State Key Laboratory of Bioreactor Engineering, East China University of Science and Technology, Shanghai, China.
Qingdao Innovation Institute of East China University of Science and Technology, Qingdao, China.
Biotechnol J. 2024 Mar;19(3):e2300683. doi: 10.1002/biot.202300683.
Acremonium chrysogenum is the major industrial producer of cephalosporin C (CPC), which is used as raw material for the production of significant cephalosporin antibiotics. Due to the lack of diverse promoter elements, the development of metabolic engineering transformation is relatively slow, resulting in a limited improvement on CPC production. In this study, based on the analysis of the transcriptome profile, 27 candidate promoters were selected to drive the expression of the reporter genes. The promoter activities of this library ranged from 0.0075 to 101 times of the control promoter P . Simultaneously, a rapid screening method for potential bidirectional promoters was developed and 4 strong bidirectional promoters from 27 candidate options were identified and validated. Finally, the Golden Gate method was employed to combine promoter modules from the library with various target genes. Through a mixed transformation and screening process, high-yielding strains AG-6, AG-18, and AG-41 were identified, exhibiting an increase in CPC production of 30%, 35%, and 29%, respectively, compared to the control strain Ac-∆axl2:: eGFP. Therefore, the utilization of this promoter library offers a broader range of synthetic biology toolkits for the genetic engineering transformation of A. chrysogenum, thus establishing a solid foundation for the precise regulation of gene expression.
顶头孢霉（Acremonium chrysogenum）是头孢菌素 C（cephalosporin C，CPC）的主要工业生产菌，CPC 是生产重要头孢菌素抗生素的原料。由于缺乏多样化的启动子元件，代谢工程改造的发展相对缓慢，导致 CPC 产量的改善有限。在这项研究中，基于转录组谱的分析，选择了 27 个候选启动子来驱动报告基因的表达。该文库的启动子活性范围从对照启动子 P 的 0.0075 到 101 倍不等。同时，开发了一种快速筛选潜在双向启动子的方法，从 27 个候选选项中鉴定并验证了 4 个强双向启动子。最后，采用 Golden Gate 方法将文库中的启动子模块与各种靶基因结合。通过混合转化和筛选过程，鉴定出高产菌株 AG-6、AG-18 和 AG-41，与对照菌株 Ac-∆axl2::eGFP 相比，CPC 的产量分别提高了 30%、35%和 29%。因此，该启动子文库的利用为顶头孢霉的遗传工程改造提供了更广泛的合成生物学工具包，为基因表达的精确调控奠定了基础。
