稳定突变通过扩大突变景观来提高BlaCβ-乳糖酶的可变性
Marko Radojković1, Anouk Bruggeling van Ingen1, Monika Timmer1
1Leiden Institute of Chemistry, Leiden University, Einsteinweg 55, 2333 CC Leiden, the Netherlands.
Journal of molecular biology
|February 19, 2025
概括
稳定β-乳酸酶酶BlaC的突变,通过提高蛋白质稳定性来恢复抗生素耐药性. 这减轻了进化约束,增强了酶的适应性,扩大了进化可能性.
科学领域:
- 生物化学 生物化学
- 分子生物学分子生物学
- 进化生物学 进化生物学
背景情况:
- 抗微生物耐药性是一个日益增长的威胁,由像β-lactamases这样的酶的进化驱动.
- 获得新的酶功能可能会损害蛋白质的稳定性,这构成了进化挑战.
- 稳定突变可以弥补稳定性损失,从而实现新的酶功能.
研究的目的:
- 识别和描述BlaC中的稳定突变,这是Mycobacterium结核病菌中的β-乳糖酶.
- 研究这些突变如何影响酶的稳定性,功能和可进化性.
- 了解稳定突变在酶进化过程中克服功能稳定性权衡方面的作用.
主要方法:
- 在一个不稳定的BlaC变种上使用随机突变和选择.
- 鉴定出了三个特定突变 (I105F,H184R,V263I).
- 在体外表征评估了埃舍里奇亚大肠杆菌的化学和热稳定性,以及蛋白质水平.
主要成果:
- 鉴定到的突变完全恢复了对抗生素卡本西林的耐药性.
- 这些突变显著增加了BlaC酶的化学和热稳定性.
- 稳定突变导致细菌周等离子体中蛋白质水平更高.
- 稳定突变大大提高了酶的可变性.
结论:
- 稳定突变对于克服酶进化中的功能稳定性权衡至关重要.
- 这些突变可以恢复丢失的功能并增强酶的适应性.
- 这些发现扩大了我们对酶可访问的进化景观的理解.
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