Ribosomal Protein RpsL Regulates Butenyl-spinosyn Synthesis in Saccharopolyspora pogona
SU Chang1,3, WANG Jing-Nan1,2, GUO Chao1, XU Zhou-Qin1, LI Chun2,4, ZHANG Gen-Lin3, WANG Chao1,*
1 Academy of National Food and Strategic Reserves Administration, Beijing 100037, China; 2 School of Chemistry and Chemical Engineering, Beijing Institute of Technology, Beijing 100081, China; 3 School of Chemistry and Chemical Engineering, Shihezi University, Shihezi 832003, China; 4 Department of Chemical Engineering, Tsinghua University, Beijing 100084, China
Abstract：Butenyl-spinosyn, a secondary metabolite produced by fermentation of Saccharopolyspora pogona, is a green bio-insecticide. However, due to the low yield of fermentation and can not meet the needs of industrial production, the breeding of strains with high production performance of butenyl-spinosyn is the key to realize industrial production. In this study, ribosomal protein S12 (RpsL) and RpsL mutant gene overexpression engineering strains Q8 (rpsl)、W43 (rpslK88R)、W44 (rpslR86P)、W45 (rpslK88E)和W46 (rpslK43N) were constructed were constructed using Saccharopolyspora pogona as the base cells. The effect of RpsL on the synthesis of butenyl-spinosyn was studied by shaking flask fermentation and comparison of RpsL protein structure. The results showed that the engineered strains Q8, W43, W44, W45, W46 increased the yield of butenyl-spinosyn to 275%, 146%, 111%, 122% and reduced to 75%, respectively. The overexpression of rpsl and rpslK88R genes increased the transcription level of the synthetic pathway genes, and thus significantly increased the yield of butenyl-spinosyn. At the same time, homologous modeling of the two was conducted, and it was found that RpsLK88R mutant structure had a larger deflection of the side chain compared with RpsL, which affected the binding effect to mRNA and thus reduced the ribosome translation efficiency, so its yield extraction effect was slightly lower than RpsL. This study provides a reference for protein engineering RpsL to improve the yield of butenyl-spinosyn.
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