通过异质表达和工艺优化,通过高度生产稳素
Zhengyu Zhang1, Songbai Yang2, Zhenxin Li2
1Department of Biological Medicines & Shanghai Engineering Research Center of Immunotherapeutics, Fudan University School of Pharmacy, 826 Zhangheng Road, Pudong, Shanghai, 201203, People's Republic of China.
Applied microbiology and biotechnology
|July 22, 2023
概括
通过优化其在异质宿主中的生物合成基因集群 (BGC) 来实现抗瘤药物前体稳素的高水平生产. 这一策略使得产量增加了30倍,为工业应用铺平了道路.
科学领域:
- 生物技术是生物技术.
- 微生物工程 微生物工程
- 自然产品的合成自然产品的合成
背景情况:
- 稳素,一个关键的印洛卡巴醇化物,是一种强大的蛋白激酶抑制剂,也是抗瘤药物开发的重要化合物.
- 从历史上看,本地生产商的低产量限制了稳定体的生产,阻碍了其治疗潜力.
- 有效的生产方法对于利用氨酸的抗癌性质至关重要.
研究的目的:
- 开发一种高产量生产系统,用于稳定菌素.
- 确定一种最佳的异质宿主来表达稳体生物合成基因集群 (BGC).
- 通过基因集群放大和发酵优化来增强稳态氨酸的生产.
主要方法:
- 使用CRISPR/Cas9介导的转化关联重组 (TAR) 来捕获稳体BGC.
- 该BGC被引入异种宿主,包括Streptomyces avermitilis,Streptomyces lividans,以及Streptomyces albus. 这三种宿主.
- 多个BGC拷贝被整合到宿主染色体中,并且优化了发酵参数 (pH,葡萄糖养).
主要成果:
- 杆菌 (Streptomyces albus J1074) 成为最有效的宿主,产量为750mg/L.
- 整合额外的BGC副本显著提高了稳定体的产量.
- 发酵的优化导致了4568mg/L的显着最终产量,比本地生产商增加了30倍.
结论:
- 白斑菌 (Streptomyces albus J1074) 是一种非常适合于稳定体蛋白生物合成的宿主.
- 放大稳素BGC是提高产量的有效策略.
- 开发的方法实现了创纪录的产量,使得可以进行可扩展的工业生产,用于合成抗癌药物.
相关概念视频
01:27Upstream Processing
Upstream processing represents a critical phase in biomanufacturing, wherein biological systems such as microorganisms, mammalian cells, or insect cells are cultivated to produce therapeutic proteins, vaccines, enzymes, or other biologically derived products. This phase encompasses all steps from the selection and genetic manipulation of the production organism to the cultivation of cells in bioreactors under tightly controlled environmental conditions.Host Selection and Genetic OptimizationThe...
01:22Bioreactor Controls-III
Strain improvement is a foundational strategy in industrial microbiology aimed at maximizing microbial productivity, particularly because natural isolates typically yield commercially valuable products in very low concentrations. Although optimizing the culture medium and environmental conditions can improve yields, these adjustments are inherently limited by the organism’s genetic potential. As a result, the focus shifts toward genetic modifications to enhance biosynthetic capacity. The...
01:30Production of Pharmaceuticals
Industrial insulin production uses genetically engineered E. coli expressing a proinsulin gene controlled by a tryptophan promoter and containing a methionine linker for later cleavage. The cells also carry ampicillin resistance for selective growth. Seed cultures are stored at −80 °C and production begins by thawing a small amount to inoculate starter cultures, which are progressively scaled to a 50,000-L bioreactor. In the bioreactor, E. coli grow in nutrient-rich media under sterile, tightly...