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定制多个序列对齐作为一种有效的策略,以提高Taq DNA聚合酶的性能

Xinjia Li^1,2, Binbin Chen^1,2, Wanyi Chen^1,2

¹Institute of Bioengineering, College of Chemical and Biological Engineering, Zhejiang University, Hangzhou, 310027, Zhejiang, China.

Applied microbiology and biotechnology

|September 1, 2023

概括

工程Taq DNA聚合酶 (TaqPol) 使用定制的多重序列对齐 (MSA) 显著改善了酶活性和稳定性. 定制的MSA方法在热友酶工程中比广泛的序列池更有效.

科学领域:

生物化学生化学
分子生物学分子生物学
酶工程是什么? 酶工程是什么?

背景情况:

Taq DNA聚合酶 (TaqPol) 工程对于其应用至关重要.
多重序列对齐 (MSA) 是一种用于酶改进的常见方法.

研究的目的:

为增强活动,稳定性和灵敏性设计TaqPol.
评估不同MSA策略对酶工程的有效性.

主要方法:

使用2756个序列的MSA (热友和非热友) 设计的TaqPol突变.
引入了双重突变和针对共同进化残留物的突变.
应用了一个定制的MSA,仅使用来自热友生物的序列.
利用分子动力学模拟来分析突变的影响.

主要成果:

最初的广泛的MSA产生了混合结果的变体;一些活动/稳定性下降.
向突变部分挽救了丢失的属性.
定制的MSA识别了E507Q和E734N变种,同时增加了稳定性,活性和灵敏度.
分子动力学揭示了对蛋白质动力学和基质相互作用的突变效应.

结论:

用于MSA的序列池对酶工程结果产生了重大影响.
定制的MSA对工程热友酶如TaqPol.Pol更有效.
改善DNA结合增强了聚合酶的稳定性和活性.

关键词:

共同发展的共同发展.分子动力学模拟的模拟.多个序列对齐的调整.Taq DNA聚合酶的使用情况

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