Zhejiang Provincial Key Laboratory of Biometrology and Inspection & Quarantine, College of Life Sciences, China Jiliang University, Xueyuan Street, Xiasha Higher Education District, Hangzhou, 310018, Zhejiang Province, People's Republic of China.
Institute for Pharmaceutical Sciences, Pharmaceutical Biology and Biotechnology, University of Freiburg, 79104, Freiburg, Germany.
Curr Microbiol. 2022 Apr 30;79(6):174. doi: 10.1007/s00284-022-02867-9.
Precursor engineering is an effective strategy for the overproduction of secondary metabolites. The polyene macrolide rimocidin, which is produced by Streptomyces rimosus M527, exhibits a potent activity against a broad range of phytopathogenic fungi. It has been predicted that malonyl-CoA is used as extender units for rimocidin biosynthesis. Based on a systematic analysis of three sets of time-series transcriptome microarray data of S. rimosus M527 fermented in different conditions, the differentially expressed acc gene that encodes acetyl-CoA carboxylase (ACC) was found. To understand how the formation of rimocidin is being influenced by the expression of the acc gene and by the concentration of malonyl-CoA, the acc gene was cloned and over-expressed in the wild-type strain S. rimosus M527 in this study. The recombinant strain S. rimosus M527-ACC harboring the over-expressed acc gene exhibited better performances based on the enzymatic activity of ACC, intracellular malonyl-CoA concentrations, and rimocidin production compared to S. rimosus M527 throughout the fermentation process. The enzymatic activity of ACC and intracellular concentration of malonyl-CoA of S. rimosus M527-ACC were 1.0- and 1.5-fold higher than those of S. rimosus M527, respectively. Finally, the yield of rimocidin produced by S. rimosus M527-ACC reached 320.7 mg/L, which was 34.0% higher than that of S. rimosus M527. These results confirmed that malonyl-CoA is an important precursor for rimocidin biosynthesis and suggested that an adequate supply of malonyl-CoA caused by acc gene over-expression led to the improvement in rimocidin production.
前体工程是次级代谢产物过量生产的有效策略。多烯大环内酯类化合物里莫西丁由嗜热链霉菌 M527 产生，对广泛的植物病原真菌具有很强的活性。据预测，丙二酰辅酶 A 被用作里莫西丁生物合成的延伸单位。基于对嗜热链霉菌 M527 在不同条件下发酵的三组时间序列转录组微阵列数据的系统分析，发现了编码乙酰辅酶 A 羧化酶 (ACC) 的差异表达 acc 基因。为了了解 acc 基因的表达和丙二酰辅酶 A 的浓度如何影响里莫西丁的形成，本研究在野生型嗜热链霉菌 M527 中克隆并过表达了 acc 基因。与野生型嗜热链霉菌 M527 相比，携带过表达 acc 基因的重组菌嗜热链霉菌 M527-ACC 在整个发酵过程中表现出更好的性能，其 ACC 的酶活性、细胞内丙二酰辅酶 A 浓度和里莫西丁产量均有所提高。ACC 的酶活性和细胞内丙二酰辅酶 A 浓度分别比嗜热链霉菌 M527 高 1.0 倍和 1.5 倍。最终，重组菌嗜热链霉菌 M527-ACC 产生的里莫西丁产量达到 320.7mg/L，比嗜热链霉菌 M527 提高了 34.0%。这些结果证实了丙二酰辅酶 A 是里莫西丁生物合成的重要前体，并且 acc 基因过表达导致丙二酰辅酶 A 的充足供应，从而提高了里莫西丁的产量。
