Improved thermostability of proteinase K and recognizing the synergistic effect of Rosetta and FoldX approaches.

IF 4.6 Q2 MATERIALS SCIENCE, BIOMATERIALS
Yang Zhao, Daixi Li, Xue Bai, Manjie Luo, Yan Feng, Yilei Zhao, Fuqiang Ma, Guang-Yu Yang
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引用次数: 4

Abstract

Proteinase K (PRK) is a proteolytic enzyme that has been widely used in industrial applications. However, poor stability has severely limited the uses of PRK. In this work, we used two structure-guided rational design methods, Rosetta and FoldX, to modify PRK thermostability. Fifty-two single amino acid conversion mutants were constructed based on software predictions of residues that could affect protein stability. Experimental characterization revealed that 46% (21 mutants) exhibited enhanced thermostability. The top four variants, D260V, T4Y, S216Q, and S219Q, showed improved half-lives at 69°C by 12.4-, 2.6-, 2.3-, and 2.2-fold that of the parent enzyme, respectively. We also found that selecting mutations predicted by both methods could increase the predictive accuracy over that of either method alone, with 73% of the shared predicted mutations resulting in higher thermostability. In addition to providing promising new variants of PRK in industrial applications, our findings also show that combining these programs may synergistically improve their predictive accuracy.

改善蛋白酶K的热稳定性,并认识到Rosetta和FoldX方法的协同效应。
蛋白酶K (PRK)是一种广泛应用于工业的蛋白水解酶。然而,稳定性差严重限制了核dprk的使用。在这项工作中,我们使用了两种结构导向的合理设计方法,Rosetta和FoldX来修饰PRK的热稳定性。基于软件预测可能影响蛋白质稳定性的残基,构建了52个单氨基酸转化突变体。实验表征显示46%(21个突变体)表现出增强的热稳定性。D260V、T4Y、S216Q和S219Q在69°C时的半衰期分别比亲本酶提高了12.4倍、2.6倍、2.3倍和2.2倍。我们还发现,选择两种方法预测的突变比单独使用任何一种方法都能提高预测精度,73%的共享预测突变导致更高的热稳定性。除了在工业应用中提供有希望的PRK新变体外,我们的研究结果还表明,将这些程序结合起来可能会协同提高它们的预测准确性。
本文章由计算机程序翻译,如有差异,请以英文原文为准。
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来源期刊
ACS Applied Bio Materials
ACS Applied Bio Materials Chemistry-Chemistry (all)
CiteScore
9.40
自引率
2.10%
发文量
464
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