for breeding and modifying high-yield butenyl-spinosyn strains, including mutagenesis methods and precise genetic engineering methods such as metabolic flux regulation, pathway genes regulation, transcriptional regulation, heterologous expression, which may provide ideas for further research of butenyl-spinosyn.