Rational stabilization of enzymes by computational redesign of surface charge-charge interactions
Rational stabilization of enzymes by computational redesign of surface charge-charge interactions
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DOI：
10.1073/pnas.0808220106
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发表时间：
2009-02-24
影响因子：
11.1
通讯作者：
Makhatadze, George I.
中科院分区：
文献类型：
--
作者：
Gribenko, Alexey V.;Patel, Mayank M.;Makhatadze, George I.
在此，我们报道了一种计算方法的应用，该方法能够合理设计具有增强热稳定性同时保留全部酶活性的酶。这种方法基于对蛋白质表面电荷 - 电荷相互作用能量的优化。我们通过实验在两种人源酶——酰基磷酸酶（AcPh）和Cdc42 GTP酶上测试了该方法的有效性，这两种酶在大小（分别为98个氨基酸残基与198个氨基酸残基）和三级结构上有所不同。我们表明，所设计的蛋白质比相应的野生型蛋白质稳定性显著提高。稳定性的提高并没有伴随着所设计的AcPh或Cdc42在结构、寡聚化状态，或者最重要的是在活性方面的显著变化。这种设计方法的成功表明，它可以单独或者与基于蛋白质核心相互作用重新设计的其他策略相结合，普遍应用于其他酶。
Here, we report the application of a computational approach that allows the rational design of enzymes with enhanced thermostability while retaining full enzymatic activity. The approach is based on the optimization of the energy of charge-charge interactions on the protein surface. We experimentally tested the validity of the approach on 2 human enzymes, acylphosphatase (AcPh) and Cdc42 GTPase, that differ in size (98 vs. 198-aa residues, respectively) and tertiary structure. We show that the designed proteins are significantly more stable than the corresponding WT proteins. The increase in stability is not accompanied by significant changes in structure, oligomerization state, or, most importantly, activity of the designed AcPh or Cdc42. This success of the design methodology suggests that it can be universally applied to other enzymes, on its own or in combination with the other strategies based on redesign of the interactions in the protein core.