通过结构引导理性设计的P450工程实现了在类固醇生物合成中的高C21选择性和生物转化
Jian Yang1, Rong Li1, Qilin Gao1
1College of Life Sciences and Medicine, Zhejiang Sci-Tech University, Hangzhou, Zhejiang, 310018, China.
Synthetic and systems biotechnology
|February 27, 2026
概括
为高效的皮质类固醇合成设计了一种细菌酶,实现了对孕激素C21-基化的区域选择性转移. 这种生物催化剂提供了增强的类固醇生产和对酶工程的洞察力.
科学领域:
- 生物催化和酶工程 生物催化和酶工程
- 合成化学 合成化学
- 分子生物学分子生物学
背景情况:
- C21-基化是合成皮质醇和阿尔多斯特等皮质类固醇的一个关键步骤.
- 来自Nocardia farcinica的酶CYP154C5是这种生物转化的一个潜在候选者.
- 工程P450酶对于提高它们在类固醇合成中的效率和选择性至关重要.
研究的目的:
- 设计一种高效和区域选择性C21-基酶用于孕激素 (PRO) 转化.
- 为了增强工程酶的催化活性和基质范围.
- 用计算方法阐明观察到的区域选择性转移的结构基础.
主要方法:
- 结构引导的理性设计和集中的理性代部位定向突变发生 (FRISM) 用于设计野生型CYP154C5.5.
- 一种氧化还原融合变体 (M6a-RhFRED L3) 被构建以提高催化性能.
- 用分子对接和分子动力学 (MD) 模拟来分析基质结合和活性位点动力学.
主要成果:
- 工程变体M6a在PRO的区域选择性从C16α转变为C21氧化,具有98%的选择性和99%以上的转化.
- 与M6a-RhFRED相比,M6a-RhFRED L3变种的催化活性提高了1.43倍.
- M6a-RhFRED L3成功催化了三种类固醇类别的C21-氧化,具有98%的选择性和>60%的转化.
结论:
- 工程M6a变体及其氧化还原融合衍生物M6a-RhFRED L3是用于类固醇C21-氧化的高效生物催化剂.
- 关键的残留物修饰对于调节P450酶的区域选择性至关重要,使从C16α转向C21基化.
- 这项研究为在类固醇合成中工业应用的P450酶的理性工程提供了宝贵的见解.
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