功能和结构的洞察力,一个热稳定的 (S) 选择性氨基转氨酶及其通过蛋白质工程改进的基质范围
Stefania Patti1,2, Simone A De Rose3, Michail N Isupov3
1Istituto Di Scienze E Tecnologie Chimiche "G. Natta" (SCITEC), CNR, Milan, Italy.
Applied microbiology and biotechnology
|August 12, 2025
概括
一种高度热稳定的 (S) 选择性氨基转氨酶 (Sbv333-ATA) 在有机溶剂中表现出广泛的基质特异性和稳定性. 局部特异性突变产生了一种W89A突变,对重的氨基酸有增强的活性.
科学领域:
- 生物催化和酶工程 生物催化和酶工程
- 蛋白质结构和功能 蛋白质结构和功能
- 有机化学 有机化学
背景情况:
- 氨基转胺酶是合成奇拉氨基的关键生物催化剂.
- 提高酶稳定性和基质范围对于工业应用至关重要.
- 来自Streptomyces的转氨酶Sbv333-ATA显示出高温稳定性和广泛的基质接受性.
研究的目的:
- 在生物化学和结构上表征Sbv333-ATA.
- 设计Sbv333-ATA以获得扩展的基质特异性,特别是对于重的氨基.
- 通过结构分析阐明酶的活性部位和机制.
主要方法:
- 生物化学测试以确定酶活性和稳定性.
- 结晶学以确定本地酶和突变的3D结构.
- 特定于位点的突变发生,以设计酶特性.
主要成果:
- Sbv333-ATA的化温度为85°C,在各种有机溶剂和双相系统中具有稳定性.
- 该酶接受一系列氨基供体,包括氨基胺和氨基酸,但不包括无菌阻碍的芳氨基胺.
- W89A突变体对像1,2-diphenylethylamine这样的大型二芳氨酸胺的活性显著增加.
- 高分辨率的晶体结构揭示了活性部位的细节,并告知了理性突变发生.
结论:
- Sbv333-ATA是一种强大的生物催化剂,具有各种合成应用的潜力.
- 理性工程,以结构洞察为指导,可以有效地扩大转氨酶的基质特异性.
- 突变W89A代表了一种有前途的工具,用于合成以前无法获得Sbv333-ATA的氨基.
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