双重作用的单一工程混合架构促进器增强并将表达方式转换为Komagataella phaffii中的多碳源调节系统
1Department of Chemical Engineering, Biochemical Reaction Engineering Laboratory, Middle East Technical University, 06800 Ankara, Türkiye.
Enzyme and microbial technology
|July 30, 2025
概括
设计的双作用单基因促进剂 (DASEPs) 能够使用多个可持续碳来源 (SCSs) 进行酵母发酵. 这些新型促进剂通过改善对复杂的碳原料的转录控制来显著增强生化生产.
科学领域:
- 合成生物学和代谢工程合成生物学和代谢工程
- 微生物生物技术 微生物生物技术
- 生化生产生化生产
背景情况:
- 像乙醇,甘油,甲醇和酸盐这样的可持续碳来源 (SCS) 对于生物化学生产至关重要.
- 酵母细胞经常同时遇到多个SCS,需要复杂的代谢调节.
- 现有的发起人缺乏灵活性,在发酵过程中有效利用各种SCS.
研究的目的:
- 设计能够响应多个SCS的新型促进剂,以提高发酵效果.
- 开发双重作用的单一工程促进剂 (DASEPs),用于对Komagataella phaffii (Pichia pastoris) 的精确转录控制.
- 通过设计具有多个调节开关的转录机器,将SCS集成到发酵中.
主要方法:
- 基于ADH2促进器架构设计的DASEP,包含合成转录因子结合位 (TFBS).
- 利用预测的主转录因子 (TFs) Cat8,Hap1和Hap2/3/4/5复合体进行促进体设计.
- 通过在各种SCS条件下测量增强的绿色光蛋白 (eGFP) 表达来评估DASEP的性能.
主要成果:
- 与单一作用单一工程促进剂 (SASEPs) 相比,DASEPs在糖醇,乙醇,甲醇和乙酸盐中表现出优异的性能.
- 观察到eGFP表达的显著倍增,例如,糖醇8.2倍,甲醇高达4.2倍.
- 较低的细胞度表明了代谢负担,这表明细胞表达和细胞生长之间存在权衡.
结论:
- DASEPs代表了工程转录机械的重大进步,用于利用多个SCSs.
- 开发的促进剂提供增强和重新连接的转录,以提高生化生产效率.
- 需要进一步优化,以平衡与工程酵母中高水平基因表达相关的代谢负担.
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