甲基转化酵母对单氧化酶的分泌表达,用于高效的硫化烯醇生物氧化
Ya-Jing Li1, Yu-Cong Zheng1, Qiang Geng1
1State Key Laboratory of Bioreactor Engineering, East China University of Science and Technology, Shanghai, 200237, People's Republic of China.
Bioresources and bioprocessing
|April 23, 2024
概括
设计的环松单氧化酶 (CHMOAcineto-Mut) 在Pichia pastoris中成功地在细胞外产生,提高了 (S) -omeprazole合成的催化效率. 这促进了这种质子抑制剂前体的工业应用.
科学领域:
- 生物催化剂是一种生物催化剂.
- 蛋白质工程是指蛋白质工程.
- 工业微生物学 工业微生物学
背景情况:
- 普罗基拉 pyrmetazole 是不对称的氧化到 (S) -omeprazole,一个质子抑制剂,使用工程循环素单氧化酶 (CHMOAcineto-Mut).
- 目前大肠杆菌中CHMOAcineto-Mut的生产是细胞内,需要复杂的净化步骤,限制了工业可扩展性.
- 对于治疗胃食道逆流性疾病而言, (S) - 欧美的有效和立体选择性合成至关重要.
研究的目的:
- 为了首次在Pichia pastoris中实现CHMOAcineto-Mut的细胞外生产.
- 提高工程酶的催化效率和工业适用性.
- 开发一种简化生产 (S) -omeprazole的流程.
主要方法:
- 在甲基营养酵母Pichia pastoris中CHMOAcineto-Mut的异质表达.
- 优化发酵条件以最大限度地提高酶分泌.
- 直接使用pH调整的无细胞超浮剂作为pyrmetazole氧化过程中的生物催化剂.
主要成果:
- 在P. pastoris中实现了CHMOAcineto-Mut的细胞外生产,该酶在1,700 U/L时代表了85%的分泌蛋白质.
- 与E. coli产生的酶相比,P. pastoris产生的酶表现出更高的黄素占用率和3.2倍的催化效率.
- 通过直接使用上浮剂,实现了10 g/L基质转化为 (S) - 硫氧化物,其中99%的反体过剩.
结论:
- 皮奇亚牧草是一种适合于高水平的功能性CHMOAcineto-Mut.的细胞外生产的宿主.
- 细胞外生产简化了下游加工,使生物催化剂更容易适用于工业应用.
- 增强的酶和精简的过程为 (S) -omeprazole的高效和立体选择性合成提供了一个有前途的途径.
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