高质量和高效的生物合成系统的合理构建和A82846B的发酵优化,基于Amycolatopsis orientalis中的组合策略
Xinyi Zhao1,2, Chenyang Zhu1,2, Wenli Gao1,2
1First Affiliated Hospital and Institute of Pharmaceutical Biotechnology, Zhejiang University School of Medicine, Hangzhou, 310058, China.
Microbial cell factories
|June 29, 2024
概括
这项研究开发了一种多步骤策略,以增强A82846B的微生物生产,这是抗生素oritavancin的前体. 优化的工艺显著提高了A82846B的产量,达到725 mg/L,没有杂质.
科学领域:
- 微生物生物技术 微生物生物技术
- 合成生物学 合成生物学
- 代谢工程是代谢工程.
背景情况:
- 奥里塔万辛是一种半合成的糖抗生素,对于治疗阳性细菌感染至关重要.
- 甲素的直接前体A82846B,产量低,含有同类杂质.
- 目前的微生物发酵方法在生产高质量的A82846B时遇到瓶.
研究的目的:
- 为A82846B.建立一个高质量和高效的生物合成系统.
- 为了优化发酵过程以增加A82846B的产量.
- 为了克服自然产品合成微生物发酵的局限性.
主要方法:
- 基因组测序和分析以删除竞争性途径并构建一个改进的A82846B生产菌株.
- 引入使用CRISPR-Cas12a的PhiC31整合酶系统,用于特定站点的整合.
- 过度表达特定路径调节器StrR和糖合成基因evaE.
- 发酵过程优化和扩大规模的研究.
主要成果:
- 在代谢背景改善后,A82846B的产量从92 mg/L增加到174 mg/L.
- 在引入PhiC31系统和过度表达StrR后,发酵产量达到226 mg/L.
- 过度表达evaE进一步提高了产量,达到332 mg/L.
- 在15升发酵器中扩大规模的生产实现了725毫克/升的创纪录产量,没有同类杂质.
结论:
- 一个多步骤的组合策略成功地为A82846B.创造了一种高产菌株 (AO-6).
- 开发的策略显著提高了A82846B.的微生物发酵.
- 组合方法适用于改善其他微生物二次代谢物的产生.
相关概念视频
01:29Bioreactor Design and Operational System
Bioreactors are engineered vessels designed to cultivate microorganisms under controlled conditions for industrial bioprocessing. They maintain sterility and allow precise regulation of pH, temperature, oxygen, and nutrient levels to optimize microbial growth and metabolite production. Bioreactors range from small laboratory units of 1 liter to industrial systems holding up to 500,000 liters, though only about 75% of their volume is actively used for fermentation. The remaining headspace...
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...
01:27Upstream 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...
01:27Production of Alcohol
Continuous fermentation is a key strategy in industrial ethanol production, particularly when efficiency, scalability, and high yields are essential. This approach allows for uninterrupted operation and optimized resource utilization. The primary feedstock, corn starch, undergoes enzymatic hydrolysis facilitated by α-amylase and glucoamylase. These enzymes break down the starch into fermentable sugars such as glucose, which are readily assimilated by fermentative microorganisms.Fermentation...
01:27Production of Antibiotics
Penicillin, one of the earliest and most widely used antibiotics, is produced industrially by the filamentous fungus Penicillium chrysogenum. Large stirred-tank bioreactors ranging from tens to hundreds of thousands of liters maintain tightly controlled temperature, pH, and dissolved oxygen conditions to support fungal metabolism and maximize antibiotic yield. Penicillin is a secondary metabolite, synthesized primarily during the stationary growth phase, which requires a carefully managed...
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...