通过代谢和调节工程来定制大肠杆菌BL21 (DE3) 适用于通过代谢和调节工程优先利用草甘
Eliseo R Molina-Vázquez1, Luis Caspeta1, Guillermo Gosset1
1Departamento de Ingeniería Celular y Biocatálisis, Instituto de Biotecnología, Universidad Nacional Autónoma de México, Av. Universidad 2001, Col. Chamilpa, 62210, Cuernavaca, Morelos, Mexico.
Applied microbiology and biotechnology
|February 28, 2025
概括
工程化大肠杆菌通过修改XylR激活器,有效地消耗氧化糖,即使存在葡萄糖. 这种菌株表现出改变的碳催化剂抑制,首选使用西洛斯.
科学领域:
- 微生物学 微生物学
- 代谢工程是代谢工程.
- 合成生物学 合成生物学
背景情况:
- 克西洛斯是一种丰富的粉素,通常与葡萄糖一起发现,在微生物中引起碳催化剂抑制 (CCR).
- 大肠杆菌菌株的新陈代谢和调节变异需要为西洛斯利用量身定制的工程.
- 之前对大肠杆菌BL21 (DE3) 进行过工程改造,将其转化为一种选择性菌株 (ES02) 导致在混合糖条件下减少了生长和糖类消耗.
研究的目的:
- 设计一种选择性西洛斯的大肠杆菌菌株,在混合西洛斯和葡萄糖上生长时克服碳催化剂抑制 (CCR).
- 为了研究一个修改的树脂酶响应转录激活剂 (XylR) 在增强树脂酶利用中的作用.
- 阐明工程菌株中改变葡萄糖同化和CCR背后的机制.
主要方法:
- 通过删除glk,ptsG和manZ基因来对大肠杆菌BL21 (DE3) 进行基因工程.
- 引入一种修改后的酶反应转录激活剂,XylRQ31K.
- 转录组学分析以确定参与葡萄糖同化的基因.
- 为了确认其作用,Galactitol操作子 (gatDCBAZY) 的基因失活.
主要成果:
- 工程菌株ES04证明了高效的西洛斯消耗,单独或与葡萄糖混合.
- 与野生型BL21(DE3) 相比,ES04的特定氧化糖消耗率高出75%.
- ES04显示了改变的CCR,优先摄入西洛斯而不是葡萄糖,同时保持与银基醇操作子 (gatDCBAZY) 过度表达相关的低速度葡萄糖同化.
结论:
- 修改后的XylRQ31K 有效地减轻了选择西洛斯的大肠杆菌中碳催化剂的抑制作用.
- 过度表达 galactitol 操作子有助于在工程菌株中部分使用葡萄糖.
- ES04菌株表现出一种新型表型,即偏好食用糖,而糖代谢发生变化,为混合糖利用提供了一个强大的平台.
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